Rosavin modulates CX3CL1 secretion and osteoblast morphology under mineralizing conditions in vitro

Rosavin, a bioactive compound derived from Rhodiola rosea, has been proposed to modulate osteoblast-associated signaling pathways. Emerging evidence highlights the role of CX3CL1 (fractalkine) in bone-immune interactions. This study aimed to investigate whether rosavin modulates CX3CL1 secretion and osteoblast morphology and confluence in human osteoblasts (HOB) in vitro, thereby providing descriptive cellular evidence relevant to CX3CL1-associated osteoblast responses under mineralizing conditions.

Prior to biological testing, the chemical identity of rosavin was verified by high-resolution electrospray ionization quadrupole time-of-flight mass spectrometry (ESI-QTOF-MS). HOB were cultured in growth medium (DMEM-g), mineralization medium (DMEM-m), or DMEM-m supplemented with rosavin at 50 µM (R50) or 100 µM (R100). Cell morphology and confluence were assessed by phase-contrast microscopy, supported by semi-quantitative image-based estimation of cell-covered area using Fiji/ImageJ. Sirius Red staining was performed only in the DMEM-m mineralization control group and was evaluated qualitatively as a supportive assessment of extracellular matrix maturation.

Rosavin was associated with dose- and time-dependent changes in CX3CL1 secretion and morphology-related observations in HOB cultures. The 50 µM rosavin condition showed significantly increased CX3CL1 secretion at days 14 and 21 compared with DMEM-m alone and was associated with higher cell confluence and predominance of cuboidal cell morphology. In contrast, the 100 µM condition was associated with lower apparent confluence, mixed cellular morphology, and reduced CX3CL1 secretion at later time points.

These observations are descriptive, preliminary, and hypothesis-generating. Rosavin was associated with concentration-dependent modulation of CX3CL1 secretion and osteoblast morphological characteristics in vitro; however, the biological significance of CX3CL1 modulation in this monoculture model remains uncertain. Further mechanistic and functional studies are required.