## 3 Results and Discussion
### Determination of Collagen-Related Proteins in the Dorsal Skin of Nude Mice
UVB: ultraviolet B; NC: normal control; PC: positive control; LPLD, LPLD and LPLD: administration of LPTWK10 viable cells at 2 \(\times\) 10\({}^{8}\) CFU/kg bw/day, 2 \(\times\) 10\({}^{9}\) CFU/kg bw/day and 2 \(\times\) 10\({}^{10}\) CFU/kg bw/day, respectively; SPLD, SPMD and SPHD: administration of LPTWK10 spray-dried supernatant powders at 0.30 g/kg bw/day, 0.60 g/kg bw/day, and 1.20 g/kg bw/day, respectively. is a major collagenase that breaks down collagen and also degrades other ECM components such as elastin, laminin, and fibronectin [43]; hence, it is considered an indicator of photoaging. Increased MMP-1 levels accelerate skin aging, leading to wrinkle formation and compromised skin integrity [41]. MMP-9 is a gelatinase that degrades type IV collagen, a major component of the basement membrane of the skin, during photoaging [44]. Prolonged exposure of the skin to UV radiation upregulates ROS production, leading to the peroxidation of cell membranes, which in turn damages keratinocytes and fibroblasts. For UV-irradiated collagen, only MMP-1 can cleave type I and type III procollagen and denature collagen, which is further catabolized by MMP-3 and MMP-9 [44; 45]. The following findings related to MMP levels were observed at week 13 of UVB and LPTWK10 treatments. Compared with the NC group, the UVB-treated group showed 67.86% higher MMP-1 levels, which significantly decreased by 33.85% on treatment with a high dose of LPTWK10 (\(p<0.05\)) (Figure 5D). MMP-9 levels were significantly higher in the UVB group than those in the NC group by 33.20%, which significantly decreased by approximately 24.41 and 23.99%, respectively, on treatment with a medium or high dose of LPTWK10 (\(p<0.05\)) (Figure 5E). These results show that high doses of LPTWK10 effectively lowered the levels of MMP-1 and MMP-9, thereby inhibiting collagen cleavage. AP-1, which is composed of phosphorylated c-Jun and c-Fos as its subunits, plays an essential role in photoaging by upregulating the gene expression levels of MMP-1, MMP-3, and MMP-9, which in turn leads to collagen cleavage in the dermis [46]. The UVB-induced increase in ROS levels stimulates MAPK phosphorylation, which activates the transcription factors AP-1 and NF-kB and regulates MMP-1, MMP-3, and MMP-9, leading to collagen degradation [44]. At week 13, AP-1 levels were significantly higher in the UVB group than those in the NC group by 26.00% (Figure 5F), which significantly decreased by 21.98% on treatment with a high dose of LPTWK10 (\(p<0.05\)). After 8 weeks treated with mixed probiotics, the expressions of MMPs were significantly decreased [47]. Moreover, HR-1 hairless male mice, which induced photoaging by UVB irradiation, were treated with _L. acidophilus_ and exhibited significant downregulation of MMP-1 and MMP-9 expressions [48]. This shows that high doses of LPTWK10 are effective in inhibiting AP-1 levels, which in turn mitigates the increases in MMP-1 and MMP-9 levels, thereby downregulating collagen degradation. By regulating MMPs and enhancing procollagen production, LPTWK10 may help maintain skin elasticity and reduce wrinkle formation, which aligns with the previously reported results on wrinkle count (Figure 1). This mechanism is crucial for understanding how LPTWK10 contributes to skin health and mitigates the effects of photoaging. The studies indicated that _B. breve_ improved UVB-induced photoaging by increasing the activities of anti-oxidation enzymes and decreasing the production of pro-inflammation cytokines [22; 23]. _L. plantarum_ HY7714 has been confirmed through animal studies to improve skin wrinkles by reducing the expressions and activities of MMPs associated with collagen degradation [24]. It is worth mentioning that LPTWK10 and LPTWK10 spray-dried supernatant powders mitigate the photoaging process not only through antioxidant and anti-inflammatory activities but also by reducing collagen degradation via the regulation of MMP expressions.