*2.2. D Cell Culture Cytotoxicity of Glutathione Responsive* β*-cyclodextrin-Based Nanosponges*

The basal level of reduced glutathione was measured in each of the different cell lines, which were grouped according to cancer type. The results show that HT-29 (Figure 1A) and PC-3 cells (Figure 1B) display significantly lower GSH levels than those in HCT116 (Figure 1A) and DU145 cells (Figure 1B). Dose-response curves were then performed by exposing human colorectal cancer, HCT116 and HT-29, and human prostatic carcinoma, DU145 and PC-3, cell monolayers to various concentrations (0.5, 1.0, 2.0, and 3.0 mg/mL) of the two types of GSH-NSs with increasing disulfide bridge content (GSH-NS B and D), for 24, 48, and 72 h. Table 2 reports the IC1 and IC50 values of GSH-NS, which were determined at 24, 48, and 72 h of exposure.

Figure 2 reports GSH-NS IC50 values according to disulfide-bond content and exposure time. These data highlight the remarkable cytotoxicity difference between the two nanosponge formulations in colorectal cancer cell lines, as lower IC50 values were observed for the nanosponge formulation with the lower disulfide bridge content (GSH-NS B) at 24, 48, and 72 h (Figure 2A,B). In prostatic cancer cell lines, no significant differences in IC50 values were detected for the two nanosponge formulations in DU145 cells (Figure 2C), which were characterized by the highest GSH cell content (Figure 1). Meanwhile, in PC-3 cells, a significant cytotoxicity difference was observed between the two nanosponge formulations, as the IC50 values determined by the GSH-NS B formulation were lower compared with what was observed in colorectal cancer cells, even if only at 24 and 48 h (Figure 2D).

in prostatic cancer cells than in colorectal cancer cells.

GSH-NS D, in all of the cell lines, with the exception of DU145 cell line (Table 2 and Figure 2). Moreover, lower GSH-NS D IC50 values were observed on DU145 and PC-3 cells than on HCT116 and HT-29 cells at 48 and 72 h of exposure (Table 2 and Figure 2), suggesting that the nanosponge

**Figure 1.** Intracellular glutathione-responsive β-cyclodextrin (GSH) level according to cell type. The reduced glutathione content of human colorectal cancer cell lines, HCT 116 and HT-29 (**A**), and human prostatic carcinoma cell lines, DU145 and PC-3 (**B**), was measured at a basal level, that is, in untreated cells, and was expressed as nmol/µg protein. The results are mean values ± SD of three independent experiments performed in triplicate. Statistically significant difference between cell lines: \* *p* < 0.05. **Figure 1.** Intracellular glutathione-responsive β-cyclodextrin (GSH) level according to cell type. The reduced glutathione content of human colorectal cancer cell lines, HCT 116 and HT-29 (**A**), and human prostatic carcinoma cell lines, DU145 and PC-3 (**B**), was measured at a basal level, that is, in untreated cells, and was expressed as nmol/µg protein. The results are mean values ± SD of three independent experiments performed in triplicate. Statistically significant difference between cell lines: \* *p* < 0.05.

**Table 2.** Glutathione-responsive β-cyclodextrin-based nanosponge (GSH-NS) inhibition concentration (IC) values on 2D cell cultures. **Table 2.** Glutathione-responsive β-cyclodextrin-based nanosponge (GSH-NS) inhibition concentration (IC) values on 2D cell cultures.

Statistically significant difference between the two nanosponge formulations: \* *p* < 0.05. Statistically significant difference between the two nanosponge formulations: \* *p* < 0.05.

Therefore, the nanosponge formulation with the lower disulfide-bond content, GSH-NS B, was found to be more cytotoxic than the nanosponge formulation with the higher disulfide-bond content, GSH-NS D, in all of the cell lines, with the exception of DU145 cell line (Table 2 and Figure 2). Moreover, lower GSH-NS D IC50 values were observed on DU145 and PC-3 cells than on HCT116 and HT-29 cells at 48 and 72 h of exposure (Table 2 and Figure 2), suggesting that the nanosponge formulation with the higher disulfide bridge concentration, GSH-NS D, has a higher cytotoxic effect in prostatic cancer cells than in colorectal cancer cells.

**Figure 2.** Cytotoxicity of glutathione responsive β-cyclodextrin-based nanosponges in 2D cell cultures. HCT116 (**A**), HT-29 (**B**), DU145 (**C**), and PC-3 (**D**) cells were incubated with GSH-NS B and GSH-NS D at different concentrations (0.5, 1.0, 2.0, 3.0 mg/mL) for 24, 48, and 72 h. Cell proliferation was evaluated by WST-1 assay and the values of the concentration required for a 50% cell growth inhibition (IC50) were determined by the proliferation curves obtained using the CalcuSyn 2.11 software (Biosoft, Cambridge, UK). The results are mean values ± St.Dev of three independent experiments, replicated eight times for each condition. Statistically significant difference between **Figure 2.** Cytotoxicity of glutathione responsive β-cyclodextrin-based nanosponges in 2D cell cultures. HCT116 (**A**), HT-29 (**B**), DU145 (**C**), and PC-3 (**D**) cells were incubated with GSH-NS B and GSH-NS D at different concentrations (0.5, 1.0, 2.0, 3.0 mg/mL) for 24, 48, and 72 h. Cell proliferation was evaluated by WST-1 assay and the values of the concentration required for a 50% cell growth inhibition (IC50) were determined by the proliferation curves obtained using the CalcuSyn 2.11 software (Biosoft, Cambridge, UK). The results are mean values ± St.Dev of three independent experiments, replicated eight times for each condition. Statistically significant difference between nanosponge formulations: \* *p* < 0.05.


"With Great Power Comes Great Impressionability": A Study of the Relation between Stereotypes and Superheroes

*Samuel C. Van Vleet, Everrett Moore, Alvin Akibar, Azlynn Osborne and Yolanda Flores Niemann*


*3.7. Li's MoN*

For a scalar random variable *x*, the un-normalized MoN proposed by Li [43,44] represents the square root of the minimum mean square distance between the nonlinear measurement function *h* and the set of all affine functions *L*,

$$J = \min \left( E\{ (L(\mathbf{x}) - h(\mathbf{x}))^2 \} \right)^{1/2},\tag{59}$$

where *L*(*x*) = *Ax* + *B*. The scalar parameters *A* and *B* are determined in the minimization process. For the current problem, where *x* is non-random, the un-normalized MoN *J* and normalized MoN *ν* ar given, respectively, by

$$J = \sigma\_h \sqrt{1 - \frac{c\_{hx}^2}{\sigma\_h^2 \sigma\_x^2}}.\tag{60}$$

$$\nu = \mathbf{J}/\sigma\_{\mathbf{h}} = \sqrt{1 - \frac{c\_{\mathbf{h}\mathbf{x}}^2}{\sigma\_{\mathbf{h}}^2 \sigma\_{\mathbf{x}}^2}}.\tag{61}$$

Given *x*ˆ and *σx* (30), the unscented transformation (UT) [14,15], cubature transformation (CT) [16], or Monte Carlo method [8] can be used to compute *σ*2 *h* and *chx*. We find that the UT gives good results in calculating the two MoNs. Next we dscribe computing *J* and *ν* using the UT. We use *κ*UT = 2 [14]. The three weights and sigma points are given, respectively, by

$$w\_0 = 2/3, \quad w\_1 = 1/6, \quad w\_2 = 1/6,\tag{62}$$

$$
\chi\_0 = \mathfrak{x}, \quad \chi\_1 = \mathfrak{x} + \sqrt{3}\sigma\_{\mathfrak{x}\prime} \quad \chi\_2 = \mathfrak{x} - \sqrt{3}\sigma\_{\mathfrak{x}\prime} \tag{63}
$$

The measurement transformed points are

$$h\_i = a \,\chi\_i^n, \quad i = 0, 1, 2. \tag{64}$$

Then the mean and variance of *h* a given by

$$\bar{h} = \sum\_{i=0}^{2} w\_i h\_{i\prime} \tag{65}$$

$$
\sigma\_h^2 = \sum\_{i=0}^2 w\_i (h\_i - \bar{h})^2,\tag{66}
$$

The cross-covariance *chx* is computed by

$$\boldsymbol{\omega}\_{\text{hx}} = \sum\_{i=0}^{2} w\_i (\boldsymbol{h}\_i - \boldsymbol{\bar{h}}) (\boldsymbol{\chi}\_i - \boldsymbol{\mathfrak{k}}).\tag{67}$$