Epigallocatechin-3-gallate-induced tolerance to cadmium stress involves increased flavonoid synthesis and nutrient homeostasis in tomato roots.

Cadmium (Cd) is a toxic heavy metal, increasingly accumulating in the environment and its presence in various environmental compartments represents a significant risk to human health via the food chain. Epigallocatechin-3-Gallate (EGCG) is a prominent secondary metabolite, which can safeguard plants from biotic and abiotic stress. However, the role of EGCG in flavonoid synthesis, nutrient acquisition and reactive oxygen species (ROS) metabolism under Cd stress remains unclear. Here, we examined the effects of EGCG and Cd treatment on leaf photochemical efficiency, cell ultrastructure, essential element acquisition, antioxidant system, and secondary metabolism in tomato (Solanum lycopersicum L.). The results showed that O2•-, H2O2, and malondialdehyde levels increased after Cd treatment, but Fv/Fm decreased significantly, suggesting that Cd induced oxidative stress and photoinhibition. However, EGCG mitigated the adverse effects of Cd-induced phytotoxicity in both the roots and leaves. A decrease in ROS accumulation under EGCG + Cd treatment was mainly attributed to the significant enhancement in antioxidant enzyme activity, flavonoid content, and PHENYLALANINE AMMONIA-LYASE expression in roots. Moreover, EGCG reduced Cd content but increased some essential nutrient contents in tomato plants. Transmission electron microscopy-based observations revealed that EGCG treatment safeguards leaf and root cell ultrastructure under Cd stress. This implies that tomato plants subjected to Cd stress experienced advantageous effects upon receiving EGCG treatment. The present work elucidated critical mechanisms by which EGCG induces tolerance to Cd, thereby providing a basis for future investigations into environmentally sustainable agricultural practices in areas contaminated with heavy metals, for utilizing naturally occurring substances found in plants.

Effect of Human Milk Oligosaccharides on Learning and Memory in Mice with Alzheimer's Disease.

Alzheimer's disease (AD) is distinguished by cognitive dysfunction and neuroinflammation in the brain. 2'-Fucosyllactose (2'-FL) is a major human milk oligosaccharide (HMO) that is abundantly present in breast milk and has been demonstrated to exhibit immunomodulatory effects. However, the role of 2'-FL and HMO in gut microbiota modulation in relation to AD remains insufficiently investigated. This study aimed to elucidate the preventive effect of the 2'-FL and HMO impact of AD and the relevant mechanism involved. Here, the behavioral results showed that 2'-FL and HMO intervention decreased the expression of Tau phosphorylation and amyloid-ß (Aß), inhibited neuroinflammation, and restored cognitive impairment in AD mice. The metagenomic analysis proved that 2'-FL and HMO intervention restored the dysbiosis of the gut microbiota in AD. Notably, 2'-FL and HMO intervention significantly enhanced the relative abundance of Clostridium and Akkermansia. The metabolomics results showed that 2'-FL and HMO enhanced the oleoyl-l-carnitine metabolism as potential drivers. More importantly, the levels of oleoyl-l-carnitine were positively correlated with the abundances of Clostridium and Akkermansia. These results indicated that 2'-FL and HMO had therapeutic potential to prevent AD-induced cognitive impairment, which is of great significance for the treatment of AD.

Effect of Thermal Inactivation on Antioxidant, Anti-Inflammatory Activities and Chemical Profile of Postbiotics.

Inactivation is a crucial step in the production of postbiotics, with thermal inactivation being the prevailing method employed. Nevertheless, the impact of thermal treatment on bioactivity and chemical composition remains unexplored. The objective of this study was to assess the influence of heating temperature on the antioxidant, anti-inflammatory properties and the chemical composition of ET-22 and BL-99 postbiotics. The findings revealed that subjecting ET-22 and BL-99 to thermal treatment ranging from 70 °C to 121 °C for a duration of 10 min effectively deactivated them, leading to the disruption of cellular structure and release of intracellular contents. The antioxidant and anti-inflammatory activity of ET-22 and BL-99 postbiotics remained unaffected by mild heating temperatures (below 100 °C). However, excessive heating at 121 °C diminished the antioxidant activity of the postbiotic. To further investigate the impact of thermal treatments on chemical composition, non-targeted metabolomics was conducted to analyze the cell-free supernatants derived from ET-22 and BL-99. The results revealed that compared to mild inactivation at temperatures below 100 °C, the excessive temperature of 121 °C significantly altered the chemical profile of the postbiotic. Several bioactive components with antioxidant and anti-inflammatory properties, including zomepirac, flumethasone, 6-hydroxyhexanoic acid, and phenyllactic acid, exhibited a significant reduction in their levels following exposure to a temperature of 121 °C. This decline in their abundance may be associated with a corresponding decrease in their antioxidant and anti-inflammatory activities. The cumulative evidence gathered strongly indicates that heating temperatures exert a discernible influence on the properties of postbiotics, whereby excessive heating leads to the degradation of heat-sensitive active constituents and subsequent diminishment of their biological efficacy.

Influence of Temperature during Freeze-Drying Process on the Viability of Bifidobacterium longum BB68S.

Maintaining optimum temperature during freeze-drying is crucial to ensuring the viability of strains. In this study, we evaluated the effect of pre-freezing, sublimation and desorption temperatures on the viability of Bifidobacterium longum BB68S (BB68S). Moreover, we examined the water content, water activity, enzyme activities, and scanning electron microscope of BB68S to explore mechanisms underpinning the effect of temperature on viability. Our analyses revealed the highest survival rates of BB68S collected after pre-freezing and sublimation drying at -40 °C (94.9 ± 2.2%) and -10 °C (65.4 ± 3.8%), respectively. Additionally, response surface methodology demonstrated that the optimum conditions for freeze-drying of BB68S were pre-freezing temperature at -45.52 °C and sublimation temperature at -6.58 °C, and the verification test showed that survival rates of BB68S could reach 69.2 ± 3.8%. Most of the vitality loss occurred during the sublimation drying phase. Further studies showed that different sublimation temperatures affected water content and activity, ß-galactosidase, lactate dehydrogenase, Na+-K+-ATP and Ca2+-Mg2+-ATP activities. In conclusion, the temperature during freeze-drying, especially sublimation temperature, is a key factor affecting the survival rate of BB68S, and the vitality loss during freeze-drying process might be due to compromised cell membrane integrity and permeability.

Research on bacterial community characteristics of traditional fermented yak milk in the Tibetan Plateau based on high-throughput sequencing.

Background: The Tibetan Plateau has an abundance of yak milk resources. The complex microbiota found in traditional fermented yak milk produced and sold by local Tibetans endows the yak milk with unique quality characteristics such as tissue morphology, flavor, and function. However, the diversity of bacterial flora in traditional fermented yak milk have not been elucidated. Methods: In this study, 15 samples of fermented yak milk were collected for 16S rRNA high-throughput sequencing to analyze the bacterial community composition and function. Results: After filtering for quality, 792,642 high-quality sequences were obtained, and 13 kinds of different phyla and 82 kinds of different genera were identified, of which the phylum Firmicutes (98.94%) was the dominant phylum, Lactobacillus (64.73%) and Streptococcus (28.48%) were identified as the dominant genus, in addition, the bacterial community richness and diversity were higher in Manang Village, followed by Bola Village. Bacterial community richness and diversity in Huage Village were relatively low. Based on the Kyoto Encyclopedia of Genes and Genomes (KEGG) functional classification, the microorganisms in traditional fermented yak milk have rich metabolic functions (77.60%). These findings suggest that a large number of bacteria in traditional fermented yak milk contain abundant metabolic genes and can carry out a variety of growth and metabolic activities. This study established a theoretical foundation for further exploring the microbial flora of traditional fermented yak milk in Gannan.

3,5,6-trichloro-2-pyridinol intensifies the effect of chlorpyrifos on the paracrine function of Sertoli cells by preventing binding of testosterone and the androgen receptor.

3,5,6-Trichloro-2-pyridinol (TCP) is an important biomarker and one of the final metabolites of chlorpyrifos (CPF). TCP inhibits secretion of sex hormones. Similar to CPF, TCP can bind to sex steroid hormone receptors and decrease the secretion of sex hormones. However, little attention has been paid to the ability of TCP and CPF to interfere with androgen receptor (AR) in Sertoli cells. This study aimed to explain how TCP promotes the inhibitory effect of CPF on the paracrine function of Sertoli cells. Western blotting indicated that after 20 weeks of exposure, expression of AR in testes was significantly reduced by CPF. An in vitro assay measured the cytotoxicity of CPF, TCP and diethylphosphate (DEP) on viability of Sertoli cells by Cell Counting Kit-8. CPF cytotoxicity was greater than that of TCP, and TCP cytotoxicity was greater than that of DEP at concentrations of 1000 µmol/L. Western blotting indicated that TCP and CPF both decreased expression of AR and cAMP-response element binding protein phosphorylation, while DEP had no effect in Sertoli cells, which are important in regulating paracrine function of Sertoli cells. The fluorescence measurements and docking studies revealed that testosterone, CPF and TCP showed four types of intermolecular interactions with AR, highlighting alkyl bonds with some of the same amino acids. Compared with testosterone, CPF and TCP also showed significant synergistic interaction with AR. CPF interacted with more amino acids and interaction energy than TCP did. This research elucidates TCP in the antiandrogenic effect of CPF on the paracrine function and suggests that TCP or chemicals with a trichloropyridine structure must be considered during reproductive toxicity assessment of potential environmental pollutants.

Visfatin induces the apoptosis of endothelial progenitor cells via the induction of pro-inflammatory mediators through the NF-κB pathway.

Endothelial progenitor cells (EPCs) are an independent factor predicting cardiovascular events. Visfatin plays an important role in the pathogenesis of various metabolic disorders. In this study, we examined the effects of visfatin on the apoptosis of EPCs and the mechanisms underlying these effects. Cultured EPCs pre-treated with various concentrations of visfatin, FK866 (visfatin inhibitor) and BAY11-7085 [referred to as BAY11; nuclear factor-κB (NF-κB) inhibitor] were used to investigate the association between visfatin and EPC apoptosis. Following treatment with visfatin for 48 h, the EPCs exhibited a dose-dependent increase in apoptosis and an upregulated expression of Bax, caspase-3 and NF-κB at both the mRNA and protein level, and a decreased protein expression of Bcl-2. Compared with the untreated control group, the increase in EPC apoptosis, as well as in Bax and caspase-3 expression was significant following treatment with 150 ng/ml visfatin, which also induced a dose-dependent and significant increase in the protein expression of interleukin-6 (IL-6) and intercellular adhesion molecule-1 (ICAM-1). All the visfatin-induced effects were suppressed by pre-treatment with FK866. Pre-incubation of the EPCs with BAY11 for 1 h followed by treatment with visfatin (150 ng/ml) for 48 h also abolished visfatin-induced apoptosis; it also abolished the promoting effects of visfatin on the expression of caspase-3, Bax, ICAM-1 and IL-6, and its suppressive effects on the protein expression of Bcl-2. On the whole, our data indicate that visfatin induces EPC apoptosis by increasing the expression of pro-inflammatory mediators partly through the regulation of NF-κB.

Immortalized bovine mammary epithelial cells express stem cell markers and differentiate in vitro.

The bovine mammary epithelial cell is a secretory cell, and its cell number and secretory activity determine milk production. In this study, we immortalized a bovine mammary epithelial cell line by SV40 large T antigen gene using a retrovirus based on Chinese Holstein primary mammary epithelial cells (CMEC) cultured in vitro. An immortalized bovine mammary epithelial cell line surpassed the 50-passage mark and was designated the CMEC-H. The immortalized mammary epithelial cells grew in close contact with each other and exhibited the typical cobblestone morphology characteristic with obvious boundaries. The telomerase expression of CMEC-H has consistently demonstrated the presence of telomerase activity as an immortalized cell line, but the cell line never induced tumor formation in nude mice. CMEC-H expressed epithelial (cytokeratins CK7, CK8, CK18, and CK19), mesenchymal (vimentin), and stem/progenitor (CD44 and p63) cell markers. The induced expression of milk proteins, αS1 -casein, ß-casein, κ-casein, and butyrophilin, indicated that CMEC-H maintained the synthesis function of the mammary epithelial cells. The established immortalized bovine mammary epithelial cell line CMEC-H is capable of self-renewal and differentiation and can serve as a valuable reagent for studying the physiological mechanism of the mammary gland.

Leucine and histidine independently regulate milk protein synthesis in bovine mammary epithelial cells via mTOR signaling pathway.

The aim of this study is to investigate the effects of leucine (Leu) and histidine (His) on the expression of both the mammalian target of rapamycin (mTOR) signaling pathway-related proteins and caseins in immortalized bovine mammary epithelial cells (CMEC-H), using a single supplement through Western blotting. The Earle's balanced salt solution (EBSS) was set as the control group and other treatment groups, based on the EBSS, were added with different concentrations of Leu or His, respectively. The results showed that, compared with the control group, the expression of caseins and the phosphorylation of mTOR (Ser(2481)), Raptor (Ser(792)), eIF4E (Ser(209)), and eEF2 (Thr(56)) increased with the Leu concentrations ranging from 0.45 to 10.80 mmol/L (P<0.01). The P-4EBP1 (Thr(37)) at 10.80 mmol/L Leu, and P-RPS6 (Ser(235/236)) at 5.40 to 10.80 mmol/L Leu all decreased. Similarly, the His supplementation from 0.15 to 9.60 mmol/L increased the expression of αs2-casein, ß-casein, κ-casein, P-mTOR (Ser(2481)), P-Raptor (Ser(792)), P-S6K1 (Thr(389)), P-4EBP1 (Thr(37)), P-eIF4E (Ser(209)), and P-eEF2 (Thr(56)) (P<0.01) in CMEC-H, whereas the αs1-casein expression was only reduced at 9.60 mmol/L His, G protein ß subunit-like protein (GßL) at 0.15 and 9.60 mmol/L His, and P-RPS6 at 4.80 to 9.60 mmol/L His. Our linear regression model assay suggested that the αs1-casein expression was positively correlated with P-mTOR (P<0.01), P-S6K1 (P<0.01), and P-eEF2 (P<0.01) for the addition of Leu, while the expressions of ß-casein (P<0.01) and κ-casein (P<0.01) were positively correlated with P-eEF2 for the addition of His. In conclusion, the milk protein synthesis was up-regulated through activation of the mTOR pathway with the addition of Leu and His in CMEC-H.

Visfatin and oxidative stress influence endothelial progenitor cells in obese populations.

UNLABELLED: PURPOSE/AIMS OF THE STUDY: This study sought to study the levels of circulating endothelial progenitor cells (EPCs), serum visfatin, and oxidative stress in obese individuals, and their respective correlations. MATERIALS AND METHODS: The circulating levels of EPCs were measured through detecting CD309 and CD34 by flow cytometry. Serum visfatin concentration was measured by enzyme-linked immunosorbent assay in 31 obese men [obese group: BMI 28.9 ± 0.86 kg/m(2); age 44.93 ± 1.78 years (range 40 to 47)] and 30 normal-weight men [control group: BMI 22.7 ± 1.22 kg/m(2); age 44.03 ± 1.87 years (range 41 to 47)]. Indexes of oxidative stress were assayed by a colorimetric method. The relationships between circulating EPCs, serum visfatin, and oxidative stress markers were further analyzed by multiple linear regression analysis. Statistical significance was set at p < 0.05. RESULTS: The obese group showed higher levels of serum visfatin and a lower level of circulating EPCs compared with controls. Serum superoxide dismutase (SOD) activity, total antioxidant capacity (T-AOC), and glutathione peroxidase (GSH-px) activity were significantly lower in obese subjects than in controls, while levels of serum malondialdehyde (MDA) were significantly higher. Circulating EPCs were positively associated with SOD (ß = 0.306) and LnHOMA-IR (ß = 0.223) and negatively associated with BMI (ß = -0.321), serum visfatin (ß = -0.236), and MDA (ß = -0.293). CONCLUSIONS: The quantity of circulating EPCs decreases in obese individuals, along with increased serum visfatin and oxidative stress product. Visfatin and oxidative stress might therefore impact on the circulating EPCs in obese populations.