Figure 1.
Morphological appearance and endogenous HA localization of basic fibroblast growth factor (bFGF) and macrophage-colony-stimulating factor (MCSF) treated FLSC cultures. Phase contrast images (a,d) Confocal localization of HA (green fluorescence) and CD44 (red fluorescence) (b,c,e,f). Panels (c,f) show images of cells which were digested with S. Hyase prior to fixation, to remove extracellular HA, revealing abundant intracellular HA (marked with white * in panels c,f). Panel g shows the % of cells containing intracellular HA from a total of n = 50–70 cells, in images taken from 5 separate areas of coverslips prepared from 3 individual cultures. Cell fixation, and staining protocols for HA (with bHABP) and CD44 (with anti-CD44) were carried out as described in the Methods Section. Space bar = 10 μm.
Figure 1.
Morphological appearance and endogenous HA localization of basic fibroblast growth factor (bFGF) and macrophage-colony-stimulating factor (MCSF) treated FLSC cultures. Phase contrast images (a,d) Confocal localization of HA (green fluorescence) and CD44 (red fluorescence) (b,c,e,f). Panels (c,f) show images of cells which were digested with S. Hyase prior to fixation, to remove extracellular HA, revealing abundant intracellular HA (marked with white * in panels c,f). Panel g shows the % of cells containing intracellular HA from a total of n = 50–70 cells, in images taken from 5 separate areas of coverslips prepared from 3 individual cultures. Cell fixation, and staining protocols for HA (with bHABP) and CD44 (with anti-CD44) were carried out as described in the Methods Section. Space bar = 10 μm.
Figure 2.
Localization of endogenous intracellular hyaluronic acid (HA) to endoplasmic reticulum (ER) or Lysosomal compartments in bFGF and MCSF treated FLSC cultures prior to treatment with LPS or exogenous high molecular weight hyaluronic acid (HMW HA). Cells were dual labelled with bHABP and either anti-Calnexin (a,b,e,f) or anti-LAMP1 (c,d,g,h) antibodies without (a,c,e,g) or with S Hyaluronidase pretreatment (b,d,f,h). Intracellular HA was partially co-localized with anti-LAMP1 (lysosomes, white arrow heads in d,h). There was no evidence for localization of intracellular HA to the ER compartments. Panels (i,j) show higher magnifications of the boxed areas in panels (d,h), respectively to illustrate the co-localization of HA with LAMP1-+ve intracellular vesicles. The bar-graph shows the quantitation of cells positive for intracellular HA, which was determined as described in the Methods Section. Space bars = 10 μm.
Figure 2.
Localization of endogenous intracellular hyaluronic acid (HA) to endoplasmic reticulum (ER) or Lysosomal compartments in bFGF and MCSF treated FLSC cultures prior to treatment with LPS or exogenous high molecular weight hyaluronic acid (HMW HA). Cells were dual labelled with bHABP and either anti-Calnexin (a,b,e,f) or anti-LAMP1 (c,d,g,h) antibodies without (a,c,e,g) or with S Hyaluronidase pretreatment (b,d,f,h). Intracellular HA was partially co-localized with anti-LAMP1 (lysosomes, white arrow heads in d,h). There was no evidence for localization of intracellular HA to the ER compartments. Panels (i,j) show higher magnifications of the boxed areas in panels (d,h), respectively to illustrate the co-localization of HA with LAMP1-+ve intracellular vesicles. The bar-graph shows the quantitation of cells positive for intracellular HA, which was determined as described in the Methods Section. Space bars = 10 μm.
Figure 3.
LPS Stimulation has no effect on intracellular localization and distribution of endogenous HA. Cells treated with bFGF (a,c,e) or MCSF (b,d,f), then stimulated with LPS for 4 h, followed by 16 h incubation in basal serum containing medium, as described in the Methods. Fixed cells were then dual labelled with bHABP and either anti-CD44 (a,b), anti-Calnexin (c,d) or anti-LAMP1 (e,f). Panels (i,j) show higher magnifications of the boxed areas in panels (d,h), respectively, to illustrate the co-localization of HA with LAMP1-+ve intracellular vesicles. Space bars = 10 μm.
Figure 3.
LPS Stimulation has no effect on intracellular localization and distribution of endogenous HA. Cells treated with bFGF (a,c,e) or MCSF (b,d,f), then stimulated with LPS for 4 h, followed by 16 h incubation in basal serum containing medium, as described in the Methods. Fixed cells were then dual labelled with bHABP and either anti-CD44 (a,b), anti-Calnexin (c,d) or anti-LAMP1 (e,f). Panels (i,j) show higher magnifications of the boxed areas in panels (d,h), respectively, to illustrate the co-localization of HA with LAMP1-+ve intracellular vesicles. Space bars = 10 μm.
Figure 4.
Effect of the addition of exogenous HMW HA on association of endogenous HA with CD44, Calnexin and LAMP1. Cells treated with bFGF (a–d) or MCSF (e–h) and incubated for 16 h in basal FBS containing medium with 100 μg/mL HMW HA. Fixed cells were then dual labelled with bHABP and either anti-CD44 (a,e), anti-Calnexin (c,g), or anti-LAMP1 (d,h). Additional cells were pretreated with S Hyaluronidase prior to fixation, to remove extracellular prior to staining with bHAPB and anti-CD44 (b,f). The bar-graph in panel (i) shows the quantitation of cells positive for intracellular HA, which was determined as described in the Methods Section. Space bars = 10 μm. White arrowheads indicate the deposition of extracellular HA in between cell groups.
Figure 4.
Effect of the addition of exogenous HMW HA on association of endogenous HA with CD44, Calnexin and LAMP1. Cells treated with bFGF (a–d) or MCSF (e–h) and incubated for 16 h in basal FBS containing medium with 100 μg/mL HMW HA. Fixed cells were then dual labelled with bHABP and either anti-CD44 (a,e), anti-Calnexin (c,g), or anti-LAMP1 (d,h). Additional cells were pretreated with S Hyaluronidase prior to fixation, to remove extracellular prior to staining with bHAPB and anti-CD44 (b,f). The bar-graph in panel (i) shows the quantitation of cells positive for intracellular HA, which was determined as described in the Methods Section. Space bars = 10 μm. White arrowheads indicate the deposition of extracellular HA in between cell groups.
Figure 5.
Effect of addition of exogenous HMW HA on association of endogenous HA with CD44, Calnexin and LAMP1 in LPS stimulated cultures. Cells treated with bFGF (a–d) or MCSF (e–h) were stimulated with LPS for 4 h, followed by 16 h incubation in basal FBS containing medium with 100 μg/mL HMW HA. The fixed cells were then dual labelled with bHABP and either anti-CD44 (a,e), anti-Calnexin (c,g) or anti-LAMP1 (d,h). Additional cells were pretreated with S Hyaluronidase prior to fixation, to remove extracellular prior to staining with bHAPB and anti-CD44 (b,f). The bar-graph in panel (i) shows the quantitation of cells positive for intracellular HA, which was determined as described in the Methods Section. Space bars = 10 μm. White arrowheads indicate the deposition of extracellular HA in between cell groups.
Figure 5.
Effect of addition of exogenous HMW HA on association of endogenous HA with CD44, Calnexin and LAMP1 in LPS stimulated cultures. Cells treated with bFGF (a–d) or MCSF (e–h) were stimulated with LPS for 4 h, followed by 16 h incubation in basal FBS containing medium with 100 μg/mL HMW HA. The fixed cells were then dual labelled with bHABP and either anti-CD44 (a,e), anti-Calnexin (c,g) or anti-LAMP1 (d,h). Additional cells were pretreated with S Hyaluronidase prior to fixation, to remove extracellular prior to staining with bHAPB and anti-CD44 (b,f). The bar-graph in panel (i) shows the quantitation of cells positive for intracellular HA, which was determined as described in the Methods Section. Space bars = 10 μm. White arrowheads indicate the deposition of extracellular HA in between cell groups.
Figure 6.
Heatmap of gene expression in the TLR signaling pathway in FLSC cultures maintained in bFGF or MCSF and their modulation by LPS, exogenous HMW HA, or a combination thereof. Cell layers from three independent culture preparations were assayed as described in the Methods. Fold changes relative to bFGF or MSCF only treated cultures were calculated from the respective ΔΔCt values as 2−ΔCt and evaluated for statistical significance as described in the Methods. B2m was used as housekeeping gene. ↑ or ↓ = 2–5-fold increase or decrease; ↑↑ or ↓↓ = 5–50-fold increase or decrease; ↑↑↑ or ↓↓↓ > 50 fold increase.
Figure 6.
Heatmap of gene expression in the TLR signaling pathway in FLSC cultures maintained in bFGF or MCSF and their modulation by LPS, exogenous HMW HA, or a combination thereof. Cell layers from three independent culture preparations were assayed as described in the Methods. Fold changes relative to bFGF or MSCF only treated cultures were calculated from the respective ΔΔCt values as 2−ΔCt and evaluated for statistical significance as described in the Methods. B2m was used as housekeeping gene. ↑ or ↓ = 2–5-fold increase or decrease; ↑↑ or ↓↓ = 5–50-fold increase or decrease; ↑↑↑ or ↓↓↓ > 50 fold increase.
Figure 7.
Heatmap of gene expression in the phagocytosis pathway in FLSC cultures maintained in bFGF or MCSF and modulation of their expression by LPS, Exogenous HA or a combination thereof. Cell layers from three independent cultures (as for
Figure 6) were assayed as described in the Methods. Fold changes relative to bFGF or MSCF only treated cultures were calculated from the respective ΔΔCt values as 2
−ΔΔCt and evaluated for statistical significance as described in
Section 2.
B2m was used as housekeeping gene. ↑ or ↓ = 2–10-fold increase or decrease; ↑↑ or ↓↓ = 11–50-fold increase or decrease; ↑↑↑ or↓↓↓ > 50 fold increase. * Marks genes that were changed by exogenous HMW HA from LPS modulated levels to baseline levels (bFGF or MCSF).
Figure 7.
Heatmap of gene expression in the phagocytosis pathway in FLSC cultures maintained in bFGF or MCSF and modulation of their expression by LPS, Exogenous HA or a combination thereof. Cell layers from three independent cultures (as for
Figure 6) were assayed as described in the Methods. Fold changes relative to bFGF or MSCF only treated cultures were calculated from the respective ΔΔCt values as 2
−ΔΔCt and evaluated for statistical significance as described in
Section 2.
B2m was used as housekeeping gene. ↑ or ↓ = 2–10-fold increase or decrease; ↑↑ or ↓↓ = 11–50-fold increase or decrease; ↑↑↑ or↓↓↓ > 50 fold increase. * Marks genes that were changed by exogenous HMW HA from LPS modulated levels to baseline levels (bFGF or MCSF).
Figure 8.
Western analysis of VCAN, ACAN, and PRG4 from FLSC cultures maintained in bFGF or MCSF, stimulated or not with LPS and exposed or not to exogenous HMW HA. Conditioned media (panels a–c) and cell extracts (panels d–f) were collected and prepared for SDS PAGE/Western blotting as described in the Methods. Membranes were probed for VCAN with anti-DPE (a,d), and for with a mix of anti-CDAG and anti-DLS (1 μg/mL each) for ACAN (b,e). The later were reprobed for PRG4 (panels c,f) with MAb 9G3 as described in the Methods. To confirm equivalent loading between samples, all membranes from media were finally with anti-HC1 and cell extract samples with anti-β-actin (bottom panels). Identified VCAN species 1and 2 represent the high molecular weight core protein and the ADAMTS-generated G1 fragment, respectively. Identified ACAN species include the full length and C-terminally processed core protein, respectively, and the ADAMTS-generated G1 fragment (5). Non-specific reactive bands are indicated by a (*).
Figure 8.
Western analysis of VCAN, ACAN, and PRG4 from FLSC cultures maintained in bFGF or MCSF, stimulated or not with LPS and exposed or not to exogenous HMW HA. Conditioned media (panels a–c) and cell extracts (panels d–f) were collected and prepared for SDS PAGE/Western blotting as described in the Methods. Membranes were probed for VCAN with anti-DPE (a,d), and for with a mix of anti-CDAG and anti-DLS (1 μg/mL each) for ACAN (b,e). The later were reprobed for PRG4 (panels c,f) with MAb 9G3 as described in the Methods. To confirm equivalent loading between samples, all membranes from media were finally with anti-HC1 and cell extract samples with anti-β-actin (bottom panels). Identified VCAN species 1and 2 represent the high molecular weight core protein and the ADAMTS-generated G1 fragment, respectively. Identified ACAN species include the full length and C-terminally processed core protein, respectively, and the ADAMTS-generated G1 fragment (5). Non-specific reactive bands are indicated by a (*).
Figure 9.
Densitometry Quantitation of Immunoreactive VCAN and ACAN species present in medium and cell layer compartments of three separately prepared FLSC cultures treated as described for
Figure 8. Each cell preparation is indicated by differently shaded bars. Data were collected using Image J software and are expressed as Integrated Pixel Density of immunoreactive bands relative to Integrated Pixel density of HC1 reactive bands for medium samples or of b-actin reactive bands for cell extract samples.
Figure 9.
Densitometry Quantitation of Immunoreactive VCAN and ACAN species present in medium and cell layer compartments of three separately prepared FLSC cultures treated as described for
Figure 8. Each cell preparation is indicated by differently shaded bars. Data were collected using Image J software and are expressed as Integrated Pixel Density of immunoreactive bands relative to Integrated Pixel density of HC1 reactive bands for medium samples or of b-actin reactive bands for cell extract samples.
Figure 10.
Localization of VCAN, bFGF, and MCSF treated FLSC cultures before and after addition of exogenous HMW HA. Cells were dual labelled with anti-DPE (red fluorescence, panels a,b,e,f) and bHABP (green fluorescence panels c,d,g,h). Intracellular HA co-localized with VCAN is marked in panels c,d, and g with white arrow heads (yellow fluorescence). ~50% of cells in a given imaged area was positive for VCAN, but the degree of staining varied between cells, as illustrated in panels a,b,e,f. Higher magnification images of co-localized areas in panels (c,g) are shown in panels (i,j), respectively.
Figure 10.
Localization of VCAN, bFGF, and MCSF treated FLSC cultures before and after addition of exogenous HMW HA. Cells were dual labelled with anti-DPE (red fluorescence, panels a,b,e,f) and bHABP (green fluorescence panels c,d,g,h). Intracellular HA co-localized with VCAN is marked in panels c,d, and g with white arrow heads (yellow fluorescence). ~50% of cells in a given imaged area was positive for VCAN, but the degree of staining varied between cells, as illustrated in panels a,b,e,f. Higher magnification images of co-localized areas in panels (c,g) are shown in panels (i,j), respectively.
Figure 11.
Co-localization of Early Endosomal Marker (EEA1) and HA in bFGF and MCSF treated FLSC cultures before and after addition of exogenous HMW HA. Cells were dual labelled with bHABP (green fluorescence) and anti-EEA1 (red fluorescence). Intracellular HA localized with early endosomes is marked in panels c and g (yellow fluorescence), ~50% of cells in a given imaged area was positive for VCAN, but the degree of staining varied between cells, as illustrated in panels a,b,e,f. Higher magnification images of colocalized areas in panels (c,g) are shown in panels (i,j), respectively. Some EEA1 reactivity appeared to be localized away from the cell body, after exposure to exogenous HMW HA (marked with a white arrowhead in panels d,h).
Figure 11.
Co-localization of Early Endosomal Marker (EEA1) and HA in bFGF and MCSF treated FLSC cultures before and after addition of exogenous HMW HA. Cells were dual labelled with bHABP (green fluorescence) and anti-EEA1 (red fluorescence). Intracellular HA localized with early endosomes is marked in panels c and g (yellow fluorescence), ~50% of cells in a given imaged area was positive for VCAN, but the degree of staining varied between cells, as illustrated in panels a,b,e,f. Higher magnification images of colocalized areas in panels (c,g) are shown in panels (i,j), respectively. Some EEA1 reactivity appeared to be localized away from the cell body, after exposure to exogenous HMW HA (marked with a white arrowhead in panels d,h).
Figure 12.
Localization of ACAN and HA in bFGF and MCSF treated FLSC cultures before and after treatment with exogenous HMW HA. The cells were dual labelled with anti-DLS (red fluorescence, panels a–h).and bHABP (green fluorescence panels c,d,g,h). ~20% of cells in a given imaged area stained positive for ACAN and no co-localization between HA and ACAN was observed.
Figure 12.
Localization of ACAN and HA in bFGF and MCSF treated FLSC cultures before and after treatment with exogenous HMW HA. The cells were dual labelled with anti-DLS (red fluorescence, panels a–h).and bHABP (green fluorescence panels c,d,g,h). ~20% of cells in a given imaged area stained positive for ACAN and no co-localization between HA and ACAN was observed.
Table 1.
Baseline Expression levels of TLR4 responsive genes, Il6 and Nos2 and genes involved in HA synthesis, extracellular organization and degradation at 4 and 16 h post-medium change.
Table 1.
Baseline Expression levels of TLR4 responsive genes, Il6 and Nos2 and genes involved in HA synthesis, extracellular organization and degradation at 4 and 16 h post-medium change.
Gene | 4 h + bFGF | 4 h + MCSF | 36 h + bFGF | 36 h + MCSF |
---|
| ⚜Ct * | ⚜Ct * | ⚜Ct * | ⚜Ct * |
---|
Il6 | 10.88 (± 2.17) | 11.28 (± 2.54) | 12.78 (±1.55) | 13.15 (± 2.00) |
Nos2 | ND | ND | ND | ND |
Has 1 | 7.64 (± 1.01) | 8.11 (± 1.92) | 8.83 (±0.15) | 9.22 (± 0.93) |
Has2 | 8.33 (± 1.48) | 8.09 (± 2.31) | 8.52 (±0.92) | 7.65 (± 1.32) |
Ptx3 | 8.19 (± 2.56) | 6.07 (± 3.21) | 8.35 (±0.68) | 8.92 (± 1.29) |
Tnfaip6 | 9.96 (± 2.01) | 9.28 (± 1.89) | 12.68 (±1.49) | 13.13 (± 2.19) |
Cd44 | 1.14 (± 0.31) | 1.67 (± 0.52) | 1.26 (±0.41) | 1.41 (± 0.31) |
Hyla1 | 9.05 (± 2.11) | 8.52 (± 2.42) | 7.33 (±0.46) | 7.12 (± 0.68) |
Hyal2 | 5.39 (± 1.11) | 4.94 (± 0.76) | 4.34 (±0.28) | 3.97 (± 0.63) |
Cemip | 8.81 (± 2.25) | 6.82 (± 1.13) | 7.53 (±0.52) | 7.82 (± 0.67) |
Tmem2 | 6.26 (± 0.99) | 6.05 (± 1.26) | 9.84 (±0.69) | 8.17 (± 0.88) |
Table 2.
Fold changes in Expression of TLR4 responsive genes Il6, Nos2, and genes involved in HA synthesis, extracellular organization and degradation following a 4 h LPS stimulus or a 4 h LPS Stimulus followed by a 16 h incubation in complete medium.
Table 2.
Fold changes in Expression of TLR4 responsive genes Il6, Nos2, and genes involved in HA synthesis, extracellular organization and degradation following a 4 h LPS stimulus or a 4 h LPS Stimulus followed by a 16 h incubation in complete medium.
| 4 h bFGF + LPS | 4 h MCSF + LPS | +16 h bFGF | +16 h MCSF |
---|
Gene | * FOLD | FOLD | FOLD | FOLD |
---|
Il6 | 101.5 (± 0.92) | 93.4 (± 3.44) | 155.1 (± 22.13) | 76.64 (± 11.11) |
Nos2 | ** >>100 | >>100 | >>100 | >>100 |
Has1 | 1.98 (± 0.21) | 2.90 (± 0.87) | 4.85 (± 0.58) | 8.90 (± 0.13) |
Has2 | 1.62 (± 0.03) | 1.65 (± 0.11) | 0.69 (± 0.11) | 0.36 (± 0.08) |
Ptx3 | 12.40 (± 2.33) | 5.32 (± 0.82) | 66.67 (± 4.88) | 132.5 (± 25.3) |
Tnfaip6 | 6.09 (± 1.20) | 12.62 (± 2.11) | 4.97 (± 0.81) | 7.89 (± 0.39) |
Cd44 | 1.04 (± 0.32) | 1.49 (± 0.46) | 0.95 (± 0.28) | 1.23 (± 0.41) |
Hyla1 | 0.86 (± 0.14) | 0.59 (± 0.32) | 0.45 (± 0.12) | 0.28 (± 0.03) |
Hyal2 | 0.85 (± 0.29) | 0.43 (± 0.08) | 0.28 (± 0.11) | 0.11 (± 0.06) |
Cemip | 1.57 (± 0.22) | 0.88 (± 0.71) | 1.30 (± 0.21) | 0.78 (± 0.39) |
Tmem2 | 1.50 (± 0.41) | 1.14 (± 0.15) | 0.24 (± 0.08) | 0.30 (± 0.06) |
Table 3.
Fold changes in the expression of TLR4 responsive genes, Il6, Nos2 genes involved in HA synthesis, extracellular organization and degradation following a 4 h incubation in basal or LPS supplemented medium, followed by a 16 h incubation in complete medium containing 100 μg/mL HMW HA.
Table 3.
Fold changes in the expression of TLR4 responsive genes, Il6, Nos2 genes involved in HA synthesis, extracellular organization and degradation following a 4 h incubation in basal or LPS supplemented medium, followed by a 16 h incubation in complete medium containing 100 μg/mL HMW HA.
| 4 h Basal Medium | 4 h LPS Treatment |
---|
| 16 h bFGF + HA | 16 h MCSF + HA | 16 h bFGF + HA | 16 h MCSF + HA |
---|
Gene | * FOLD | FOLD | FOLD | FOLD |
---|
Il6 | # 2.58 (± 1.07) | # 2.29 (± 0.84) | ### 21.4 (± 4.5) | ### 44.3 (± 5.99) |
Nos2 | ND | ND | ### 0.03 (± 0.06) | ### 0.14 (± 0.03) |
Has1 | 0.86 (± 0.41) | 1.38 (± 0.51) | 1.06 (± 0.21) | 1.04 (± 0.23) |
Has2 | 1.08 (± 0.33) | 1.44 (± 0.59) | # 1.97 (± 0.31) | # 1.87 (± 0.29) |
Ptx3 | 0.87 (± 0.64) | 1.01 (± 0.14) | 1.08 (± 0.09) | 1.13 (± 0.25) |
Tnfaip6 | 0.58 (± 0.44) | 1.10 (± 0.61) | 1.33 (± 0.26) | 1.25(± 0.44) |
Cd44 | 1.23 (± 0.29) | 1.14 (± 0.17) | 0.97 (± 0.31) | 1.19 (± 0.29) |
Hyal1 | 1.12 (± 0.44) | 1.13 (± 0.22) | # 2.01 (± 0.14) | ## 2.68(± 0.27) |
Hyal2 | 0.60 (± 0.71) | 1.27 (± 0.41) | # 1.95 (± 0.54) | ## 3.29 (± 0.98) |
Cemip | 1.09 (± 0.32) | 0.97 (± 0.31) | 0.96 (± 0.33) | 1.15 (± 0.66) |
Tmem2 | 0.68 (± 0.58) | 0.74 (± 0.29) | # 1.78 (± 0.11) | ## 2.32 (± 0.28) |
Table 4.
Baseline expression of Aggrecan (Acan), versican (Vcan), and Prg4 at 4 and 16 h post-medium change.
Table 4.
Baseline expression of Aggrecan (Acan), versican (Vcan), and Prg4 at 4 and 16 h post-medium change.
Gene | 4 h + bFGF | 4 h + MCSF | 36 h + bFGF | 36 h + MCSF |
---|
| ⚜Ct * | ⚜Ct * | ⚜Ct * | ⚜Ct * |
---|
Acan | 9.12 (± 1.1) | 6.93 (± 0.82) | 8.97 (± 1.6) | 6.42 (± 0.61) |
Vcan | 6.91 (± 0.48) | 4.76 (± 1.01) | 6.29 (± 0.57) | 4.74 (± 2.01) |
Prg4 | 3.55 (± 0.51) | 3.92 (± 0.21) | 3.67 (± 0.86) | 3.19 (± 0.20) |
Table 5.
Fold changes in Expression of Acan, Vcan, and Prg4 after 4 h of LPS stimulation followed by 16 h incubation in complete medium.
Table 5.
Fold changes in Expression of Acan, Vcan, and Prg4 after 4 h of LPS stimulation followed by 16 h incubation in complete medium.
Gene | 4 h bFGF + LPS | 4 h MCSF + LPS | + 16 h bFGF | + 16 h MCSF |
---|
| * FOLD | FOLD | FOLD | FOLD |
---|
Acan | ## 0.24 (± 0.11) | ## 0.29 (± 0.03) | ## 0.31 (± 0.08) | ## 0.09 (± 0.03) |
Vcan | 0.84 (± 0.37) | 1.44 (± 0.38) | # 2.44 (± 0.41) | 1.03 (± 0.38) |
Prg4 | 0.74 (± 0.18) | 0.78 (± 0.34) | # 2.07 (± 4.88) | 0.94 (± 25.3) |
Table 6.
Fold changes in Expression of Acan, Vcan, and Prg4 following a 4 h incubation in basal or LPS supplemented medium, followed by a 16 h incubation in complete medium containing 100 μg/mL HMW HA.
Table 6.
Fold changes in Expression of Acan, Vcan, and Prg4 following a 4 h incubation in basal or LPS supplemented medium, followed by a 16 h incubation in complete medium containing 100 μg/mL HMW HA.
Gene | bFGF + HA | MCSF + HA | bFGF + LPS + HA | MCSF + LPS + HA |
---|
| * FOLD | FOLD | FOLD | FOLD |
---|
Acan | 1.64 (± 0.22) | 0.62 (± 0.62) | 1.56 (± 0.51) | 2.26 (± 0.61) |
Vcan | 1.76 (± 0.21) | 2.59 (± 0.11) | 0.78 (± 0.32) | 0.40 (± 0.63) |
Prg4 | 1.44 (± 0.25) | 1.56 (± 0.31) | 1.04 (± 0.08) | 1.69 (± 0.21) |