Increased neurotrophin activity in degenerative intervertebral discs (IVDs) is one potential cause of chronic low-back pain (LBP). The aim of the study was to assess if nerve growth factor (NGF) ...might alter gene expression of IVD cells and contribute to disc degeneration by enhancing expression or activity of factors that cause breakdown of IVD matrix.
Rat-tail IVD cells were stimulated by NGF and subjected to microarray analysis. Real-time polymerase chain reaction, Western blotting, and immunocytochemistry of rat and human IVD cells and tissues treated with NGF in vitro in the absence or presence of the NGF inhibitor Ro 08-2750 were used to confirm findings of the microarray studies. Phosphorylation of mitogen-activated protein kinase (MAPK) was used to identify cell signaling pathways involved in NGF stimulation in the absence or presence of Ro 08-2750.
Microarray analysis demonstrated increased expression of chitinase 3-like 1 (Chi3l1), lipocalin 2 (Lcn2), and matrix metalloproteinase-3 (Mmp3) following NGF stimulation of rat IVD cells in vitro. Increased gene expression was confirmed by real-time polymerase chain reaction with a relative increase in the Mmp/Timp ratio. Increased expression of Chi3l1, Lcn2, and Mmp3 following NGF stimulation was also demonstrated in rat cells and human tissue in vitro. Effects of NGF on protein expression were blocked by an NGF inhibitor and appear to function through the extracellular-regulation kinase 1/2 (ERK1/2) MAPK pathway.
Nerve growth factor has potential effects on matrix turnover activity and influences the catabolic/anabolic balance of IVD cells in an adverse way that may potentiate IVD degeneration. Anti-NGF treatment might be beneficial to ameliorate progressive tissue breakdown in IVD degeneration and may lead to pain relief.
Knee osteoarthritis (OA) is a complex disease involving both biomechanical and metabolic factors that alter the tissue homeostasis of articular cartilage and subchondral bone. The catabolic ...activities of extracellular matrix degradation products, especially fibronectin (FN), have been implicated in mediating cartilage degradation. Chondrocytes express several members of the integrin family which can serve as receptors for FN including integrins α5β1, αvβ3, and αvβ5. The purpose of this study was to determine whether polymorphisms in the FN (FN-1) and integrin genes are markers of susceptibility to, or severity of, knee OA in a Han Chinese population.
Two independent case-control studies were conducted on 928 patients with knee OA and 693 healthy controls. Ten single nucleotide polymorphisms (SNPs) of FN-1 and the integrin αV gene (ITGAV) were detected using the ABI 7500 real-time PCR system.
The AT heterozygote in FN-1 (rs940739A/T) was found to be significantly associated with knee OA (adjusted OR = 1.44; 95% CI = 1.16-1.80) in both stages of the study. FN-1 rs6725958C/A and ITGAV rs10174098A/G SNPs were only associated with knee OA when both study groups were combined. Stratifying the participants by Kellgren-Lawrence (KL) score identified significant differences in the FN-1 rs6725958C/A and rs940739 A/T genotypes between patients with grade 4 OA and controls. Haplotype analyses revealed that TGA and TAA were associated with a higher risk of OA, and that TAG conferred a lower risk of knee OA in the combined population.
Our study suggests that the FN-1 rs940739A/T polymorphism may be an important risk factor of genetic susceptibility to knee OA in the Han Chinese population.
Pathology of soft tissue tumours Oniscu, Anca; Salter, Donald
Surgery (Oxford),
April 2023, 2023-04-00, Letnik:
41, Številka:
4
Journal Article
Recenzirano
Soft tissue tumours are a heterogeneous group of neoplasms with differentiation towards mesenchymal tissue. They may arise anywhere in the body and show similar clinical presentation. Traditional ...histopathological diagnosis is now complemented by molecular diagnostic techniques that have become firmly established ancillary diagnostic methods. This article provides a short overview of the aetiology and clinical features of soft tissue sarcomas with an update on how molecular genetics is influencing classification and management of these rare tumours.
In previous work we demonstrated that the matrix-forming phenotype of cultured human cells from whole meniscus was enhanced by hypoxia (5% oxygen). Because the meniscus contains an inner region that ...is devoid of vasculature and an outer vascular region, here we investigate, by gene expression analysis, the separate responses of cells isolated from the inner and outer meniscus to lowered oxygen, and compared it with the response of articular chondrocytes. In aggregate culture of outer meniscus cells, hypoxia (5% oxygen) increased the expression of type II collagen and SOX9 (Sry-related HMG box-9), and decreased the expression of type I collagen. In contrast, with inner meniscus cells, there was no increase in SOX9, but type II collagen and type I collagen increased. The articular chondrocytes exhibited little response to 5% oxygen in aggregate culture, with no significant differences in the expression of these matrix genes and SOX9. In both aggregate cultures of outer and inner meniscus cells, but not in chondrocytes, there was increased expression of collagen prolyl 4-hydroxylase (P4H) alpha (I) in response to 5% oxygen, and this hypoxia-induced expression of P4H alpha (I) was blocked in monolayer cultures of meniscus cells by the hypoxia-inducible factor (HIF)-1 alpha inhibitor (YC-1). In fresh tissue from the outer and inner meniscus, the levels of expression of the HIF-1 alpha gene and downstream target genes (namely, those encoding P4H alpha (I) and HIF prolyl 4-hydroxylase) were significantly higher in the inner meniscus than in the outer meniscus. Thus, this study revealed that inner meniscus cells were less responsive to 5% oxygen tension than were outer meniscus cells, and they were both more sensitive than articular chondrocytes from a similar joint. These results suggest that the vasculature and greater oxygen tension in the outer meniscus may help to suppress cartilage-like matrix formation.
Sarcomas are rare forms of cancer with a high unmet clinical need that develop in connective tissue, such as muscle, bone, nerves, cartilage, and fat. The outcome for patients is poor, with surgery ...and postoperative radiotherapy the standard treatment for patients. A better understanding of the molecular pathology of sarcoma may allow for the development of novel therapeutics. There are dozens of sarcoma subtypes where there is a need for targetted therapeutics, with the most commonly studied including Ewing’s sarcoma and osteosarcoma. Here we initiate a proteomics-based target-discovery program to define “dominant” pro-oncogenic signaling targets in the most common sarcoma in adults: high-grade pleiomorphic soft tissue sarcoma. We have carried out a proteome screen using tandem mass tag isobaric labeling on three high-grade undifferentiated pleomorphic sarcoma biopsies from different tissue sites. We identified the commonly dysregulated proteins within the three sarcomas and further validated the most penetrant receptor as CLIC1, using immunohistochemistry arising from two different population cohorts representing over 300 patients. The dominant expression of CLIC1 in a broad range of human sarcomas suggests that studying this relatively unexplored signaling pathway might provide new insights into disease mechanism and facilitate the development of new CLIC1 targeted therapeutics.
We have performed finite element electromagnetic analysis of 'apertureless' scanning near-field optical microscope tips. A variety of radii was considered, from 40 nm down to 1 nm, and the ...enhancement of optical scattering from the region beneath the probe apex was modelled in air and water. We observed sizeable spectral shifts when the radius decreases, together with a surprising increase in the amount of scattering from small tips. The lateral resolution is considered, which is shown to be always smaller than the tip radius, and for 1-nm-radius tips can allow atomic resolution imaging with sufficient optical enhancement. Gold, silver, copper and aluminium were modelled as tip materials.
In addition to being the primary organ involved in redox cycling, the liver is one of the most highly innervated tissues in mammals. The interaction between hepatocytes and sympathetic, ...parasympathetic, and peptidergic nerve fibers through a variety of neurotransmitters and signaling pathways is recognized as being important in the regulation of hepatocyte function, liver regeneration, and hepatic fibrosis. However, less is known regarding the role of the sympathetic nervous system (SNS) in modulating the hepatic response to oxidative stress. Our aim was to investigate the role of the SNS in healthy and oxidatively stressed liver parenchyma. Mice treated with 6-hydroxydopamine hydrobromide were used to realize chemical sympathectomy. Carbon tetrachloride (CCl4) injection was used to induce oxidative liver injury. Sympathectomized animals were protected from CCl4 induced hepatic lipid peroxidation-mediated cytotoxicity and genotoxicity as assessed by 4-hydroxy-2-nonenal levels, morphological features of cell damage, and DNA oxidative damage. Furthermore, sympathectomy modulated hepatic inflammatory response induced by CCl4-mediated lipid peroxidation. CCl4 induced lipid peroxidation and hepatotoxicity were suppressed by administration of an α-adrenergic antagonist. We conclude that the SNS provides a permissive microenvironment for hepatic oxidative stress indicating the possibility that targeting the hepatic α-adrenergic signaling could be a viable strategy for improving outcomes in patients with acute hepatic injury.
ObjectiveTransforming growth factor/Smad family member 3 (TGF)-β/Smad3 signalling is essential for maintaining articular cartilage. A relationship between the genetic variants of TGF-β itself, TGF-β ...signalling and binding molecules, and osteoarthritis (OA) has been reported. Although variants of candidate genes have become prime targets for genetic analysis, their detailed interplay has not been documented. Our goal was to establish whether single nucleotide polymorphisms (SNPs) of TGF-β1, TGF-βRI, Smad3 and tissue inhibitor of metalloproteinases 3 (TIMP3), and their interactions, are associated with knee OA.DesignWe performed a case–control association study and genotyped 518 knee patients with OA and 468 healthy controls. All participants were genotyped for TGF-β1 (rs1800469C/T), TGF-βRI (rs1590A/G), Smad3 (rs12901499A/G and rs6494629T/C), and TIMP3 (rs715572G/A and rs1962223G/C) polymorphisms by polymerase chain reaction–restriction fragment length polymorphism analysis. Multifactor dimensionality reduction (MDR) was used to identify gene–gene interactions.ResultsSignificant associations were observed for TIMP3 rs715572G/A polymorphisms in knee patients with OA and healthy individuals. The GA heterozygote in TIMP3 (rs715572G/A) was significantly associated with OA (p=0.007). Patient stratification using the Kellgren–Lawrence grading scale showed significant differences in TIMP3 rs715572G/A genotypes between grade 4 knee OA and controls. By MDR analysis, a two-locus model (Smad3 rs6494629T/C and TIMP3 rs715572G/A) of gene–gene interaction was the best for predicting knee OA risk, and its maximum testing accuracy was 57.55% and maximum cross-validation consistency was 10/10.ConclusionsTIMP3 rs715572G/A is a candidate protective gene for severe knee OA. Gene–gene interactions between Smad3 rs6494629T/C and TIMP3 rs715572G/A polymorphisms may play more important protective roles in knee OA.
Recent studies provide evidence of roles for integrins in mechanical signalling in bone and cartilage. Integrin signalling is modulated by various mechanisms, including interaction with other ...transmembrane proteins. We aimed to identify whether one such protein, integrin-associated protein (CD47/IAP), is expressed by chondrocytes and whether it may regulate integrin-dependent mechanotransduction.
Chondrocytes, isolated from macroscopically normal and osteoarthritic articular cartilage of human knee joints, were studied in a resting state or following mechanical stimulation at 0.33 Hz. CD47/IAP expression and associations were confirmed by immunohistology, reverse transcription-polymerase chain reaction, Western blotting, and immunoprecipitation. Roles in mechanotransduction were studied by assessing effects of function-blocking antibodies on a range of electrophysiological, cellular, and molecular responses of primary chondrocytes and responses of CD47/IAP-null cell lines transfected with CD47/IAP.
Human articular chondrocytes were shown to express CD47/IAP, predominantly the type 2 isoform. Immunoprecipitation showed association of CD47/IAP with alpha5 integrin and thrombospondin but not SIRPalpha (signal-regulatory protein-alpha). The function-blocking anti-CD47/IAP antibody Bric 126 inhibited changes in membrane potential, tyrosine phosphorylation, and elevation of relative levels of aggrecan mRNA induced by mechanical stimulation, whereas in the presence of B6H12, an antibody that has partial agonist activity, a membrane depolarisation rather than a membrane hyperpolarisation response was induced by mechanical stimulation. CD47-null cell lines did not show changes in cell membrane potential following mechanical stimulation. Changes in cell membrane potential following mechanical stimulation were seen when CD47-null cells were transfected with CD47/IAP expression vectors but were not seen following mechanical stimulation of cells transfected with vectors for the extracellular immunoglobulin variable (IgV) domain of CD47/IAP in the absence of the transmembrane and intracellular domains.
CD47/IAP is necessary for chondrocyte mechanotransduction. Through interactions with alpha5beta1 integrin and thrombospondin, CD47/IAP may modulate chondrocyte responses to mechanical signals.
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