Effects of L-arginine and arginine-arginine dipeptide on amino acids uptake and αS1-casein synthesis in bovine mammary epithelial cells.

Arginine (Arg), as an important functional amino acids (AA), is essential for milk protein synthesis in lactating ruminants. Arg shares transporters with cationic and neutral AA in mammary epithelial cells. Therefore, competitive inhibition might exist among these AA in uptake by mammary epithelial cells. In this study, cultured bovine mammary epithelial cells (BMEC) were used as the model to investigate whether the availability of L-Arg (0.7, 1.4, 2.8, 5.6, and 11.2 mM) affects the uptake of other AA and if this related to αS1-casein synthesis, and whether Arginine-Arginine (Arg-Arg) substituting part of free L-Arg can alleviate competitive inhibition among Arg and other AA, so as to promote αS1-casein synthesis. Our results showed that 2.8 mM L-Arg generated the greatest positive effects on αS1-casein synthesis and the activation of mammalian target of rapamycin (mTOR) signaling pathway (P < 0.01). With L-Arg supply increasing from 0.7 to 11.2 mM, the net-uptake of other AA (except Glu and Ala) decreased linearly and quadratically (Plinear < 0.01; Pquadratic < 0.01). Compared with 2.8 mM, the net-uptake of essential amino acids (EAA) and total amino acids (TAA) were lower at 11.2 mM L-Arg group, while greater at 1.4 mM L-Arg group (P < 0.01). Arg-Arg dipeptide replacing 10% free L-Arg increased αS1-casein synthesis (P < 0.05), net-uptake of EAA and TAA, as well as phosphorylation level of mTOR and p70 ribosomal protein S6 kinase (P70S6K) and mRNA expression of oligopeptide transporter 2 (PepT2; P < 0.01). These observations suggested that the increased αS1-casein synthesis by 10% Arg-Arg dipeptide might be related to the increase of AA availability and the activation of mTOR signaling pathway in BMEC.

Arginine (Arg) availability has been demonstrated to affect milk protein synthesis in dairy cows. Competitive inhibition exists among amino acids (AA) in uptake by mammary epithelial cells. This study aims to explore whether the availability of L-Arg affects the uptake of other AA by bovine mammary epithelial cells (BMEC) and if this is related to αS1-casein synthesis, and whether Arginine-Arginine (Arg-Arg) dipeptide substituting part of free L-Arg can alleviate competitive inhibition among Arg and other AA, so as to promote αS1-casein synthesis in BMEC. Our results showed that 2.8 mM L-Arg is the appropriate concentration for αS1-casein synthesis. With L-Arg supply increasing from 0.7 to 11.2 mM, the net-uptake of most AA decreased linearly and quadratically. Arg-Arg dipeptide substituting 10% of free L-Arg increased αS1-casein synthesis and the net-uptake of AA as well as expression of proteins related to mammalian target of rapamycin (mTOR) signaling pathway and mRNA expression of oligopeptide transporter 2 (PepT2). The positive effects of 10% Arg-Arg dipeptide on αS1-casein synthesis may be related to the increase of AA availability and the activation of mTOR signaling pathway.

Effects of glucose availability on αS1-casein synthesis in bovine mammary epithelial cells.

Glucose has been demonstrated to affect milk protein synthesis in dairy cows. However, its potential mechanisms has not been thoroughly studied. The objective of this study was to investigate the effects of glucose availability on αS1-casein synthesis, glucose uptake, metabolism, and the expression of proteins involved in AMP-activated protein kinase (AMPK)/mammalian target of rapamycin (mTOR) signaling pathway in bovine mammary epithelial cells (BMEC). BMEC were treated for 24 h with different concentrations of glucose (0, 7, 10.5, 14, 17.5, and 21 mM). The results showed that 10.5 and 14 mM glucose supply increased the expression of αS1-casein, glucose uptake, cellular ATP content, and the phosphorylation of mTOR and P70S6K, but repressed AMPK phosphorylation in BMEC. Compared with 10.5 and 14 mM glucose supply, 17.5 and 21 mM glucose decreased the expression of αS1-casein, P70S6K phosphorylation as well as the activity of hexokinase (HK) and pyruvate kinase (PK), but increased the activity of glucose-6-phosphate dehydrogenase (G6PD). These results indicate that 10.5 to 14 mM glucose supply is the proper range for αS1-casein synthesis, and the promotion effects may be related to the increase of glucose uptake, ATP content and the changes of key proteins' phosphorylation in AMPK/mTOR signaling pathway. However, the inhibition of the expression of αS1-casein by 17.5 and 21 mM glucose may be associated with the changes of key enzymes' activity involved in glucose metabolism.

Glucose play an important role in milk protein synthesis in dairy cows. But the effects of glucose availability on casein synthesis and its underlying mechanisms has not been thoroughly studied. To elucidate the underlying mechanisms of glucose availability affecting casein synthesis, the effects of glucose availability on αS1-casein synthesis, glucose uptake, metabolism, and the expression of proteins involved in AMP-activated protein kinase (AMPK)/mammalian target of rapamycin (mTOR) signaling pathway in bovine mammary epithelial cells were measured. We found that the expression of αS1-casein increased with 10.5 and 14 mM glucose supplementation, which may be associated with the increase of glucose uptake, ATP content and the changes of key proteins' phosphorylation in AMPK/mTOR signaling pathway. The inhibition of αS1-casein expression with 17.5 and 21 mM glucose supplementation may be related to the changes of key enzymes' activity involved in glucose metabolism. This study provided an insight into the potential mechanisms of glucose availability affecting milk protein synthesis.

Minimally invasive treatment of forearm double fracture in adult using Acumed forearm intramedullary nail: A case report.

BACKGROUND: Currently, open reduction internal fixation is the conventional surgical method for treatment of double ulna and radius fracture. However, open reduction is associated with a high risk of complications. This case of forearm double fracture involved a patient treated using an Acumed intramedullary nail. The patient experienced good follow-up outcomes. The Acumed forearm intramedullary nail enables early functional exercise and hastens healing of the fracture. Few studies have reported on the use of this approach for the treatment of fractures. CASE SUMMARY: A 23-year-old male patient was admitted to hospital after 5 h of pain, swelling, and limited activity of left forearm caused by a careless fall. Physical examination showed stable basic vital signs, swelling of the left forearm, and severe pain when pressing on the injured part of the forearm. Further, friction was felt at the broken end of the bone; the skin was not punctured. Movement of the left hand was normal, and the left radial artery pulse was normal. Three-dimensional computed tomography examination showed an ulna fracture of the left forearm and comminuted fracture of the radius. The fracture was located in the upper third of the radius, with significant displacement on the fracture side. Clinical diagnosis further confirmed the left radius comminuted fracture and ulna fracture. After analyzing the fracture pattern, age, and other patient characteristics, we chose an Acumed nail for treatment and achieved good follow-up outcomes. CONCLUSION: Acumed forearm intramedullary nail for fixation of ulna and radius fracture reduced complication risk and resulted in good follow-up outcomes.

ß sheets not required: combined experimental and computational studies of self-assembly and gelation of the ester-containing analogue of an Fmoc-dipeptide hydrogelator.

In our work toward developing ester-containing self-assembling peptides as soft biomaterials, we have found that a fluorenylmethoxycarbonyl (Fmoc)-conjugated alanine-lactic acid (Ala-Lac) sequence self-assembles into nanostructures that gel in water. This process occurs despite Fmoc-Ala-Lac's inability to interact with other Fmoc-Ala-Lac molecules via ß-sheet-like amide-amide hydrogen bonding, a condition previously thought to be crucial to the self-assembly of Fmoc-conjugated peptides. Experimental comparisons of Fmoc-Ala-Lac to its self-assembling peptide sequence analogue Fmoc-Ala-Ala using a variety of microscopic, spectroscopic, and bulk characterization techniques demonstrate distinct features of the two systems and show that while angstrom-scale self-assembled structures are similar, their nanometer-scale size and morphological properties diverge and give rise to different bulk mechanical properties. Molecular dynamics simulations were performed to gain more insight into the differences between the two systems. An analysis of the hydrogen-bonding and solvent-surface interface properties of the simulated fibrils revealed that Fmoc-Ala-Lac fibrils are stronger and less hydrophilic than Fmoc-Ala-Ala fibrils. We propose that this difference in fibril amphiphilicity gives rise to differences in the higher-order assembly of fibrils into nanostructures seen in TEM. Importantly, we confirm experimentally that ß-sheet-type hydrogen bonding is not crucial to the self-assembly of short, conjugated peptides, and we demonstrate computationally that the amide bond in such systems may act mainly to mediate the solvation of the self-assembled single fibrils and therefore regulate a more extensive higher-order aggregation of fibrils. This work provides a basic understanding for future research in designing highly degradable self-assembling materials with peptide-like bioactivity for biomedical applications.

Experimental and computational studies reveal an alternative supramolecular structure for fmoc-dipeptide self-assembly.

We have investigated the self-assembly of fluorenylmethoxycarbonyl-conjugated dialanine (Fmoc-AA) molecules using combined computational and experimental approaches. Fmoc-AA gels were characterized using transmission electron microscopy (TEM), circular dichroism (CD), Fourier transform infrared (FTIR), and wide-angle X-ray scattering (WAXS). Computationally, we simulated the assembly of Fmoc-AA using molecular dynamics techniques. All simulations converged to a condensed fibril structure in which the Fmoc groups stack mostly within in the center of the fibril. However, the Fmoc groups are partially exposed to water, creating an amphiphilic surface, which may be responsible for the aggregation of fibrils into nanoscale fibers observed in TEM. From the fibril models, radial distribution calculations agree with d-spacings observed in WAXS for the fibril diameter and π-stacking interactions. Our analyses show that dialanine, despite its short length, adopts a mainly extended polyproline II conformation. In contrast to previous hypotheses, these results indicate that ß-sheet-like hydrogen bonding is not prevalent. Rather, stacking of Fmoc groups, inter-residue hydrogen bonding, and hydrogen bonding with water play the important roles in stabilizing the fibril structure of supramolecular assemblies of short conjugated peptides.