alpha-modified minimal essential medium α-mem  (FUJIFILM)

Journal: Scientific Reports

Article Title: Role of CXCL10 released from osteocytes in response to TNF-α stimulation on osteoclasts

doi: 10.1038/s41598-025-87092-7

Figure Lengend Snippet: Gene expression in tumor necrosis factor-alpha (TNF-α)-stimulated osteocytes are comprehensively analyzed using RNA-sequencing (RNA-Seq) and TNF-α induces C-X-C motif chemokine ligand 10 (CXCL10) expression in osteocytes. ( a ) The number of Topaz-positive cells isolated using fluorescence-activated cell sorting (FACS). ( b ) Morphology of isolated Topaz-positive cells. ( c ) Heatmap of 422 genes with differentially expressed genes (DEGs) in osteocytes treated with TNF-α. ( d ) Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis of 422 DEGs (control vs. TNF-α). The vertical axis indicates the KEGG pathway, and the horizontal axis indicates the Q-value. ( e ) Top 20 genes with increased expression (red) and bottom 20 genes with decreased expression (blue). ( f ) Expression of Cxcl10 mRNA in osteocytes, analyzed using real-time polymerase chain reaction (qPCR). Total RNA is isolated from osteocytes cultured with or without TNF-α (100 ng/mL) for 12 h. ( g ) CXCL10 in the supernatant of osteocytes cultured with or without TNF-α (100 ng/mL) for 12 h measured by ELISA. ( h ) Heatmap of osteoclast differentiation in TNF-α-treated osteocytes. Data are expressed as the mean ± standard deviation (SD). The statistical significance of differences is determined using the Tukey–Kramer test (n = 4; ** p < 0.01). Scale bar = 100 µm.

Article Snippet: All cell fractions, except fraction 1, were cultured overnight in alpha-modified minimal essential medium (α-MEM) (Fujifilm, Osaka, Japan) containing 10% fetal bovine serum (FBS) and 1% penicillin–streptomycin (P/S) at 37 °C under 5% CO 2 .

Techniques: Gene Expression, RNA Sequencing, Expressing, Isolation, Fluorescence, FACS, Control, Real-time Polymerase Chain Reaction, Cell Culture, Enzyme-linked Immunosorbent Assay, Standard Deviation

Journal: Scientific Reports

Article Title: Role of CXCL10 released from osteocytes in response to TNF-α stimulation on osteoclasts

doi: 10.1038/s41598-025-87092-7

Figure Lengend Snippet: Migration of osteoclast precursors is facilitated by tumor necrosis factor-alpha (TNF-α)-induced C-X-C motif chemokine ligand 10 (CXCL10) expression in osteocytes. ( a ) Schematic diagram of transwell migration assay of the osteoclast precursors under the influence of CXCL10. Osteoclast precursors are seeded in the upper layer, and IgG (100 ng/mL), IgG + CXCL10 (100 ng/mL), CXCL10 + CXCL10 antibody (100 ng/mL), or CXCL10 antibody are added in the lower layer. Osteoclast precursors are cultured for 12 h. ( b ) Microscopic images of osteoclast precursors migrated by CXCL10. ( c ) The number of migrated osteoclasts precursors by CXCL10. ( d ) Schematic diagram of transwell migration assay of the osteoclast precursors within a co-culture of osteoclasts precursors and osteocytes. Osteoclast precursors derived from TNF receptors-deficient (TNFRsKO) mice are seeded in the upper layer, and osteocytes isolated from dentin matrix protein 1 (DMP1)-Topaz mice are seeded in the lower layer. IgG (100 ng/mL), IgG + TNF-α (100 ng/mL), TNF-α + CXCL10 antibody (100 ng/mL), or CXCL10 antibody are added in the lower layer. Osteoclast precursors and osteocytes are incubated in the transwell co-culture system for 12 h. ( e ) Microscopic images of osteoclast precursors migrated by CXCL10 in osteocytes with TNF-α. ( f ) The number of migrated osteoclast precursors within a co-culture of osteoclast precursors and osteocytes. Data are expressed as the mean ± standard deviation (SD). The statistical significance of differences is determined using the Tukey–Kramer test (n = 4; * p < 0.05, ** p < 0.01). Scale bar = 200 µm.

Article Snippet: All cell fractions, except fraction 1, were cultured overnight in alpha-modified minimal essential medium (α-MEM) (Fujifilm, Osaka, Japan) containing 10% fetal bovine serum (FBS) and 1% penicillin–streptomycin (P/S) at 37 °C under 5% CO 2 .

Techniques: Migration, Expressing, Transwell Migration Assay, Cell Culture, Co-Culture Assay, Derivative Assay, Isolation, Incubation, Standard Deviation

Journal: Scientific Reports

Article Title: Role of CXCL10 released from osteocytes in response to TNF-α stimulation on osteoclasts

doi: 10.1038/s41598-025-87092-7

Figure Lengend Snippet: C-X-C motif chemokine ligand 10 (CXCL10) is expressed in osteocytes by tumor necrosis factor-alpha (TNF-α), and anti-CXCL10 antibody inhibits TNF-α-induced osteoclast formation in vivo. ( a ) The microscopic images of immunohistochemical staining of CXCL10 of calvariae are obtained from wild-type (WT) mice after 5 days of daily subcutaneous administration of phosphate-buffered saline (PBS) or TNF-α (3 µg/day). The sections are immunostained using CXCL10-specific antibodies and counterstained with hematoxylin. ( b ) The percentage of CXCL10-positive osteocytes in the calvariae. ( c ) Expression of Cxcl10 mRNA in fractions and osteocyte-enriched calvariae, analyzed using real-time polymerase chain reaction (qPCR). Total RNA is isolated from fractions and osteocyte-enriched calvariae. Pf; PBS-treated fractions group, Tf; TNF-α-treated fractions group, Pc; PBS-treated calvariae group, and Tc; TNF-α-treated calvariae group. ( d ) The microscopic images of tartrate-resistant acid phosphatase (TRAP) staining of calvariae obtained from WT mice after 5 days of daily subcutaneous administration of IgG (3 µg/day), IgG + TNF-α (3 µg/day), TNF-α + CXCL10-neutralizing antibody (3 µg/day), or CXCL10-neutralizing antibody. Cells stained red are counted as TRAP-positive cells. ( e ) The number of TRAP-positive cells are stained through TRAP staining. Data are expressed as mean ± standard deviation (SD). The statistical significance of differences is determined using the Tukey–Kramer test (n = 4; * p < 0.05, ** p < 0.01). Scale bar = 100 µm.

Article Snippet: All cell fractions, except fraction 1, were cultured overnight in alpha-modified minimal essential medium (α-MEM) (Fujifilm, Osaka, Japan) containing 10% fetal bovine serum (FBS) and 1% penicillin–streptomycin (P/S) at 37 °C under 5% CO 2 .

Techniques: In Vivo, Immunohistochemical staining, Staining, Saline, Expressing, Real-time Polymerase Chain Reaction, Isolation, Standard Deviation

Journal: Scientific Reports

Article Title: Role of CXCL10 released from osteocytes in response to TNF-α stimulation on osteoclasts

doi: 10.1038/s41598-025-87092-7

Figure Lengend Snippet: Anti-C-X-C motif chemokine ligand 10 (CXCL10) antibody inhibits tumor necrosis factor-alpha (TNF-α)-induced bone resorption in vivo . ( a ) Three-dimensional (3D) mouse calvariae images using micro-computed tomography (micro-CT). IgG (3 µg/day), IgG (3 µg/day) + TNF-α (3 µg/day), TNF-α (3 µg/day) + CXCL10-neutralizing antibody (3 µg/day), or CXCL10-neutralizing antibody (3 µg/day) are injected subcutaneously into the calvariae of mice for five consecutive days. The red areas indicate bone resorption areas. ( b ) The ratio of bone resorption area. Data are expressed as the mean ± standard deviation (SD). The statistical significance of differences is determined using the Tukey–Kramer test (n = 4; ** p < 0.01).

Article Snippet: All cell fractions, except fraction 1, were cultured overnight in alpha-modified minimal essential medium (α-MEM) (Fujifilm, Osaka, Japan) containing 10% fetal bovine serum (FBS) and 1% penicillin–streptomycin (P/S) at 37 °C under 5% CO 2 .

Techniques: In Vivo, Micro-CT, Injection, Standard Deviation

Journal: Scientific Reports

Article Title: Role of CXCL10 released from osteocytes in response to TNF-α stimulation on osteoclasts

doi: 10.1038/s41598-025-87092-7

Figure Lengend Snippet: Illustration of potential C-X-C motif chemokine ligand 10 (CXCL10) of osteocyte induced by tumor necrosis factor-alpha (TNF-α). TNF-α enhances CXCL10 expression in osteocytes. CXCL10 expression in osteocytes through TNF-α facilitates the migration of osteoclast precursors. Therefore, the number of osteoclasts and bone resorption may increase by TNF-α induced CXCL10 expression in osteocytes.

Article Snippet: All cell fractions, except fraction 1, were cultured overnight in alpha-modified minimal essential medium (α-MEM) (Fujifilm, Osaka, Japan) containing 10% fetal bovine serum (FBS) and 1% penicillin–streptomycin (P/S) at 37 °C under 5% CO 2 .

Techniques: Expressing, Migration

Journal: Scientific Reports

Article Title: Role of CXCL10 released from osteocytes in response to TNF-α stimulation on osteoclasts

doi: 10.1038/s41598-025-87092-7

Figure Lengend Snippet: Top 20 upregulated genes and bottom 20 downregulated expressed genes in tumor necrosis factor-alpha (TNF-α) stimulated osteocytes.

Article Snippet: All cell fractions, except fraction 1, were cultured overnight in alpha-modified minimal essential medium (α-MEM) (Fujifilm, Osaka, Japan) containing 10% fetal bovine serum (FBS) and 1% penicillin–streptomycin (P/S) at 37 °C under 5% CO 2 .

Techniques: Control