## 4 Discussion
For instance, a study indicated that different HPL formulations derived from various preparation methods, such as He-HPL and Fd-HPL, were similarly effective in supporting the growth of mesenchymal stem cells [19]. The absence of a statistically significant difference could also be attributed to the inherent biological variability in osteoblast response or the saturation of growth-promoting factors in both HPL preparations. Previous research has shown that growth factors like platelet-derived growth factor (PDGF), transforming growth factor-beta (TGF-\(\beta\)), and VEGF, which are present in HPL, play crucial roles in cellular proliferation and differentiation [20]. Since both He-HPL and Fd-HPL are likely to contain similar concentrations of these growth factors, as indicated by the comparable levels of BDNF and VEGF in this study as well as TGF-\(\beta\)1 and PDGF-BB in the previous study by [PERSON] et al. [14], it is plausible that the cells reached their proliferative capacity under both conditions, leading to the observed similarities. These observations indicate that the addition of calcium chloride to prepare Fd-HPL does not significantly alter the concentration of key bioactive components. Thus, Fd-HPL retains the bioactive properties necessary to support osteoblast proliferation to a degree comparable to He-HPL. This may explain why no differences were observed in the proliferation rate, total cell count, or viability between the two groups. The morphological variations observed between hFOB cells cultured in He-HPL and Fd-HPL on day 1 provide insights into the initial cellular response to these two different platelet lysate preparations. Cells in He-HPL initially exhibited irregular, flattened morphologies and appeared larger compared to those in Fd-HPL, which displayed a more consistent and fibrobl shape. This early difference suggests that He-HPL may induce a distinct early cellular response, potentially due to variations in the concentration of bioactive molecules in the lysate. Growth factors, such as PDGF, TGF-\(\beta\), and VEGF, are known to influence cell adhesion, morphology, and proliferation, and slight variations in their availability between He-HPL and Fd-HPL could explain the different morphologies observed on day 1 [14]. Despite these initial differences, by day 4, hFOB cells in both He-HPL and Fd-HPL cultures exhibited similar spindle-shaped fibroblatic morphologies, indicating that both methods support normal osteoblast behavior over time. The convergence of cell shape by Day 4 suggests that both He-HPL and Fd-HPL provide an adequate environment for osteoblastic growth and proliferation, despite their initial differences in bioactivity. This is consistent with studies that have demonstrated the effectiveness of platelet lysates in promoting mesenchymal stem cell and osteoblast-like cell proliferation and differentiation, regardless of the specific preparation method used [14; 21; 22]. The spindle-shaped fibroblatic morphologies observed in both groups by day 4 align with the expected morphology of osteoblasts in culture, reflecting healthy proliferation and the cells' ability to adapt to their environment. The fact that these morphological differences did not persist beyond day 1 suggests that the initial stress or adaptation period in He-HPL conditions was overcome, leading to normal cellular behavior in the long term. Similar studies have shown that while different platelet lysate preparations can induce varied short-term responses, long-term cell growth and differentiation tend to converge as cells adjust to the culture environment [21]. These findings suggest that both He-HPL and Fd-HPL provide sufficient bioactive factors to support osteoblast-like cell growth, despite slight initial variations in cellular morphology. This also highlights the versatility of hFOB cells in adapting to different culture conditions, as well as the robustness of both forms of HPL in supporting cell viability and morphology over extended culture periods.