*2.4. Fetuin-A Leads to Disorganized Microtubule Nucleation*

*p* = 0.001). Thus, fetuin-A induces centrosome amplification.

*2.4. Fetuin-A Leads to Disorganized Microtubule Nucleation* Microtubule nucleation activity is mainly regulated by the centrosome, and fetuin-A leads to aberrant centrosome homeostasis. Thus, the microtubule nucleation ability was examined by the microtubule regrowth assay. Microtubules were disrupted by 1 h of nocodazole treatment. Then, the cells were washed by PBS and grown in the fresh medium for 10 min to allow the microtubule to regrow. Nocodazole treatment efficiently disrupted microtubule arrays in both the control and fetuin-A-treated cells (Figure 5A). After microtubule regrowth for 10 min, the microtubule concentrated around the centrosome and the array started to emanate from the centrosome to the Microtubule nucleation activity is mainly regulated by the centrosome, and fetuin-A leads to aberrant centrosome homeostasis. Thus, the microtubule nucleation ability was examined by the microtubule regrowth assay. Microtubules were disrupted by 1 h of nocodazole treatment. Then, the cells were washed by PBS and grown in the fresh medium for 10 min to allow the microtubule to regrow. Nocodazole treatment efficiently disrupted microtubule arrays in both the control and fetuin-A-treated cells (Figure 5A). After microtubule regrowth for 10 min, the microtubule concentrated around the centrosome and the array started to emanate from the centrosome to the periphery of the cells (Figure 5B, upper panel), however, in fetuin-A-treated cells, the emanated microtubule density was less than that of in the control cells, and these microtubule arrays did not extend to the cell periphery (Figure 5B, lower panel). Thus, fetuin-A treatment leads to the disorganization of microtubule nucleation.

periphery of the cells (Figure 5B, upper panel), however, in fetuin-A-treated cells, the emanated microtubule density was less than that of in the control cells, and these microtubule arrays did not extend to the cell periphery (Figure 5B, lower panel). Thus, fetuin-A treatment leads to the

**Figure 5.** Fetuin-A treatment leads to disorganized microtubule arrays. Microtubules were depolymerized by nocodazole treatment (**A**), and, then, cells were cultured in the drug-free medium for 10 min (**B**), in the absence (CTL) or presence of fetuin-A (FA). Centrosomes and microtubules (MT) were immunostained with antibodies against pericentrin (centrosome marker) and γ-tubulin **Figure 5.** Fetuin-A treatment leads to disorganized microtubule arrays. Microtubules were depolymerized by nocodazole treatment (**A**), and, then, cells were cultured in the drug-free medium for 10 min (**B**), in the absence (CTL) or presence of fetuin-A (FA). Centrosomes and microtubules (MT) were immunostained with antibodies against pericentrin (centrosome marker) and γ-tubulin (microtubule marker). DNA was stained with DAPI. Scale bar 5 µM.
