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Comparing effects of perfusion and hydrostatic pressure on human chondrocytes using gene profiles

ABSTRACT: Hydrostatic pressure and perfusion have been shown to alter the chondrogenic potential of articular chondrocytes. In order to compare the effects of hydrostatic pressure plus perfusion (HPP) and perfusion (P) we investigated the complete gene expression profiles of human chondrocytes under HPP and P. A simplified bioreactor was constructed applying loading (0.1 MPa for 2 h) and perfusion (2ml) through the same piping by pressurizing the medium directly. High-density monolayer cultures of human chondrocytes were exposed to HPP or P for 4 days. Controls were maintained in static culture. Gene expression was evaluated by sequencing (RNAseq) and quantitative real-time PCR analysis. RNAseq identified similarities between the two treatments. Specifically, HPP and P increased COL2A1 expression and decreased COL1A1 and MMP-13 expression. Despite of the similarities, RNAseq revealed a list of cartilage genes including ACAN, ITGA10 and TNC, which were differentially expressed by HPP and P. Of these candidates adhesion related molecules were found to be upregulated in HPP. Both HPP and P treatment had beneficial effects on chondrocyte differentiation and decreased catabolic enzyme expression. The study provides new insight into how hydrostatic pressure and perfusion enhance cartilage differentiation and inhibit catabolic effects

ORGANISM(S): Homo sapiens

PROVIDER: GSE69206 | GEO | 2016/03/21

SECONDARY ACCESSION(S): PRJNA284885

REPOSITORIES: GEO

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Project description:IL-1B is an important cytokine that is often found to be up-regulated during osteoarthritic and rheumatoid joint diseases. It is viewed as a catabolic factor, inducing enzymes that allow for the degradation of the cartilage extracellular matrix and also has essential roles as an autocrine and paracrine factor in fibronectin fragment-mediated degradation. It can also reduce the synthesis of the major cartilage components, type II collagen and aggrecan. On the other hand, IL-1B also has the ability to induce the growth and morphogenic factor BMP-2. During joint diseases, IL-1B is synthesized by both synovial cells and chondrocytes. Addition of IL-1 biological antagonists such as IL-1 receptor antagonists can suppress cartilage degradation in vitro. Thus, the production of IL-1B could act as the first step in mediating a cascade of other mediators in cartilage which could be relevant to the fate of the cartilage. In order to obtain a global picture of the effect of IL-1B production on human adult articular chondrocytes, we analyzed changes in gene expression induced by IL-1B by microarray analysis. We found that IL-1B has a diverse effect on gene expression profile in chondrocytes. One of the predominant responses that we observed in adult human articular chondrocytes on exposure to IL-1B is a dramatic increase in a large set of chemokines and other genes related to the inflammatory cascade. Keywords: Gene response to IL-1B (10 ng/ml) Cartilage was obtained from adult human tissue donors with above the knee amputations due to chondrosarcoma or traumatic injury or from autopsy. Chondrocytes were isolated following established protocols, maintained in high density, and treated with IL-1B (10 ng/ml). Chondrocytes treated with buffer only served as the untreated control. The experiment was carried out in duplicate. Total RNA was extracted from these chondrocytes, labeled with fluorophores (Cy3 or Cy5) and analyzed for expression changes using the Human Operon/Qiagen v3.0 oligonucleotide array. The analysis was repeated with the fluorophore dyes exchanged between the untreated and experimental RNAs.

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Comparing effects of perfusion and hydrostatic pressure on human chondrocytes using gene profiles

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