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Effects of Drugs
Published in Stephen W. Carmichael, Susan L. Stoddard, The Adrenal Medulla 1986 - 1988, 2017
Stephen W. Carmichael, Susan L. Stoddard
Jacobs and Stevens (1986b) examined the effect of the microtubule-depolymerizing drug Nocodazole on the form of neurites extending from PC12 cells. Microtubule polymerization led to the formation of varicosities due to a clustering of membranous organelles in young neurites. Neurites older than 7 days appeared to be resistant to microtubule depolymerization. The authors were able to predict the volume and shape of a neurite quantitatively.
The use of Spheroids in the Study of Invasion
Published in Rolf Bjerkvig, Spheroid Culture in Cancer Research, 2017
Marc Bracke, Hans Romijn, Vakaet Luc, Barbara Vyncke, Marc de Mets, Marc Mareel
Microtubule inhibitors interfere with the assembly of microtubules of both the cytoplasmic microtubule complex (CMTC) and the mitotic spindle.13 Some molecules bind to tubulin (vinca alkaloids, nocodazole, cis-tubulozole, podophyllotoxin) and lead to a net disassembly of the microtubules. Other molecules decrease the critical concentration for microtubule assembly and lead to an overassembly with a chaotic CMTC pattern (taxol). Still others (estramustin) have their target in the microtubule-associated proteins. All treatments have at least two functional consequences for the cell: loss of direction finding during cell translocation due to dysfunction of the CMTC, and inhibition of mitosis because a normal spindle cannot be assembled. With MO4, cells it was shown that the inhibition of invasion is not the result of the effect on the spindle. MO4 spheroids arrested in S-phase by 5-fluorouracil were able to invade into PHF, although they did not divide. Addition of microtubule inhibitors to these nondividing cell populations prevents invasion. These and other experiments have led to the conclusion that a functionally intact CMTC is crucial for invasion and that microtubule inhibitors block invasion via their effect on the CMTC.
Pharmacologic Ascorbate Influences Multiple Cellular Pathways Preferentially in Cancer Cells
Published in Qi Chen, Margreet C.M. Vissers, Cancer and Vitamin C, 2020
Qi Chen, Kishore Polireddy, Ping Chen, Ramesh Balusu, Tao Wang, Ruochen Dong
Acetylation of Lys-40 on an α-tubulin subunit stabilizes lateral interactions between neighboring tubulin protofilaments, whereas deacetylation enhances tubulin depolymerization [41]. Therefore, the acetylation status of α-tubulin confers functional diversity of tubulin polymers by altering its dynamics. In pancreatic cancer cells (PANC-1 and BxPC-3), ascorbate at both subcytotoxic and cytotoxic concentrations (1.25–2.5 mM) increased tubulin acetylation in a dose-dependent manner. Stable microtubules were generated and failed to undergo depolymerization, even in the presence of a tubulin depolymerizing agent, nocodazole [8]. Interestingly, the increase of α-tubulin acetylation in a normal pancreatic ductal epithelial cell was minimal. H2O2 treatment mimicked the effects of ascorbate, and catalase completely prevents the effects of ascorbate. These data indicate that ascorbate stabilizes α-tubulin, and the action is dependent on H2O2 [8] (Figure 3.3). The same study also reported that ascorbate treatment increased expression of epithelial markers (E-cadherin) in pancreatic cancer cells and decreased mesenchymal markers (N-Cad and vimentin), which ultimately increases cell-cell contact and reduces cancer cell metastasis [8]. Polireddy's study also found decreased metalloproteinases (MMPs) and increased collagen in pancreatic cancers treated with ascorbate. Whether the changes in the epithelial-mesenchymal transition (EMT) and cell-cell contact are mechanistically independent or interrelated to tubulin dynamics is not known. A similar phenomenon was observed in a study conducted in endothelial cells [42]. It was found that when ascorbate stabilized microtubules and increased acetylation of α-tubulin, it also increased cell-cell contact and tightened the endothelial permeability barrier [42]. Nevertheless, as a result, the migration and invasion ability of these pancreatic cancer cells decreased, as well as the proliferation [8].
An overview of the recent progress in Middle East Respiratory Syndrome Coronavirus (MERS-CoV) drug discovery
Published in Expert Opinion on Drug Discovery, 2023
A cell-based enzyme-linked immunosorbent assay (ELISA) has been used to screen and evaluate many FDA-approved drugs for their ability to target MERS-CoV. The drugs include neurotransmitter blockers, DNA synthesis/repair inhibitors, lipid and sterol metabolism antagonists, estrogen receptor antagonists, protein-processing blockers, ion transport blockers, cytoskeletal drugs (particularly tubulin), kinase-signaling blockers, and apoptosis inhibitors [22]. Despite not having a clear connection to CoVs, two groups of pharmaceuticals – antiparasitics and antibiotics – showed effectiveness against MERS-CoV infection. Specifically, nocodazole acted on the cytoskeleton to halt spindle development. It is an antimitotic drug originally developed to fight cancer, but studies have shown that it is also highly effective against MERS-CoV [22,130]. Two of nine ion channel blockers tested, monensin and salinomycin sodium, had inhibitory effects against MERS-CoV. There is experimental evidence that MERS-CoV is vulnerable to ionophore activities [131]. The EC50 and CC50 values for monensin were 3.2 and greater than 20 µM, respectively [131]. Thus, it was effective against MERS-CoV at low concentrations and had little toxicity.
Intercellular transfer of mitochondria via tunneling nanotubes protects against cobalt nanoparticle-induced neurotoxicity and mitochondrial damage
Published in Nanotoxicology, 2021
Fuli Zheng, Zhousong Luo, Xinpei Lin, Wei Wang, Michael Aschner, Ping Cai, Yuan-Liang Wang, Wenya Shao, Guangxia Yu, Zhenkun Guo, Siying Wu, Huangyuan Li
To corroborate the effect of astrocyte-derived mitochondria upon CoNPs induced neuronal damage, we analyzed apoptosis of the two cell types by means of flow cytometry. First, we identified U251 and SHSY5Y cells: P6 indicated U251-Mito-EGFP (right cluster) and P3 denoted SHSY5Y cells (left cluster) (Figure 10(a), left panel). Next, the two cell types were analyzed with Annexin V-PE/7-AAD apoptosis detection kit (Figure 10(a), middle: SHSY5Y and right panel: U251-Mito-EGFP). Upon CoNPs exposure, SHSY5Y showed a significant increase in late apoptosis (Figure 10(b)). As shown in Figure 8(c), TNTs were actin rich structures but lacking tubulin. Therefore, the actin and microtubule inhibitors (LAT-B and Nocodazole) were applied to the co-culture system. Both inhibitors failed to elicit a significant effect on apoptosis in both cell types (Figure 10(b,c)). Notably, when exposed to CoNPs, SHSY5Y treated with the actin depolymerizing LAT-B were more prone to both early and late apoptosis (Figure 10(b)). On the other hand, Nocodazole, a microtubule depolymerizing drug, showed no effect on apoptosis-induced by CoNPs in SHSY5Y cells (Figure 10(b)). However, no statistical significance was observed in the U251 cells groups (Figure 10(c)), suggesting that the co-culturing of apoptotic SHSY5Y cells did not affect the viability of U251 cells. In summary, these data demonstrate that blockage of mitochondrial transfer through TNTs inhibition increases CoNPs-induced apoptosis, especially late apoptosis, in SHSY5Y cells.
Delivery of pDNA to lung epithelial cells using PLGA nanoparticles formulated with a cell-penetrating peptide: understanding the intracellular fate
Published in Drug Development and Industrial Pharmacy, 2020
Larissa Gomes dos Reis, Wing-Hin Lee, Maree Svolos, Lyn M. Moir, Rima Jaber, Andrea Engel, Norbert Windhab, Paul M. Young, Daniela Traini
Chlorpromazine, an inhibitor that disrupts the formation of clathrin-coated pits, hence inhibiting clathrin-mediated endocytosis, led to the most significant decrease in NP–DNA–CPP uptake in both cell lines, with only 22% of positive cells in A549 and 67% in Beas-2B, respectively. To a lesser extent, the disruption of microtubules formation via the microtubule depolymerizing agent nocodazole also significantly decreases internalization to ∼63% in both cell lines. The inhibition of GTPase activity dynamin was also able to significantly decrease NP–DNA–CPP uptake in both cell lines. Dynamin is essential for membrane fission of clathrin-coated vesicles [18] and also plays a role on caveola-mediated and lipid-raft endocytosis [16]. Wortmannin, a pharmacological inhibitor that affects the formation of the double-shell of the endosomes, hence inhibiting clathrin-mediated endocytosis, led to a significant decrease by 20 and 25% of NP–DNA–CPP uptake in A549 and Beas-2B respectively. When colchicine was used to bind tubulin, thereby inhibiting cytoskeleton function, a significant decrease in A549 internalization was observed. Similarly, incubation with cytochalasin D decreased internalization to 61% of NP–DNA–CPP in A549. While the depletion of cholesterol in the cellular membrane caused by MβCD and the inhibition of scavenger receptors by Polyisinosinic acid did not affect internalization in A549, a decrease by 16 and 14%, respectively, was observed in Beas-2B.