## 4 Discussion
HPLC analysis revealed that loligide levels remained unchanged, while pheophorbide A and a compound (\(m/z\) 277.2192, likely linolelaidic or linolenic acid) increased under heterotrophic conditions. These results suggest that the enhanced effect is not loligide-dependent but attributed to these components. Furthermore, transitioning to heterotrophic cultivation offers significant industrial advantages by reducing production costs, simplifying facility construction, and enabling large-scale biomass production without the limitations of light penetration. This approach enhances scalability, ensures higher biomass yields compared to autotrophic cultivation, and improves commercial feasibility for applications such as biofuel production and bioactive compound extraction [49; 89; 90; 91]. Furthermore, exploring the exact mechanisms through which these alternative compounds contribute to hair growth warrants further investigation. As per prior studies, increased DP sphere sizes are related to hair inductivity [72; 92]. The number of DP cells is associated with the anagen phase and influences the size and morphology of hair, forming a niche that affects progenitor cells [93]. On the other hand, the miniaturization of hair follicles is known as a typical phenotype in alopecia [42; 43]. In this study, we aimed to investigate whether _S. deserticola_ JD052 extracts have hair-inductive properties in 2D-cultured and 3D spheroids of HDP cells (Figures 1 and 2). To enhance the effect of _S. deserticola_ JD052 extracts on hair inductivity, we upgraded it via modifying the cultivation conditions of _S. deserticola_ JD052. In our results, it was demonstrated that _S. deserticola_ JD052 extracts increased the cell viability and the sphere size of HDP cells in a concentration-dependent manner. Interestingly, HE activated the transcriptional activity of \(\beta\)-catenin higher than PE did at the same dose of each extract as expected (Figures 2c and S2). The function of dermal papilla on forming spherical structures is crucial due to affecting hair induction directly. With this insight, our data indicated that HE can perhaps act as a trigger of DP spheroid growth and that increased DP spheroid size by HE is possibly related to hair inductivity. These results highlight that HE has effective hair-inductive properties, engaging spheroid formation and stimulating the expression of DP signature genes in HDP cells. Since WNT/\(\beta\)-catenin signaling plays an important role in hair growth initiation, hair follicle morphogenesis, and hair growth, diverse studies investigated this signaling pathway as a therapeutic strategy for alopecia improvement [28; 94; 95]. As shown prior, we also observed that _S. deserticola_ JD052 extracts increased the protein expression level of total \(\beta\)-catenin, whilst the phosphorylation level of \(\beta\)-catenin, which means \(\beta\)-catenin degradation, was decreased (Figures 1e and 3a,b). After this, we also found that the translocation status of \(\beta\)-catenin into the nucleus was upregulated by HE treatment (Figure 3c). These data are consistent with the increase in TCF/LEF-induced luciferase activity described earlier. Consequently, it was suggested that _S. deserticola_ JD052 extracts enhanced the stabilization of \(\beta\)-catenin followed by translocation. GSK3\(\beta\), which is phosphorylated at the serine 9 residue by AKT and ERK and at the threonine 390 residue by p38 MAPK, is known as a crucial element of the \(\beta\)-catenin signaling pathway [55; 56; 57]. In addition, AKT, also known as protein kinase B (PKB), is a serine/threonine-specific kinase in the proliferation mechanism of various kinds of cell types including DP cells. Therefore, the regulation of AKT signaling has been studied as a therapeutic strategy for alopecia, the same approach as \(\beta\)-catenin in DP cells [59; 96]. Based on this, we demonstrated that HE upregulated the phosphorylation level of AKT and subsequently phosphorylated GSK3\(\beta\) at serine 9, not threonine 390 (Figure 4a).