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Rainbow trout bone proteins were prepared by heating at 121°C for 30 min, followed by filtration, concentration, and lyophilization. Nutritional properties and flavor analyses of hydrolysates digested by five different enzymes were investigated, respectively. Results showed that the crude protein content of rainbow trout bone was 15.90% and had a well‐balanced nutritional value. The content of total amino acids was 983.64 mg/g. The amount of free amino acids of hydrolysates digested by alkaline protease, neutral protease, flavourzyme, papain, and trypsin for 3 hr was 207.83, 224.13, 1,001.59, 283.26, and 303.64 mg/g, respectively. During the hydrolysis, the main flavor compounds were identified by GC‐MS to be alcohols, aldehydes, ketones, acids, and alkanes. After hydrolysis, the main molecular weight of peptides was focused on the range of 1,000–3,000 Da in all enzymatic hydrolysates. This study provided a theoretical basis to comprehensive utilization of rainbow trout bone in food industry.

Hydrolysis of the proteins from cod bone was performed using flavourzyme and trypsin. The nutritional properties of hydrolysates by flavourzyme (HF) and trypsin (HT) were investigated. By comparison, HF exhibits a better degree of hydrolysates (DH) and nitrogen recovery (NR) than HT. The protein content of extract is 97.39% and had a good nutritional value due to the high protein digestibility-corrected amino acid score (PDCAAS) of 0.95 for adults. The content of total amino acids is 942.55 mg/g. The free amino acids content of hydrolysates derived from flavourzyme is 136.82 mg/g after hydrolyzing 3 h, while 17.12 mg/g of such hydrolysates was obtained by using trypsin. The main flavor compounds are alcohols, aldehydes, ketones, acids and alkanes. The molecular weight of predominant peptides ranged from 1000 to 3000 Da in both enzymatic hydrolysates. This study provided a theoretical basis to the preparation of nutritional components with attracting flavor in functional food industry.

Fluid shear stress (FSS) is a potent mechanical stimulus and prevents cells from TNF-a-induced apoptosis. Recently, Extracellular-signal-regulated kinase 5 (ERK5) has been found to be involved in regulation of cell survival. However, little is known about the role of ERK5 signaling pathway in FSS-mediated anti-apoptotic effects in osteoblast. In this study, we show that FSS blocks TNF-a-induced apoptosis of MC3T3-E1 cells via ERK5 signaling pathway. We found that physiological FSS for 1 h significantly decreased TNF-α-induced MC3T3-E1 cells apoptosis. After inhibition of ERK5 activity by XMD8-92, a highly-selective inhibitor of ERK5 activity, the ability of FSS to inhibit TNF-α induced apoptosis was significantly decreased. Analysis of anti-apoptotic mechanisms indicated that exposure of MC3T3-E1 cells to FSS for 1 h increased phosphorylation of Bad and inhibited caspase-3 activity. After treatment with XMD8-92, phosphorylation of Bad by FSS was significantly blocked, but caspase-3 activity was increased. In summary, these findings indicated that FSS inhibits TNF-α-mediated signaling events in osteoblast by a mechanism dependent on activation of ERK5, and Bad is a crucial downstream target for ERK5. Those results implied that ERK5 signaling pathway play a crucial role in FSS-mediated anti-apoptotic effect in osteoblast. Thus, ERK5 signaling pathway may be a new drug treatment target of osteoporosis and related bone-wasting diseases.

Alaska pollock frame is a kind of byproduct that was rich in protein, amino acids, and mineral elements. However, the unfavorite bitterness may probably be produced in enzymatic processes. In this study, the bitterness accounted from the hydrolysates prepared by neutral proteases, alkaline proteases, papain, flavourzyme, and animal proteases, was investigated. The hydrolysis conditions, amino acids composition, metal ion composition, molecular weight distribution, and peptide composition of the hydrolysates were detected to figure out the relationship between bitterness and compositions of the hydrolysates. The hydrolysate digested by alkaline protease has the highest bitterness intensity, and that amino acids composition, peptide composition, and molecular weight distribution had a significant influence on the bitterness degree. Hydrophobic amino acids and alkaline amino acids, such as leucine, isoleucine, lysine, and so on are likely to contribute to the bitterness and molecular weight distribution of peptides that affect bitterness is mainly lower than 3,000 Da. PRACTICAL APPLICATIONS: Fish bones would produce a bitter taste when reusing them by hydrolysis. Bitterness is one of unfavorable flavor as to consumers. The results of this study are of great significance for the further utilization of Alaska pollock frame. For products obtained from the hydrolysate of Alaska pollock frame, such as condiments and health care product the results of this study provide the processing technology of the lowest bitter hydrolysate, which can effectively improve the flavor and acceptability of the products.

Fluid shear stress (FSS) is a ubiquitous mechanical stimulus that potently promotes osteoblast proliferation. Previously, we reported that extracellular signal-regulated kinase 5 (ERK5) is essential for FSS-induced osteoblast proliferation. However, the precise mechanism by which FSS promotes osteoblast proliferation via ERK5 activation is poorly understood. The aim of this study was to determine the critical role of Gαq in FSS-induced ERK5 phosphorylation and osteoblast proliferation, as well as the downstream targets of the Gαq-ERK5 pathway. MC3T3-E1 cells were transfected with 50 nM Gαq siRNA, treated with 5 mM XMD8-92 (a highly selective inhibitor of ERK5 activity), and/or exposed to FSS (12 dyn/cm(2)). Cell proliferation was evaluated using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The protein expression levels of Gαq, P-ERK5, ERK5, Cyclin B1, and CDK1 were analyzed by Western blot. Physiological FSS exposure for 60 min remarkably promoted MC3T3-E1 cell proliferation, however, this effect was suppressed by siRNA-mediated Gαq knockdown or inhibition of ERK5 activity by XMD8-92 treatment, suggesting that Gαq and ERK5 might modulate FSS-increased osteoblast proliferation. Furthermore, ERK5 phosphorylation was dramatically inhibited by Gαq siRNA. In addition, our study further revealed that FSS treatment of MC3T3-E1 cells for 60 min markedly upregulated the protein expression levels of Cyclin B1 and CDK1, and this increased expression was predominantly blocked by Gαq siRNA or XMD8-92 treatment. We propose that FSS acts on the Gαq-ERK5 signaling pathway to upregulate Cyclin B1 and CDK1 expression, thereby resulting in MC3T3-E1 cell proliferation. Thus, the Gαq-ERK5 signaling pathway may provide useful information regarding the treatment of bone metabolic disease.

TNF-α is known to induce osteoblasts apoptosis, whereas mechanical stimulation has been shown to enhance osteoblast survival. In the present study, we found that mechanical stimulation in the form of fluid shear stress (FSS) suppresses TNF-α induced apoptosis in MC3T3-E1 cells. Extracellular signal-regulated kinase 5 (ERK5) is a member of the mitogen-activated protein kinase (MAPK) family that has been implicated in cell survival. We also demonstrated that FSS imposed by flow chamber in vitro leads to a markedly activation of ERK5, which was shown to be protective against TNF-α-induced apoptosis, whereas the transfection of siRNA against ERK5 (ERK5-siRNA) reversed the FSS-medicated anti-apoptotic effects. An initial FSS-mediated activation of ERK5 that phosphorylates AKT to increase its activity, and a following forkhead box O 3a (FoxO3a) was phosphorylated by activated AKT. Phosphorylated FoxO3a is sequestered in the cytoplasm, and prevents it from translocating to nucleus where it can increase the expression of FasL and Bim. The inhibition of AKT-FoxO3a signalings by a PI3K (PI3-kinase)/AKT inhibitor (LY294002) or the transfection of ERK5-siRNA led to the nuclear translocation of non-phosphorylated FoxO3a, and increased the protein expression of FasL and Bim. In addition, the activation of caspase-3 by TNF-α was significantly inhibited by aforementioned FSS-medicated mechanisms. In brief, the activation of ERK5-AKT-FoxO3a signaling pathways by FSS resulted in a decreased expression of FasL and Bim and an inhibition of caspase-3 activation, which exerts a protective effect that prevents osteoblasts from apoptosis.