[Research progress of haematopoietic stem cells to generate immune cells and its application].

Hematopoietic stem cells (HSCs) posses the potential for highly self-renewal, proliferation and multi-lineage differentiation. HSC transplantation has long been the primary method for treating hematologic disorders and autoimmune diseases, and the ability to rebuild the immune system after transplantation is a key indicator of success. To enhance the reconstruction ability of the immune system after transplantation, current research focuses on genetic engineering and the use of HSCs modified by clustered regularly interspaced short palindromic repeats (CRISPR) gene editing technology as a source of transplant cells. This article summaries the biological characteristics, regulatory mechanism, ability to differentiate into immune cells, as well as the application and advance in the treatment of blood disorders, immune deficiencies, cancers and other related diseases, aiming to provide references for the research on relevant diseases.

Effect of estradiol after bacterial infection on the Wnt/ß-catenin pathway in bovine endometrium epithelial cells and organoids.

Endometritis is a disease caused by a postpartum bacterial infection with a poor prognosis that primarily affects dairy cows. Three-dimensional organoids have been used as a model for endometritis, because they exhibit a structure comparable to that of the endometrium, demonstrating both expansibility and hormone responsiveness. These characteristics render them an ideal platform for in vitro investigations of endometrial diseases. Estradiol (E2) is an endogenous steroid hormone with demonstrated anti-inflammatory properties, and the objective of this study was to determine the mechanism by which E2 modulates the inflammatory response and the Wnt signal transduction pathway in bovine endometrial epithelial cells and organoids following E. coli infection. We present the techniques for isolating and culturing primary bovine endometrial epithelial cells (BEECs), and producing endometrial organoids. For the experiments, the endometrial epithelial cells and organoids were infected with E. coli for 1 h, followed by incubation with E2 for 12 h. The mRNA and protein expressions of the inflammation-related genes, IL-1ß, IL-6, TLR4, and NF-κB, as well as the Wnt pathway-related genes, Wnt4, ß-catenin, c-Myc, and CyclinD1, were assessed using real-time quantitative-PCR and western blotting, respectively. The CCK8 viable cell counting assay was utilized to determine the optimal concentration of the Wnt inhibitor, IWR-1. The mRNA and protein expression of Wnt pathway-related genes was assessed following IWR-1 treatment, while the expression levels of proliferation-associated genes (Ki67, PCNA) and barrier repair genes (occludin, claudin, and Zo-1) in BEECs and organoids were evaluated after E2 treatment. The results of this study show that mRNA expression of the inflammatory genes, IL-1ß, TLR4, and NF-κB (P < 0.05) decreased in BEECs following E2 treatment compared to the E. coli group. The protein expression of the IL-1ß, IL-6, TLR4 and NF-κB genes was also inhibited (P < 0.05). Similar results were observed in tests on the organoids. Our findings demonstrate that E2 significantly upregulates the expression of Wnt-related genes, including ß-catenin and c-Myc, while concurrently downregulating the expression of GSK3ß (P < 0.05). Next, we treated E. coli-infected BEECs and organoids with the Wnt inhibitor, IWR-1. Compared with E. coli and E. coli + E2, the expression of mRNA and protein from Wnt 4, ß-catenin, and CyclinD1 in E. coli + E2 and E. coli + IWR-1 was down-regulated (P < 0.05). The expression of the proliferation genes, Ki67, PCNA, and the tight junction genes, occludin, claudin1, and Zo-1, in organoids was significantly higher than that in BEECs (P < 0.05). In summary, we found strong potential for E2 mitigation of the E. coli-induced inflammatory response in BEECs and organoids, through activation of the Wnt pathway. In addition, the proliferation and repair capacity of organoids was much higher than that of BEECs.

The effect of 1,25-dihydroxyvitamin D3 on the Wnt signaling pathway in bovine intestinal epithelial cells is mediated by the DKK2 (dickkopf2) Wnt antagonist.

The Wnt/ß-catenin signaling pathway is aberrantly activated in most colorectal cancers. High-dose 1,25(OH)2D3 has anticancer effect by regulating Wnt signal pathway. However, it is not clear whether high-dose of 1,25(OH)2D3 have an effect on normal cells. The aim of the present study was to investigate the mechanism of high-dose 1,25(OH)2D3 on the Wnt signaling pathway in bovine intestinal epithelial cells. The potential mechanism of action was investigated after knockdown and overexpression of the Wnt pathway inhibitor, DKK2, in intestinal epithelial cells by observing the effects of 1,25(OH)2D3 on proliferation, apoptosis, pluripotency and the expression of genes related to the Wnt/ß-catenin signaling pathway. In the present study, we introduced the method of isolation and culture of primary bovine intestinal epithelial cells. After cells were treated with 50 ng/mL 1,25(OH)2D3 or DMSO for 48 h, total RNA was extracted, and six differentially expressed genes, including SERPINF1, SFRP2, SFRP4, FZD2, WISP1 and DKK2 were identified by transcriptome sequencing, which were related to Wnt signaling pathway. To further explore the mechanism of 1,25(OH)2D3 on the Wnt/ß-catenin signaling pathway, we constructed knockdown and overexpression plasmids of DKK2. After transfecting these plasmids into bovine intestinal epithelial cells, we measured the expression of DKK2 mRNA and protein through GFP expression, qRT-PCR and western blot analyses to verify the transfection efficiency. In addition, the CCK-8 assay was used to detect the cell proliferation rate after transfection. Subsequently, the transfected cells were treated with 1,25(OH)2D3 for 48 h, and the proliferation- (Ki67 and PCNA), apoptosis- (Bcl-2, p53, casp3 and casp8), pluripotency- (Bmi-1, Lrig1, KRT19 and TUFT1) and Wnt/ ß-catenin signaling pathway- related genes (LGR5, DKK2, VDR, ß- Catenin, SFRP2, WISP1 and FZD2) were detected by qRT-PCR and western blot analyses. Our results showed that the expression trend of some genes in bovine intestinal epithelial cells under high-dose 1,25(OH)2D3 was consistent with the sequencing results, including SFRP2 (P < 0.001), SFRP4 (P < 0.05), FZD2 (P < 0.01), WISP1 (P < 0.001) and DKK2 (P < 0.001). In addition, knockdown of DKK2 inhibited cell proliferation (P < 0.01), but DKK2 overexpression promoted cell proliferation (P < 0.01). Compared to the control group, 1,25(OH)2D3 promoted the expression of Wnt/ß-catenin signaling pathway-related proteins in bovine intestinal epithelium, thus maintaining intestinal homeostasis in normal intestinal epithelium. In addition, knockdown and overexpression of DKK2 indicated that 1,25(OH)2D3 weakened the inhibitory effect of DKK2 on the Wnt/ß-catenin signaling pathway. Together, these results suggest that high-dose 1,25(OH)2D3 has no killing effect on normal intestinal epithelial cells and regulates Wnt/ß-catenin signaling pathway through DKK2.

Effect of VD3 on cell proliferation and the Wnt signaling pathway in bovine endometrial epithelial cells treated with lipopolysaccharide.

Vitamin D (VD) deficiency plays an important role in the occurrence and development of various uterine diseases. At present, most studies on the mechanism of VD in the Wnt signaling pathway focus on cancer, while there are no relevant reports on its mechanism in endometritis. This study investigated the effect of vitamin D3 (VD3) on the Wnt signaling pathway in endometrial epithelial cells (BEECs) induced by lipopolysaccharide (LPS). BEECs obtained from bovine uteri were treated with VD3 (0, 50 ng/mL) and LPS (0, 10, 100 ng/mL) separately or in combination, and treated with the Wnt signaling pathway inhibitor IWR-1 to study the mechanism of action. The proliferation of BEECs was evaluated by a CCK-8 assay. qRT-PCR was used to assess the gene expression of Wnt pathway-related factors, including MYC, PCNA, LGR5, GREM1, ß-catenin, FZD7, FZD2, Wnt4 and VDR. The results showed that VD3 had no significant effect on cell proliferation (P > 0.05); LPS inhibited BEEC proliferation in a time- and dose-dependent manner, and cells treated with LPS at different concentrations for 24-48 h in combination with VD3 promoted cell proliferation to varying degrees. IWR-1 inhibited cell proliferation in a time- and concentration-dependent manner, while LPS + IWR-1 treatment also significantly promoted cell proliferation after VD3 treatment (P < 0.01). The qRT-PCR results showed that the expression of Wnt4 and PCNA genes showed different trends with different LPS concentrations for stimulation, and the expression of the MYC and GREM1 genes was only stimulated by high-dose (100 ng/mL) LPS stimulation. The expression of FZD7, LGR5, FZD2 and ß-catenin was upregulated by LPS at both concentrations. LPS + VD3 significantly downregulated the expression of the Wnt pathway-related genes MYC, PCNA, LGR5, GREM1 and ß-catenin (P < 0.001), Wnt4 and FZD2 (P < 0.01), and significantly upregulated the expression of VDR (P < 0.05). After LPS + IWR-1 treatment, the expression of the ß-catenin (P < 0.01) and LGR5 (P < 0.05) genes was significantly downregulated, while the Wnt4 (P < 0.01) and VDR (P < 0.001) genes were significantly upregulated, MYC was downregulated but without a significant difference (P > 0.05). In conclusion, VD3 treatment can mitigate the LPS-induced abnormal expression of Wnt signaling pathway genes in BEECs, showing that the Wnt pathway may be a protective pathway of VD3 against LPS-induced gene overexpression in BEECs. The results suggest that VD3 may play a regulatory role in pathways other than the Wnt signaling pathway. Whether VD3 affects the Wnt signaling pathway by affecting Wnt4 gene expression requires further study.

Animal based diets and environment: Perspective from phosphorus flow quantifications of livestock and poultry raising in China.

Identifying the key nodes of the phosphorus flows in animal raising system is fundamental to improve P utilization efficiency and reduce the P contamination. This study established a phosphorus flow analysis model for livestock and poultry raising, depicted P flows for major livestock and poultry under two raising modes, and further analyzed their spatial and temporal distributions. We find that around 15% of P input was transferred into the products, and in P output around 40% lost into the environment in 2015. The P flows have been increasing since 2000, and the main contributor is pigs followed by beef cattle. It should be noticed that P loss from livestock and poultry raising is huge with extensive prospect of recycling in some central provinces of China, and western region where ecological environment is fragile, has a higher P loss rate which need to change the dietary preference and adjust raising structure. As for diets, pork and eggs are better choices than milk or other kinds of meat in terms of reducing the P load, when producing per unit protein or energy. This study contributes to the understanding of P management in husbandry industry, the quantification of environmental loads of animal-based food and the identification of the potential of reducing P loss to realize sustainable utilization of P.

Tracing anthropogenic cadmium emissions: From sources to pollution.

Cadmium (Cd), a widely concerned heavy metal, is toxic to humans and ecosystems. In this paper, a Cd-polluted town in southeast China was selected to estimate the Cd emissions of human activities into surface water, soil and atmospheric environment. The analysis shows that the total amount of Cd emitted into the environment in 2015 was approximately 43.5 kg, and the majority of those emissions were discharged into the water, accounting for approximately 90.4% of the total Cd emissions. The remaining Cd emissions into the soil and atmosphere accounted for 9.5% and 0.1%, respectively. The industrial production (IP) is the dominant source of anthropogenic Cd emissions, which contributes to 62.1% of the total emissions. The other contributors include aquaculture (AQ), wastewater treatment (WT), living consumption (LC), crop farming (CF) and animal breeding (AB); each accounted for less than 10% of the total emissions. Pigment production is the largest source of IP emissions. According to the results of correlation analysis, the pigment enterprises are responsible for the heavy Cd pollution in local soil. By comparing the spatial position and combing with the local watercourses, the study reveals that the irrigated watercourse is the transmission channels of soil Cd pollution. This study contributes to the analysis of connecting the emission inventory, environmental media and transmission channels of the heavy metal Cd and provides policy supports for the local governments to adopt a life cycle Cd management approach.