摘要：
Highly immunogenic nucleotide fragments from 3 genes of Mycoplasma hyopneumoniae strain 232 were selected using information software technology. After repeating each fragment three times, a total of 9 nucleotide fragments were joined together to form a new nucleotide sequence called Mhp2321092bp. Mhp2321092bp was directly synthesized and cloned into a pET100 vector and expressed in Escherichia coli. After purification, the proteins were successfully validated by SDS-PAGE and Western blotting using mouse His-tag antibody and pig anti-Mhp serum. BALB/c mice were intraperitoneally injected with purified proteins in the high-dose (100 & mu;g), medium-dose group (50 & mu;g) and low-dose (10 & mu;g) groups. Mice in each group were injected on day 1, day 8 and day 15 of feeding, respectively. Serum samples were collected from all mice on the day before immunization and on day 22 after immunization. The antibody level in the mouse serum was detected using western blotting using purified expressed proteins as antigens. IL-2, TNF-& alpha; and IFN-& gamma; were simultaneously detected in the mouse serum by ELISA. The results showed that the 60 kDa protein was successfully expressed and reacted specifically with the specific serum Mhp His-Tag mouse monoclonal antibody and pig anti-Mhp serum. From day 0 to day 22 of immunization, IFN-& gamma; increased from 269.52 to 467.74 pg/mL, IL-2 increased from 14.03 to 145.16 pg/mL, and TNF-& alpha; increased from 6.86 to 12.37 pg/mL. The IgG antibody in mice increased significantly from 0 day to day 22 after immunization. This study suggests that the expressed recombinant protein may serve as one of the novel vaccine candidates for Mhp.
摘要：
Resveratrol (Res) is a bioactive dietary component and alleviates apoptosis in multiple cell types. However, its effect and mechanism on lipopolysaccharide (LPS)-induced bovine mammary epithelial cells (BMEC) apoptosis, which commonly happens in dairy cows with mastitis, is unknown. We hypothesized that Res would inhibit LPS-induced apoptosis in BMEC through SIRT3, a NAD + -dependent deacetylase activated by Res. To test the dose-response effect on apoptosis, 0-50 mu M Res were incubated with BMEC for 12 h, followed by 250 mu g/mL LPS treatment for 12 h. To investigate the role of SIRT3 in Res-mediated alleviation of apoptosis, BMEC were pretreated with 50 mu M Res for 12 h, then incubated with si-SIRT3 for 12 h and were finally treated with 250 mu g/mL LPS for 12 h. Res dose-dependently promoted the cell viability and protein levels of Bcl-2 (Linear P < 0.001) but decreased protein levels of Bax, Caspase-3 and Bax/Bcl-2 (Linear P < 0.001). TUNEL assays indicated that cellular fluorescence intensity declined with the rising doses of Res. Res also dose-dependently upregulated SIRT3 expression, but LPS had the opposite effect. SIRT3 silencing abolished these results with Res incubation. Mechanically, Res enhanced the nuclear translocation of PGC1 alpha, the transcriptional cofactor for SIRT3. Further molecular docking analysis revealed that Res could directly bind to PGC1 alpha by forming a hydrogen bond with Tyr-722. Overall, our data suggested that Res relieved LPS-induced BMEC apoptosis through the PGC1 alpha-SIRT3 axis, providing a basis for further in vivo investigations of applying Res to relieve mastitis in dairy cows.
摘要：
The present study was conducted to evaluate the regulatory actions and underlying mechanisms of butyrate on the inflammatory response and tight junction (TJ) disruption in bovine mammary epithelial cells (BMECs). Results showed that butyrate declined histone deacetylase 3 (HDAC3) expression, blocked NF-kappa B activation, and thus suppressed inflammatory cytokine production in gamma-d-glutamyl-meso-diaminopimelic acid (iE-DAP)-triggered BMECs. Butyrate also depressed the protein abundance of myosin light chain kinase (MLCK), elevated the expression of TJ proteins, and restored the cellular distribution of TJ proteins and the barrier function of epithelial cells. HDAC3 overexpression abolished the protective effects of butyrate. In conclusion, butyrate alleviated the iE-DAP-induced inflammatory response and TJ injury by blocking NF-kappa B activation and decreasing inflammatory cytokine production and MLCK expression in a HDAC3-dependent manner. Our finding provides a mechanistic basis for further exploring the regulatory effects of butyrate on the mammary inflammatory response.
摘要：
本论文采用了网络药理学的方法,以原阿片碱(Protopine)为研究对象,探究其在鸡卵巢组织中的作用靶点。通过反向分子对接网站SwissTargetPrediction筛选原阿片碱对应的靶点,通过检索NCBI和GeneC...展开更多 本论文采用了网络药理学的方法,以原阿片碱(Protopine)为研究对象,探究其在鸡卵巢组织中的作用靶点。通过反向分子对接网站SwissTargetPrediction筛选原阿片碱对应的靶点,通过检索NCBI和GeneCards数据库收集与鸡卵巢组织炎症相关的靶点,进而得到原阿片碱治疗家禽卵巢炎症的关键靶点;将获得的关键靶点导入STRING数据库进行分析,并构建蛋白质-蛋白质相互作用(protein-protein interaction,PPI)网络,通过Cytoscape软件可视化;通过STRING数据库对关键靶点进行基因本体(gene ontology,GO)功能及京都基因与基因组百科全书(Kyoto encyclopedia of genes and genomes,KEGG)通路富集分析。共筛选得到原阿片碱的42个相关的靶点,与2489个鸡卵巢组织相关靶点取交集,获得17个交集靶点。PPI网络分析显示,MAPK8、CHEK1、PIK3CA、PTGS2、JAK1、JAK3、JAK2、PIK3CB、PIK3CD、CCNE1、CDK2、APP、TYMS和CYP19A1这些基因可能为原阿片碱治疗鸡卵巢炎症的关键靶点;KEGG通路富集分析得到炎症途径和凋亡相关途径信号通路。通过基因数据库的分析,预测出了可能与原阿片碱相关的基因、蛋白质、信号通路等,对于原阿片碱产品的临床的使用有较好的指导意义。收起
摘要：
The outbreak of coronavirus disease 2019 (COVID-19) caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has shaken the global health system. Various nanotechnology-based strategies for vaccine development have played pivotal roles in fighting against SARS-CoV-2. Among them, the safe and effective protein-based nanoparticle (NP) platforms display a highly repetitive array of foreign antigens on their surface, which is urgent for improving the immunogenicity of vaccines. These platforms greatly improved antigen uptake by antigen presenting cells (APCs), lymph node trafficking, and B cell activation, due to the optimal size, multivalence, and versatility of NPs. In this review, we summarize the advances of protein-based NP platforms, strategies of antigen attachment, and the current progress of clinical and preclinical trials in the development of SARS-CoV-2 vaccines based on protein-based NP platforms. Importantly, the lessons learnt and design approaches developed for these NP platforms against SARS-CoV-2 also provide insights into the development of protein-based NP strategies for preventing other epidemic diseases.
摘要：
In goats, most follicles in the ovaries will be atresia and only a few dominant follicles (DFs) may eventually mature and ovulate at a follicular wave. To investigate the potential microRNAs (miRNAs) that regulate the expression of genes associated with follicular dominance or atresia, small RNA sequencing was performed on granulosa cells of DF and subordinate follicle at the first follicular wave in goats. A total of 108 differentially expressed miRNAs were detected in the two types of follicle granulosa cells: 16 upregulated miRNAs and 92 downregulated miRNAs. Kyoto Encyclopedia of Genes and Genomes analysis of the target genes showed that TKTL1, LOC102187810, LOC102184409 and ALDOA are closely associated with follicle dominance and are involved in the pentose phosphate pathway. Furthermore, a coexpression network of miRNAs and follicular dominance-related genes was constructed. The qPCR results well correlated with the small RNA sequencing data. Our findings provide new insight for exploring the molecular mechanism of miRNAs in regulating follicular development in goats.
摘要：
Ketosis is often accompanied by a reduction in milk production in dairy cows, but the molecular mechanism has not been fully elucidated. Ketotic cows possess systemic oxidative stress (OS), which may implicate apoptosis in mammary glands. Sirtuin 3 (SIRT3) is a vital regulator of cellular redox homeostasis and is under the control of AMP-activated protein kinase (AMPK) signaling in nonruminants. Thus, we aimed to investigate (1) the AMPK-SIRT3 and apoptosis status of mammary glands from ketotic cows, (2) the effect of SIRT3 on OS-induced apoptosis in bovine mammary epithelial cells (BMEC), and (3) the role of AMPK signaling on SIRT3-mediated effects on apoptosis. Mammary gland samples were reused from a previous study, which contained healthy and ketotic cows (both n = 15). BMEC were incubated with 0, 0.3, 0.6, or 0.9 mM H(2)O(2) for 6 h with/without a 30 min incubation of an antioxidant MitoQ (1 μM). Then BMEC were incubated with SIRT3 overexpression adenovirus (Ad-SIRT3) for 6 h followed by a 6 h incubation with 0.6 mM H(2)O(2). Finally, BMEC were treated with the AMPK inhibitor Compound C (Cd C,10 μM) for 30 min before the H(2)O(2) challenge, or cells were initially treated with the AMPK agonist MK8722 (10 μM) for 30 min followed by a 30-h culture with/without si-SIRT3 and eventually the H(2)O(2) exposure. Ketotic cows displayed higher levels of Bax, Caspase-3 and Bax/Bcl-2 but lower levels of Bcl-2 in mammary glands. H(2)O(2) incubation displayed similar results, exhibiting a dose-dependent manner between the H(2)O(2) concentration and the apoptosis degree. Mito Q pretreatment reduced cellular reactive oxygen species and rescued cells from apoptosis. Ketotic cows had a lower mammary protein abundance of SIRT3. Similarly, H(2)O(2) incubation downregulated both mRNA and protein levels of SIRT3 in a dose- and time-dependent manner. Ad-SIRT3 infection lowered levels of cellular reactive oxygen species, Bax, Caspase-3 and Bax/Bcl-2 but increased levels of Bcl-2. TUNEL assays confirmed that Ad-SIRT3 infection mitigated H(2)O(2)-induced apoptosis. Both ketotic cows and H(2)O(2)-induced BMEC had lower levels of p-AMPK and p-AMPK/AMPK. Additionally, Cd C pretreatment decreased SIRT3 and Bcl-2 expression but increased levels of Bax and Caspase-3. Contrary to the inhibitor, MK8722 had opposite effects and reduced the percentage of apoptotic cells. However, these effects of MK8722 were reversed upon SIRT3 silencing. In conclusion, in vivo data confirmed that ketosis is associated with greater apoptosis and restricted AMPK-SIRT3 signaling in mammary glands; in vitro data indicated that SIRT3 mitigates OS-induced apoptosis via AMPK signaling. As such, there may be potential benefits for targeting the AMPK-SIRT3 axis to help counteract the negative effects of mammary glands during ketosis.
通讯机构：
[Shouyu Wang] J;Joint International Research Laboratory of Animal Health and Food Safety of Ministry of Education & Single Molecule Nanobiology Laboratory (Sinmolab), Nanjing Agricultural University, Nanjing, Jiangsu 210095, China<&wdkj&>OptiX+ Laboratory, School of Electronics and Information Engineering, Wuxi University, Wuxi, Jiangsu 214105, China
关键词：
PEDV detection;FRET immunoassay;Rapid on-site detection;FRET immunoassay station (FRETIS)
摘要：
The porcine epidemic diarrhea virus (PEDV) is a major pathogen of swine enteric diseases. Various etiology and serological methods have been employed for PEDV detection, but most of their applications are limited to lab-oratories. To extend PEDV detection to on-site applications, we design a homogeneous fluorescence resonance energy transfer (FRET)-based enzyme-linked immunosorbent assay (ELISA). Both donor and acceptor fluores-cence microspheres modified PEDV antibodies can be linked only to the occurrence of PEDV antigen, thus generating FRET signals, which can be collected by our designed portable FRET immunoassay station (FRETIS). Verified by standard samples, FRET immunoassay reached high sensitivity with a detection limit as TCID50 (median tissue culture infective dose) of 10/mL, which is 10 times more sensitive than colloidal gold test strips; and verified by clinical samples, it was also proved with high accuracy, good selectivity, and repeatability. More importantly, FRET immunoassay could detect PEDV in a 96-well plate in 35 min with only one step of incubation without any further washing steps using field-portable devices and field-operable procedures, well supporting on -site applications. Considering these advantages, this reported FRET immunoassay provides a promising way for multi-sample on-site PEDV detection and can be potentially used in the swine industry.
作者机构：
[Xie, Shi-Chen; Mei, Jin-Jin; Xie, SC; Zheng, Wen-Bin; Liu, Ya-Ya; Liu, Qing; Zhu, Xing-Quan; Liang, Yao; Gao, Wen-Wei] Shanxi Agr Univ, Coll Vet Med, Lab Parasit Dis, Jinzhong 030801, Peoples R China.;[Xie, Shi-Chen; Xie, SC; Zhu, Xing-Quan] Hunan Agr Univ, Coll Vet Med, Res Ctr Parasites & Vectors, Changsha 410128, Peoples R China.;[Zhu, Xing-Quan] Yunnan Agr Univ, Coll Vet Med, Key Lab Vet Publ Hlth Higher Educ Yunnan Prov, Kunming 650201, Peoples R China.
通讯机构：
[Xie, SC ; Zhu, XQ] S;[Zhu, XQ ] Y;Shanxi Agr Univ, Coll Vet Med, Lab Parasit Dis, Jinzhong 030801, Peoples R China.;Hunan Agr Univ, Coll Vet Med, Res Ctr Parasites & Vectors, Changsha 410128, Peoples R China.;Yunnan Agr Univ, Coll Vet Med, Key Lab Vet Publ Hlth Higher Educ Yunnan Prov, Kunming 650201, Peoples R China.
关键词：
Cryptosporidium spp.;Blastocystis sp.;genotypes;subtypes;prevalence;cattle;Shanxi Province
摘要：
Both Cryptosporidium spp. and Blastocystis sp. are common intestinal protozoa, which can cause zoonotic diseases and economic losses to livestock industry. To evaluate the prevalence and genetic population structure of Cryptosporidium spp. and Blastocystis sp. in beef and dairy cattle in Shanxi Province, north China, a total of 795 fecal samples were collected from beef and dairy cattle in three representative counties in Shanxi Province, and these fecal samples were examined using molecular approaches based on 18S small-subunit ribosomal RNA (SSU rRNA) of Cryptosporidium spp. and Blastocystis sp., respectively. Among 795 cattle fecal samples, 23 were detected as Cryptosporidium-positive and 103 were detected as Blastocystis-positive, and the overall prevalence of Cryptosporidium spp. and Blastocystis sp. in cattle in Shanxi Province was 2.9% and 13.0%, respectively. For Cryptosporidium spp., DNA sequence analysis indicated that all 23 positive samples were identified as C. andersoni. Furthermore, five known subtypes (ST1, ST10, ST14, ST21 and ST26) and three unknown subtypes of Blastocystis sp. were detected among 103 positive samples using DNA sequence analysis. This study reported the occurrence and prevalence of Cryptosporidium spp. and Blastocystis sp. in cattle in Shanxi Province for the first time, which extends the geographical distribution of these two zoonotic parasites and provides baseline data for the prevention and control of these two important zoonotic parasites in cattle in Shanxi Province.
摘要：
The T-2 toxin (T2) poses a major threat to the health and productivity of animals. The present study aimed to investigate the regulatory mechanism of Nrf2 derived from broilers against T2-induced oxidative damage. DF-1 cells, including those with normal characteristics, as well as those overexpressing or with a knockout of specific components, were exposed to a 24 h treatment of 50 nM T2. The primary objective was to evaluate the indicators associated with oxidative stress and the expression of downstream antioxidant factors regulated by the Nrf2-ARE signaling pathway, at both the mRNA and protein levels. The findings of this study demonstrated a noteworthy relationship between the up-regulation of the Nrf2 protein and a considerable reduction in the oxidative stress levels within DF-1 cells (p < 0.05). Furthermore, this up-regulation was associated with a notable increase in the mRNA and protein levels of antioxidant factors downstream of the Nrf2-ARE signaling pathway (p < 0.05). Conversely, the down-regulation of the Nrf2 protein was linked to a marked elevation in oxidative stress levels in DF-1 cells (p < 0.05). Additionally, this down-regulation resulted in a significant decrease in both the mRNA and protein expression of antioxidant factors (p < 0.05). This experiment lays a theoretical foundation for investigating the detrimental impacts of T2 on broiler chickens. It also establishes a research framework for employing the Nrf2 protein in broiler chicken production and breeding. Moreover, it introduces novel insights for the prospective management of oxidative stress-related ailments in the livestock and poultry industry.
通讯机构：
[Zeng, JG; Yang, ZH ] H;Hunan Agr Univ, Hunan Key Lab Tradit Chinese Vet Med, Changsha 410128, Peoples R China.;Hunan Agr Univ, Coll Vet Med, Changsha 410128, Peoples R China.
关键词：
sanguinarine;UPLC-Q-TOF-MS/MS;metabolites;rats;in vivo
摘要：
Sanguinarine (SAN), as the main active component of a traditional Chinese veterinary medicine, has been widely used in the animal husbandry and breeding industry. However, the metabolites of SA are still uncertain. Therefore, this research aimed to investigate the metabolites of SA based on rats in vivo. The blood, feces, and urine of rats were collected after the oral administration of 40 mg/kg SAN. Ultra-high-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS/MS) was employed to identify the metabolites of SAN. The elemental composition of sanguinarine metabolites was inferred by analyzing their exact molecular weight, and the structures of the metabolites were predicted based on their fragment ions and cleavage pathways. A total of 12 metabolites were identified, including three metabolites in the plasma, four in the urine, and nine in the feces. According to the possible metabolic pathways deduced in this study, SAN was mainly metabolized through reduction, oxidation, demethylation, hydroxylation, and glucuronidation. This present research has summarized the metabolism of SAN in rats, which is helpful for further studying the metabolic mechanism of SAN in vivo and in vitro.
