RESUMEN
Septic arthritis induced by Staphylococcus aureus (S. aureus) causes irreversible cartilage degradation and subsequent permanent joint dysfunction. Recently, cartilage degradation in osteoarthritis is recognized to be associated with metabolic disorders. However, whether cholesterol metabolism is linked to septic arthritis pathology remains largely unknown. Here, we found that exposure to fermentation supernatant (FS) of S. aureus in chondrocytes resulted in a significant increase in expression of key modulators involved in cholesterol metabolism, including lectin-type oxidized low density lipoprotein receptor 1 (LOX1), cholesterol 25-hydroxylase (CH25H), 25- hydroxycholesterol 7α-hydroxylase (CYP7B1) as well as retinoic acid-related orphan receptor alpha (RORα), a binding receptor for cholesterol metabolites. We further demonstrated that enhancement of CH25H/CYP7B1/RORα axis resulted from FS exposure was mediated by activation of NF-κB signaling, along with upregulation in catabolic factors including matrix metallopeptidases (MMP3 and MMP13), aggrecanase-2 (ADAMTS5), and nitric oxide synthase-2 (NOS2) in chondrocytes. Exogenous cholesterol acts synergistically with FS in activating NF-κB pathway and increases cholesterol metabolism. While, the addition of tauroursodeoxycholic acid (TUDCA) which promotes cholesterol efflux, resulted in remarkable reduction of intracellular cholesterol level and restoration of balance between anabolism and catabolism in FS treated chondrocytes. Collectively, our data indicated that, in response to FS of S. aureus, NF-κB signaling activation coupled with increased cholesterol metabolism to stimulate catabolic factors in chondrocytes, highlighting cholesterol metabolism as a potential therapeutic target for treating septic arthritis.
Asunto(s)
Artritis Infecciosa/genética , Cartílago/crecimiento & desarrollo , Osteoartritis/genética , Staphylococcus aureus/patogenicidad , Proteína ADAMTS5/genética , Artritis Infecciosa/microbiología , Artritis Infecciosa/patología , Cartílago/metabolismo , Cartílago/microbiología , Cartílago/patología , Células Cultivadas , Colesterol/genética , Condrocitos/metabolismo , Condrocitos/microbiología , Condrocitos/patología , Familia 7 del Citocromo P450/genética , Regulación de la Expresión Génica/genética , Humanos , Metaloproteinasa 13 de la Matriz/genética , Metabolismo/genética , FN-kappa B/genética , Óxido Nítrico Sintasa de Tipo II/genética , Miembro 1 del Grupo F de la Subfamilia 1 de Receptores Nucleares/genética , Osteoartritis/microbiología , Osteoartritis/patología , Receptores Depuradores de Clase E/genética , Transducción de Señal/genética , Esteroide Hidroxilasas/genética , Ácido Tauroquenodesoxicólico/genética , Factor de Transcripción ReIA/genéticaRESUMEN
7α-Hydroxysteroid dehydrogenase (7α-HSDH) plays an important role in the efficient biotransformation of taurochenodeoxycholic acid (TCDCA) to tauroursodeoxycholic acid (TUDCA). In this paper, a novel NADP(H)-dependent 7α-HSDH (named J-1-1) was discovered, heterologously expressed in Escherichia coli and biochemically characterized. J-1-1 exhibited high enzymatic activities. The specific activities of J-1-1 toward TCDCA, glycochenodeoxycholic acid (GCDCA) and ethyl benzoylacetate (EBA) were 188.3 ± 0.2, 217.6 ± 0.4, and 20.0 ± 0.2 U·mg-1, respectively, in 50 mM Glycine-NaOH, pH 10.5. Simultaneously, J-1-1 showed high thermostability; 73% of its activity maintained after heat treatment at 40 °C for 100 h. Particularly noteworthy is that magnesium ion could stabilize the structure of J-1-1, resulting in the enhancement of its enzymatic activity and thermostability. The enzymatic activity of J-1-1 increased 40-fold in the presence of 50 mM Mg2+, and T0.5 increased by approximately 6 °C. Furthermore, after heat treatment at 40 °C for 20 min, the control group only retained 52% of the residual enzyme activity, while the residual enzyme activity of the experimental group was still 77% of the J-1-1 enzyme activity with Mg2+ and without heat treatment. These properties of 7α-HSDH would be expected to contribute to more extensive applications in the biotransformation of related substrates.
