Different Modulatory Effects of Four Methicillin‐Resistant <em>Staphylococcus aureus</em> Clones on MG-63 Osteoblast-Like Cells

Abstract

Staphylococcus aureus is a Gram-positive bacterium causing a range of mild to life-threatening infections including bone infections such as osteomyelitis. S. aureus is able to invade and persist within non-professional phagocytic cells such as osteoblasts. In the present study four different S. aureus strains, 2SA-ST239-III, 5SA-ST5-II, 10SA-ST228-I, and 14SA-ST22-IVh were tested for their ability to modulate cell viability in MG-63 osteoblast-like cells following a successful invasion and persistence. Methicillin-sensitive S. aureus (MSSA) ATCC-12598-ST30 was used as control. Despite the demonstrated similar abilities of internalization and persistence of ATCC-12598-ST30, 2SA-ST239-III, and 14SA-ST22-IVh strains in MG-63 osteoblast-like cells under our experimental conditions, we demonstrated that the decrease in cell viability was due to the different behavior of the considered strains, with the number of intracellular bacteria playing a limited role. We focused our attention on different cellular biochemical functions related to inflammation, cell metabolism, and oxidative stress during osteoblast infections. We were able to show that: 1) ATCC-12598-ST30 and 2SA-ST239-III were the only two clones able to persist and maintain their number into the cellular hostile environment during the entire period of infection; 2) 2SA-ST239-III was the only clone able to significantly increase the gene expression (3 and 24 h) and protein secretion (24 h) of both interleukin-6 (IL-6) and tumor necrosis factor alpha (TNF-&amp;alpha;) in MG-63 osteoblast-like cells; 3) the same clone determined a significant up-regulation of transforming growth factor-&amp;beta;1 (TGF-&amp;beta;1) and the metabolic marker glyceraldehyde 3-phosphate dehydrogenase (GAPDH) mRNAs at 24 h post infection; 3) neither the MSSA nor the four MRSA strains induced oxidative stress phenomena in MG-63 cells, although a very different expression pattern towards nuclear factor E2-related factor 2 (Nrf2) and its downstream gene heme oxygenase 1 (HO-1) activation was observed among the different clones. Our results can open a new way of considering therapies, going in the direction of an individualized therapeutic strategy that should take into account the difference existing between MSSA and MRSA as well as the distinctive features of the different clones. Not only, therefore, a different antibiotic approach but also a starting point for considering different host factors, i.e. the modulation of specific cytokines such as IL-6, TNF-&amp;alpha;, and TGF-&amp;beta;1.

Contemporary clinical isolates of Staphylococcus aureus from pediatric osteomyelitis patients display unique characteristics in a mouse model of hematogenous osteomyelitis

“…However, only TI2, and not LAC, displayed lower osteoblast cytotoxicity and intra-osteoclast proliferation. These findings coincide with previous work establishing that S. aureus clinical isolates differ in their virulence, pathogenicity, and toxicity, and that a lower cytotoxicity correlates with increased persistence in vivo (34,35). We found major differences in specific tissue tropism among isolates.…”

“…However, our knowledge of the clinical behavior of these isolates is limited to only one patient per isolate, so it is currently unclear how these findings translate to a broader population. Further work is needed to determine specific bacterial factors that may promote differences in infection incidence and tropism, as well as if the lower cytotoxicity of TI2 contributes to chronicity as seen with other isolates (34, 35). It will be interesting to see if the incidence of sequestra or periosteal reactions – characteristic changes of chronic osteomyelitis - increases over time past the 5 weeks we examined here.…”

Staphylococcus aureus Internalization in Osteoblast Cells: Mechanisms, Interactions and Biochemical Processes. What Did We Learn from Experimental Models?

“…Finally, in previously published works, we demonstrated that internalization, using MG-63 cells, is a pathophysiological pathway of some methicillin-resistant S. aureus (MRSA) which depends on the total number of cells infected and not on the number of bacterial cells that enter each osteoblast. Furthermore, even if our strains were not homogeneous in terms of genetic backgrounds and virulence factors, ST22-IVh and ST239-III S. aureus showed higher intracellular persistence in host cells, making them more prone to developing chronic and recurrent infections, and the different genetic background was also accompanied by a different modulation of inflammatory phenomena, metabolism and antioxidant machinery [ 64 , 65 ].…”

“…In this way, S. aureus induces inflammatory cell recruitment, leading to bone loss [ 3 ]. Cultured osteoblasts infected with bacteria secrete immune modulators of the inflammatory response, cytokines and chemokines, which trigger bone inflammation and destruction [ 65 , 101 , 102 ].…”

Lung Surfactant Decreases Biochemical Alterations and Oxidative Stress Induced by a Sub-Toxic Concentration of Carbon Nanoparticles in Alveolar Epithelial and Microglial Cells

“…After 2 h, cells were treated with ECNs or DPPC:POPG(7:3)/ECNs and incubated for 24 h, in a humidified environment (37 °C, 5% CO 2 ). At this time point, the effects of the two different experimental conditions on cell viability and cells proliferation were measured through the MTT method [ 63 , 64 ] and the Cell Proliferation Kit II (XTT), following the manufacturer’s instructions.…”