## 3 Results
### The Plant Extracts Increase the Selected Intestinal Epithelial Differentiation Markers  
Next, we aimed to check if prolonged exposure to sea buckthorn and Correlian cherry extracts is able to modulate the intestinal epithelial differentiation in two separate intestinal mucosa models using a mixture of Caco-2/HT-29-MTX cells at a 9:1 ratio: (i) a 3D spheroid culture in the non-adherent Biofloat U-shaped plate (Sarstedt) and (ii) a 2D confluent culture on the porous polycarbonate inserts. In both models the prolonged culture (2 weeks in the presence of the plant extracts or solvent as a control) enabled Caco-2's differentiation into absorptive enterocyte-like cells, whereas HT-29-MTX acquire goblet cell features, such as mucin secretion. The spheroids in all the experimental groups formed the brush border structures (Figure 1a, fluorescent \(\beta\)-actin staining panel) facing the environment (i.e., upside-out spheroid morphology). This indicated that the apical surfaces of Caco-2 cells were polarised, which is an important step in the differentiation process. The presence of the plant extracts during the culture did not significantly affect the spheroid polarisation (Figure 1a) or size (Figure 1b). Nevertheless, all the extracts, but particularly sea buckthorn (RO) and Cornelian cherry fruit (DO), enhanced the differentiation markers of goblet cells, as indicated by the darker Alcian blue staining of the mucins (Figure 2a). Similarly, the spheroids that grew in the presence of the extracts demonstrated a stronger activity of intestinal alkaline phosphatase (ALPI), a brush border enzyme regarded as a microvilli marker (Figure 2a). In the two-dimensional cultures, Cornelian cherry leaf extract (DL) significantly promoted differentiation into both enterocyte and goblet cells grown on microscopic slides (Figure 2b-d) \begin{table}
\begin{tabular}{c c c c c} \hline \hline
**Phenolic Compounds** & **Sea Buckthorn Leaves (RL)** & **Sea Buckthorn Fruit (RO)** & **Cornelian Cherry Leaves (DL)** & **Cornelian Cherry Fruit (DO)** \\ \hline Protocatechuic acid & ND & 38.63 \(\pm\) 0.31 \({}^{\text{a}}\) & 436.00 \(\pm\) 8.44 \({}^{\text{b}}\) & ND \\ _Trans_ caffeine acid & 38.64 \(\pm\) 4.38 \({}^{\text{c}}\) & 12.14 \(\pm\) 1.03 \({}^{\text{b}}\) & 64.16 \(\pm\) 0.91 \({}^{\text{d}}\) & 1.65 \(\pm\) 1.43 \({}^{\text{a}}\) \\ _Trans_\(p\)-cumaric acid & 7.74 \(\pm\) 0.06 \({}^{\text{a}}\) & ND & 63.09 \(\pm\) 2.34 \({}^{\text{b}}\) & 7.41 \(\pm\) 0.10 \({}^{\text{a}}\) \\ _Trans_ ferulic acid & ND & 1.09 \(\pm\) 0.24 \({}^{\text{a}}\) & 111.00 \(\pm\) 4.51 \({}^{\text{b}}\) & 2.72 \(\pm\) 0.82 \({}^{\text{a}}\) \\ Rutin trihydrate & 366.00 \(\pm\) 16.53 \({}^{\text{d}}\) & 86.00 \(\pm\) 1.14 \({}^{\text{b}}\) & 155.59 \(\pm\) 1.46 \({}^{\text{c}}\) & 41.41 \(\pm\) 2.51 \({}^{\text{a}}\) \\ Myricetin & 4.66 \(\pm\) 0.24 \({}^{\text{a}}\) & 2994.98 \(\pm\) 26.54 \({}^{\text{b}}\) & 3.57 \(\pm\) 0.06 \({}^{\text{a}}\) & ND \\ Resveratrol & 3.35 \(\pm\) 0.06 \({}^{\text{a}}\) & 27.86 \(\pm\) 2.90 \({}^{\text{b}}\) & 4.70 \(\pm\) 0.41 \({}^{\text{a}}\) & ND \\ Quercetin hydrate & 4.77 \(\pm\) 0.03 \({}^{\text{a}}\) & ND & 5.42 \(\pm\) 0.05 \({}^{\text{b}}\) & ND \\ _Trans_ cinnamic acid & 84.65 \(\pm\) 0.29 \({}^{\text{c}}\) & 0.67 \(\pm\) 0.24 \({}^{\text{a}}\) & 4.12 \(\pm\) 0.03 \({}^{\text{b}}\) & ND \\ Apigenin & 3.67 \(\pm\) 0.03 \({}^{\text{b}}\) & ND & 1.69 \(\pm\) 0.02 \({}^{\text{a}}\) & ND \\ Kaempfer & 4.42 \(\pm\) 0.02 \({}^{\text{a}}\) & 36.04 \(\pm\) 2.43 \({}^{\text{b}}\) & ND & ND \\ \hline \hline \end{tabular}
\end{table} 
Table 1: The content of selected polyphenols in the plant tissue extracts (\(\text{\mu g}/\text{g}\) DW sample; mean \(\pm\) SD, n = 3). ND—not detected. Figure 1: **(a)** Caco-2/HT-29-MTX spheroid growth in the presence of sea buckthorn leaf extract (RL, 50 \(\text{\mu g}/\text{mL}\)) or fruit extract (RO, 50 \(\text{\mu g}/\text{mL}\)) and Cornelian cherry leaf (DL, 50 \(\text{\mu g}/\text{mL}\)) and fruit (DO, 50 \(\text{\mu g}/\text{mL}\)) extracts.