CELLULAR AND MOLECULAR ACTIVITY OF A STANDARDIZED SMALL SEA FISH EXTRACT IN AN EXPERIMENTAL MODEL OF PRIMARY HUMAN CARTILAGE CELLS

Abstract

The inflammatory, degenerative and rheumatic diseases are triggered and evolving at osteoarticular level due to the disruption of functionality of constitutive cells and extracellular microenvironment correlated with systemic responses as synthesis of progressively degradative cytokines. Objectives. The in vitro bioactive efficacy of standardized small sea fish extract has therapeutic relevance on the strength of optimal physiological and structural biologic system rendered by primary cells isolated from human cartilage (HCH, PromoCell) unlike standardized cell lines that may have genetic and functional changes and beside the primary cells directly isolated from animal tissue that do not reproduce an authentic biological response because of metabolic stress adaptation to growing conditions in vitro. The main therapeutic targets in osteoarticular disorders are regeneration of affected cartilage and attenuation of local inflammatory processes. Materials and methods. The in vitro articular matrix reconstruction action was evaluated by molecular tests with relative quantification of gene expression (qPCR) for aggrecan, the predominant component of human cartilage, and for the enzyme responsible for degradation of aggrecan, aggrecanase (ADAMTS4) whose activities were phenotypically measured by ELISA (enzyme-linked immunosorbent assay) test. The anti-inflammatory profile was proved by techniques of quantitative evaluation of mRNA level and detection of extracellular soluble proteins by flow-cytometry (BD™ Cytometric Bead Array) for pro-inflammatory cytokines IL-6, IL-8 and gene expression for nuclear factor NF-κB. Results and discussion. The results prove the matrix regenerator effect of standardized small fish extract by stimulation at gene expression level of aggrecan synthesis in early stages of chondrocytes differentiation, directing cells to a mature functional status, as well as by inhibition of aggrecan degradation by action on gene expression and extracellular activity of ADAMTS4. The standardized small sea fish extract has an anti-inflammatory effect proven by extracellular and transcriptional inhibition of pro-inflammatory cytokines, IL-6, IL-8 and the nuclear factor NF-κB gene. Conclusions. The in vitro anti-inflammatory and regenerative action of standardized small fish extract reflects a significant therapeutic potential for osteoarticular disease sustained both through the effects at gene and phenotypic expression levels and through the reproducible characteristics of dynamics of in vivo chondrocytes evolution evidenced by primary cell lines.

Keywords

• osteoarthritis
• inflammation
• extracellular matrix
• small marine sea fish concentrate
• aggrecan
• aggrecanase
• interleukin