A Low-Phenylalanine-Containing Whey Protein Hydrolysate Stimulates Osteogenic Activity through the Activation of p38/Runx2 Signaling in Osteoblast Cells

Abstract

A phenylalanine (Phe)-restricted diet is indispensable for individuals suffering from phenylketonuria (PKU). Our previous study reported a low-Phe-containing whey protein hydrolysate (LPH) prepared from a selected whey protein hydrolysate (TA2H). This study aimed to investigate the osteogenic activity of LPH and TA2H in MC3T3-E1 preosteoblast cells and explore the underlying mechanism. Results showed that the treatment of TA2H and LPH (at the final concentrations of 100–1000 μg/mL) had a stimulatory effect on the proliferation, differentiation, and mineralization of MC3T3-E1 cells. The LPH of 1000 μg/mL significantly increased cell proliferation (2.15- ± 0.11-fold) and alkaline phosphatase activity (1.22- ± 0.07-fold), promoted the protein and mRNA levels of runt-related transcription factor 2 (Runx2, 2.50- ± 0.14-fold and 2.97- ± 0.23-fold, respectively), enhanced the expression of differentiation biomarkers (type-I collagen, osteocalcin, and osteopontin), increased calcium deposition (1.56- ± 0.08-fold), and upregulated the ratio of osteoprotegerin/receptor activator of nuclear factor-κB ligand. The exploration of signaling pathways indicated that the activated p38-dependent Runx2 signaling contributed to the LPH-induced osteogenesis. These results provided evidence, for the first time, that a prepared low-Phe whey protein hydrolysate positively modulated the activity of osteoblasts through the p38/Runx2 pathway, thereby providing a new osteoinductive protein substitute to make functional PKU food.

Whey Protein Hydrolysate Ameliorated High-Fat-Diet Induced Bone Loss via Suppressing Oxidative Stress and Regulating GSK-3β/Nrf2 Signaling Pathway

“…Our previous studies reported that the WPH could regulate the homeostasis of bone metabolism in vitro and in vivo [ 25 , 27 ]. To further identify the specific peptides with high osteogenic and antioxidative potentials, WPH was separated into 16 fractions by RP-HPLC ( Figure 1 a); among which F9 possessed both significantly higher osteogenic capacity and DPPH scavenging activity than other fractions ( Figure 1 b).…”

“…Whey-derived antioxidant peptide YVEEL ameliorated osteoporosis in an ovariectomized rat model by suppressing inflammation and enhancing osteogenesis [ 24 ]. We previously reported a whey protein enzymatic hydrolysate (WPH) exerted effects on activating osteogenesis and preventing ovariectomy-induced osteoporosis through regulating the p38/mitogen-activated protein kinase (MAPK)/Runx2 signaling pathway [ 25 , 26 , 27 ]. Although the anti-obesity/adipogenesis function of milk proteins, as well as their derived bioactive peptides, has been reported previously [ 28 , 29 ], the regulation effects of WPH on HFD-induced bone loss through redox modulation have not been sufficiently investigated.…”

Yunnan Baiyao Might Mitigate Periodontitis Bone Destruction by Inhibiting Autophagy and Promoting Osteoblast Differentiation in vivo, ex vivo and in vitro

“…7 Conversely, osteoclastogenesis inhibitory factor (osteoprotegerin, OPG), a soluble protein from osteoblasts, competes with receptor activator of nuclear factor-kappaB ligand (RANKL) to antagonize the RANK-RANKL interaction, thereby inhibiting osteoclastogenesis and preserving the osteogenic environment. 8,9 Therefore, osteogenesis is a key process in the inhibition of bone loss.…”

Endothelialization of Whey Protein Isolate-Based Scaffolds for Tissue Regeneration

“…In endothelial cells exposed to diabetic conditions, treatment with whey hydrolysate prevented mitochondrial injury and redox imbalance [25]. Increased GSH levels in plasma and erythrocytes have also been reported in rats which were fed a diet supplemented with 10% whey proteins [26,27], but the evidence for its protective effects on vascular functions in humans remains unsubstantiated [28] Regarding tissue engineering applications, several reports have indicated that WPI dissolved in cell culture medium stimulates the osteogenic differentiation of bone-forming cells [12,[29][30][31]. However, in the majority of the previous studies, WPI has been used only as a supplement for cell culture medium, in the form of lysate or dissolved powder.…”