Alfalfa xeno-miR159a regulates bovine mammary epithelial cell proliferation and milk protein synthesis by targeting PTPRF.

Milk protein content is an important index to evaluate the quality and nutrition of milk. Accumulating evidence suggests that microRNAs (miRNAs) play important roles in bovine lactation, but little is known regarding the cross-kingdom regulatory roles of plant-derived exogenous miRNAs (xeno-miRNAs) in milk protein synthesis, particularly the underlying molecular mechanisms. The purpose of this study was to explore the regulatory mechanism of alfalfa-derived xeno-miRNAs on proliferation and milk protein synthesis in bovine mammary epithelial cells (BMECs). Our previous study showed that alfalfa miR159a (mtr-miR159a, xeno-miR159a) was highly expressed in alfalfa, and the abundance of mtr-miR159a was significantly lower in serum and whey from high-protein-milk dairy cows compared with low-protein-milk dairy cows. In this study, mRNA expression was detected by real-time quantitative PCR (qRT-PCR), and casein content was evaluated by enzyme-linked immunosorbent assay (ELISA). Cell proliferation and apoptosis were detected using the cell counting kit 8 (CCK-8) assay, 5-ethynyl-2'-deoxyuridine (EdU) staining, western blot, and flow cytometry. A dual-luciferase reporter assay was used to determine the regulation of Protein Tyrosine Phosphatase Receptor Type F (PTPRF) by xeno-miR159a. We found that xeno-miR159a overexpression inhibited proliferation of BMEC and promoted cell apoptosis. Besides, xeno-miR159a overexpression decreased ß-casein abundance, and increased α-casein and κ-casein abundance in BMECs. Dual-luciferase reporter assay result confirmed that PTPRF is a target gene of xeno-miR159a. These results provide new insights into the mechanism by which alfalfa-derived miRNAs regulate BMECs proliferation and milk protein synthesis.

Alfalfa xenomiR-162 targets G protein subunit gamma 11 to regulate milk protein synthesis in bovine mammary epithelial cells.

OBJECTIVE: It was shown that microRNAs (miRNAs) play an important role in milk protein synthesis. However, the post-transcriptional regulation of casein expression by exogenous miRNA (xeno-miRNAs) in ruminants remains unclear. This study explores the regulatory roles of alfalfa xeno-miR162 on casein synthesis in bovine mammary epithelial cells (bMECs). METHODS: The effects of alfalfa xenomiR-162 and G protein subunit gamma 11 (GNG11) on proliferation and milk protein metabolism of bMECs were detected by 5-Ethynyl-2'-Deoxyuridine (EdU) staining, flow cytometry, cell counting kit-8 (CCK-8), enzyme-linked immunosorbent assay, quantitative real-time polymerase chain reaction (qRT-PCR), and Western blot. Dual-luciferase reporter assay was used to verify the targeting relationship between GNG11 and xenomiR-162. RESULTS: Results showed that over-expression of xenomiR-162 inhibited cell proliferation but promoted apoptosis, which also up-regulated the expression of several casein coding genes, including CSN1S1, CSN1S2, and CSN3, while decreasing the expression of CSN2. Furthermore, the targeting relationship between GNG11 and xenomiR-162 was determined, and it was confirmed that GNG11 silencing also inhibited cell proliferation but promoted apoptosis and reduced the expression of casein coding genes and genes related to the mammalian target of rapamycin (mTOR) pathway. CONCLUSION: Alfalfa xenomiR-162 appears to regulate bMECs proliferation and milk protein synthesis via GNG11 in the mTOR pathway, suggesting that this xeno-miRNA could be harnessed to modulate CSN3 expression in dairy cows, and increase κ-casein contents in milk.

High-Grade Ferronickel Concentrates Prepared from Laterite Nickel Ore by a Carbothermal Reduction and Magnetic Separation Method.

Nickel is widely used in industrial processes and plays a crucial role in many applications. However, most of the nickel resource mainly exists as nickel oxide in laterite nickel ore with complex composition, resulting in difficulty in upgrading the nickel content using physical separation methods. In this study, high-grade ferronickel concentrates were obtained through a carbothermal reduction and magnetic separation using laterite nickel ore and anthracite as raw materials. The effects of different types of additives (CaF2, Na2SO4, and H3BO3), carbon ratio (the molar ratio of oxygen atoms in the laterite nickel ore to carbon atoms in anthracite), and grinding time on the recoveries and grades of ferronickel concentrates were experimentally investigated, along with the microstructure and chemical composition of the products. CaF2 was proved to be the primary active additive in the aggregation and growth of the ferronickel particles and the improvement of the grade of the product. Under the optimal conditions of CaF2 addition of 9.85 wt%, carbon ratio of 1.4, and grinding time of 240 s, high-grade magnetically separable ferronickel concentrate with nickel grade 8.93 wt% and iron grade 63.96 wt% was successfully prepared. This work presents a practical method for the highly efficient recovery and utilization of iron and nickel from low-grade laterite nickel ore, contributing to the development of strategies for the sustainable extraction and utilization of nickel resources.

Alfalfa Xeno-miR168b Target CPT1A to Regulate Milk Fat Synthesis in Bovine Mammary Epithelial Cells.

It was shown that microRNAs (miRNAs) play an important role in the synthesis of milk fat; thus, this manuscript evaluated whether exogenous miRNA (xeno-miRNAs) from alfalfa could influence the milk fat content in dairy cows. At first, mtr-miR168b was screened from dairy cow milk and blood. Then, EdU staining, flow cytometry, Oil Red O staining, qRT-PCR, and WB were applied to explore the effect of xeno-miR168b on the proliferation, apoptosis, and lipid metabolism of bovine mammary epithelial cells (BMECs). Finally, in order to clarify the pathway that regulated the lipid metabolism of BMECs using xeno-miR168b, a double-luciferase reporter assay was used to verify the target gene related to milk fat. These results showed that overexpression of xeno-miR168b inhibited cell proliferation but promoted apoptosis, which also decreased the expression of several lipid metabolism genes, including PPARγ, SCD1, C/EBPß, and SREBP1, significantly inhibited lipid droplet formation, and reduced triglyceride content in BMECs. Furthermore, the targeting relationship between CPT1A and xeno-miR168b was determined and it was confirmed that CPT1A silencing reduced the expression of lipid metabolism genes and inhibited fat accumulation in BMECs. These findings identified xeno-miR168b from alfalfa as a cross-kingdom regulatory element that could influence milk fat content in dairy cows by modulating CPT1A expression.

A robust air superhydrophilic/superoleophobic diatomite porous ceramic for high-performance continuous separation of oil-in-water emulsion.

Three-dimensional (3D) porous architecture has attracted considerable attention in remediation of oil/water emulsion. In present work, an air superhydrophilic/superoleophobic diatomite porous ceramic (AS-DC) was prepared, using SiO2 whiskers modified diatomite ceramic as the substrate and FS-50 as the modifier. The interconnected SiO2 whiskers intertwined on the skeleton of ceramic block forming a 3D network structure, which not only improved the wettability of AS-DC, but also reinforced its mechanical property (about 2.5 MPa of compressive strength). The as-prepared AS-DC with intrinsically superoleophobicity (154°) and superhydrophilicity (0°) exhibited an underwater oil contact angle of 161°, suggesting a multifunctional separation capability. By simply assembling AS-DC with pipes and a pump, it could not only separate the surfactant-stabilized oil-in-water emulsion in a permeation flux as high as 107.8 kg min-1 m-2 with a selectivity of >95%, but also collect the clean water from the floating oil/water mixture in a flux of 197.4 kg min-1 m-2 and a selectivity of ∼99%. In addition, the AS-DC was resistant to the salt/acid/alkaline corrosion and temperature fluctuation. The mechanical/chemical firmness of AS-DC renders it tremendous potential as a robust 3D architecture in real application for purification of oil/water mixture.