Multi-omics reveals that alkaline mineral water improves the respiratory health and growth performance of transported calves.

BACKGROUND: Long-distance transportation, a frequent practice in the cattle industry, stresses calves and results in morbidity, mortality, and growth suppression, leading to welfare concerns and economic losses. Alkaline mineral water (AMW) is an electrolyte additive containing multiple mineral elements and shows stress-mitigating effects on humans and bovines. RESULTS: Here, we monitored the respiratory health status and growth performance of 60 Simmental calves subjected to 30 hours of road transportation using a clinical scoring system. Within the three days of commingling before the transportation and 30 days after the transportation, calves in the AMW group (n = 30) were supplied with AMW, while calves in the Control group (n = 29) were not. On three specific days, namely the day before transportation (day -3), the 30th day (day 30), and the 60th day (day 60) after transportation, sets of venous blood, serum, and nasopharyngeal swab samples were collected from 20 calves (10 from each group) for routine blood testing, whole blood transcriptomic sequencing, serology detection, serum untargeted metabolic sequencing, and 16S rRNA gene sequencing. The field data showed that calves in the AMW group displayed lower rectal temperatures (38.967 ℃ vs. 39.022 ℃; p = 0.004), respiratory scores (0.079 vs. 0.144; p < 0.001), appetite scores (0.024 vs. 0.055; p < 0.001), ocular and ear scores (0.185 vs. 0.338; p < 0.001), nasal discharge scores (0.143 vs. 0.241; p < 0.001), and higher body weight gains (30.870 kg vs. 7.552 kg; p < 0.001). The outcomes of laboratory and high throughput sequencing data revealed that the calves in the AMW group demonstrated higher cellular and humoral immunities, antioxidant capacities, lower inflammatory levels, and intestinal absorption and lipogenesis on days -3 and 60. The nasopharynx 16S rRNA gene microbiome analysis revealed the different composition and structure of the nasopharyngeal microflora in the two groups of calves on day 30. Joint analysis of multi-omics revealed that on days -3 and 30, bile secretion was a shared pathway enriched by differentially expressed genes and metabolites, and there were strong correlations between the differentially expressed metabolites and the main genera in the nasopharynx. CONCLUSIONS: These results suggest that AMW supplementation enhances peripheral immunity, nutrition absorption, and metabolic processes, subsequently affecting the nasopharyngeal microbiota and improving the respiratory health and growth performance of transported calves. This investigation provided a practical approach to mitigate transportation stress and explored its underlying mechanisms, which are beneficial for the development of the livestock industry. Video Abstract.

Antioxidant Capacity and Protective Effect of Cow Placenta Extract on D-Galactose-Induced Skin Aging in Mice.

Placental extract has been used for skin care and delaying skin aging. Cow placenta is an abundant resource with a large mass, which has not been harnessed effectively. Cow placenta extract (CPE) has the functions of antioxidation, anti-inflammatory, promoting growth and development, and promoting hair growth. However, little is known about the effect of oral administration of cow placenta extract on skin conditions. Therefore, the present study aimed to investigate the antioxidant capacity of CPE in vitro and in vivo and its protective effect on d-galactose (D-gal) induced skin aging in mice. The results showed that CPE had strong free radical scavenging, reducing and metal chelating activities. CPE can increase the activity of catalase (CAT), glutathione peroxidase (GSH-Px), peroxidase (POD), superoxide dismutase (SOD), and the content of glutathione (GSH), decrease the content of malondialdehyde (MDA). Moreover, CPE can decrease the gene and protein expression of matrix metalloproteinase 1a (MMP-1a) and matrix metalloproteinase 3 (MMP-3) and increase the expression of transforming growth factor-ß (TGF-ß) and tissue inhibitor of metalloproteinase 1 (TIMP-1) of mouse skin. Histopathological analysis showed CPE reduced the collagen damage caused by D-gal, increased collagen synthesis and reduced its degradation to delay skin aging.

Exploration of PDCoV-induced apoptosis through mitochondrial dynamics imbalance and the antagonistic effect of SeNPs.

Porcine Deltacoronavirus (PDCoV), an enveloped positive-strand RNA virus that causes respiratory and gastrointestinal diseases, is widely spread worldwide, but there is no effective drug or vaccine against it. This study investigated the optimal Selenium Nano-Particles (SeNPs) addition concentration (2 - 10 µg/mL) and the mechanism of PDCoV effect on ST (Swine Testis) cell apoptosis, the antagonistic effect of SeNPs on PDCoV. The results indicated that 4 µg/mL SeNPs significantly decreased PDCoV replication on ST cells. SeNPs relieved PDCoV-induced mitochondrial division and antagonized PDCoV-induced apoptosis via decreasing Cyt C release and Caspase 9 and Caspase 3 activation. The above results provided an idea and experimental basis associated with anti-PDCoV drug development and clinical use.

Research progress on diarrhoea and its mechanism in weaned piglets fed a high-protein diet.

In order to pursue faster growth and development of weaned piglets, increased dietary protein (CP) levels were favoured by the pig industry and the feed industry. The digestive organs of piglets were not fully developed at weaning, and the digestive absorption capacity of protein was limited. High-protein diets can cause allergic reactions in piglets, destroy intestinal structural integrity, reduce immunity, and cause intestinal flora imbalance. Undigested proteins were prone to produce toxic substances, such as ammonia and biogenic amines, after fermentation in the hindgut, which negatively affects the health of the intestine and eventually causes reduced growth performance and diarrhoea in piglets. This review revealed the mechanism of diarrhoea caused by high-protein diets in weaned piglets and provided ideas for preventing diarrhoea in weaned piglets.

Effect of Selenium on Brain Injury in Chickens with Subacute Arsenic Poisoning.

The aim of this study was to investigate the effects of different doses of selenium (Se) on oxidative damage and neurotransmitter-related parameters in arsenic (As)-induced broiler brain tissue damage. Two hundred 1-day-old avian broilers were randomly divided into five groups and fed the following diets: control group (As 0.1 mg/kg + Se 0.2 mg/kg), As group (As 3 mg/kg + Se 0.2 mg/kg), low-Se group (As 3 mg/kg + Se 5 mg/kg), medium-Se group (As 3 mg/kg + Se 10 mg/kg), and high-Se group (As 3 mg/kg + Se 15 mg/kg). Glutathione (GSH), glutathione peroxidase (GSH-PX), nitric oxide (NO), nitric oxide synthase (NOS) activity, glutamate (Glu) concentration, glutamine synthetase (GS) activity, acetylcholinesterase (TchE) activity, and the apoptosis rate of brain cells were measured. The results showed that 3 mg/kg dietary As could induce oxidative damage and neurotransmitter disorder of brain tissue, increase the apoptosis rate of brain cells and cause damage to brain tissue, decrease activities of GSH and GSH-PX, decrease the contents of NO, decrease the activities of iNOS and tNOS, increase contents of Glu, and decrease activities of Gs and TchE. Compared with the As group, the Se addition of the low-Se and medium-Se groups protected against As-induced oxidative damage, neurotransmitter disorders, and the apoptosis rate of brain cells, with the addition of 10 mg/kg Se having the best effect. However, 15 mg/kg Se not only did not produce a protective effect against As damage but actually caused similar or severe damage.

Assessment of antiviral activity and mechanism of rhein on newcastle disease virus (La Sota strain IV) in vitro.

Current research is focused on the development of drug candidates from natural products. Rhein a Traditional Chinese Medicine (TCM) from Polygonaceae (rhubarb) has exhibited antioxidant, anti-inflammatory and anticancer activities, however no work has reported its antiviral potential, thus this study was performed to investigate the antiviral activities of rhein on new castle disease virus (NDV) in vitro.NDV infection of chicken embryo fibroblasts (CEFs) was prepared using 10-day-old specific pathogen free chicken embryos. Cytotoxicity and anti-viral activities of rhein were assessed using the MTT method. The interaction between NDV and cell membrane proteins were also detected using virus overlay protein binding assay (VOPBA). In addition NDV genes expressions in CEFs were measured using real-time fluorescent quantitative (RTFQ) PCR.The results showed that rhein effectively inhibit NDV activities maximal safe concentration of 0.125 mg/ml. This finding indicated that, rhein could be used as future antiviral drug against NDV.[Formula: see text].

Effects of Selenium on the Immunotoxicity of Subacute Arsenic Poisoning in Chickens.

This study aimed to determine the effects of selenium on the immune toxicity of subacute arsenic poisoning in chickens. Two hundred 8-day-old broilers were randomly divided into 5 groups: the control group (0.1 mg/kg As + 0.2 mg/kg Se), the As group (3 mg/kg As + 0.2 mg/kg Se), As + Se group I (3 mg/kg As + 5 mg/kg Se), As + Se group II (3 mg/kg As + 10 mg/kg Se), and As + Se group III (3 mg/kg As + 15 mg/kg Se). The conclusions were drawn based on the following measurements: 3.0 mg/kg added to feed led to a decrease in the growth performance of the broilers, reduced the level and conversion rate of ANAE, reduced the blood protein content of the broilers but had no effect on the albumin/globulin ratio, and had an inhibitory effect on erythrocyte immunity. Selenium-added of 5 and 10 mg/kg in daily feed leads to increased growth performance, increases the positive rate and conversion rate of ANAE, increases the hemoglobin content of broilers, and promotes erythrocyte immunity, which indicates that the selenium-added reduces the toxic effects of arsenic; 3.0 mg/kg arsenic with 15 mg/kg selenium had the most severe toxic effects. Fifteen milligrams per kilogram of selenium added in daily feed increases the toxicity of arsenic to broilers. The dose of 10 mg/kg selenium showed the best inhibitory effect on subacute arsenic poisoning in the broilers.

Effects of Selenium on Arsenic-Induced Liver Lesions in Broilers.

The aim of the present study was to investigate the abilities of selenium to counteract the toxic damage of arsenic (As). Two hundred 1-day-old healthy male broilers were randomly divided into five groups and fed the following diets: control group (0.1 mg/kg As + 0.2 mg/kg Se), As group (3 mg/kg As + 0.2 mg/kg Se), As + Se group I (3 mg/kg As + 5 mg/kg Se), As + Se group II (3 mg/kg As + 10 mg/kg Se), and As + Se group III (3 mg/kg As + 15 mg/kg Se), respectively. The relative weight of the liver, hepatic protein content, GSH-Px levels, SOD activities, NO contents, iNOS and tNOS activities, and increased malondialdehyde contents, ALT and AST activities, and the apoptotic hepatocytes were analyzed. Adding 3 mg/kg arsenic to the diet caused the growth and development of chicken liver to be blocked, resulting in decrease of protein contents in liver tissue, decrease of SOD and GSH-Px activities, increase of MDA contents, decrease of NO contents, decrease of iNOS and TNOs activities, increase of ALT and AST activities, increase of apoptosis rates of liver cells. Compared to the 3-mg/kg arsenic group, adding 5 mg/kg and 10 mg/kg selenium, respectively, could repair the liver growth retardation and steatosis caused by arsenic, increase the protein contents in liver tissue, increase the activities of SOD and GSH-Px, reduce the contents of MDA, increase the contents of NO, enhance the activities of iNOS and TNOs, reduce the activities of ALT and AST, and reduce the rates of apoptosis of liver cells, in which the best effects are to add 10 mg/kg selenium. While 15 mg/kg of sodium selenite may induce progression of As-induced hepatic lesions, the results indicated that 5 and 10 mg/kg of sodium selenite supplied in the diet, through mechanisms of oxidative stress and apoptosis regulation, may ameliorate As-induced hepatic lesions in a dose-dependent manner.

Comparative iTRAQ Proteomics Reveals Multiple Effects of Selenium Yeast on Dairy Cows in Parturition.

The effects of prepartum dietary supplementation with selenium yeast on low abundant plasma proteins in postpartum dairy cows are not known. In this study, 24 healthy parturient dairy cows were divided into two groups (group C, a control group, and group T, a selenium treatment group). Low abundance proteins were extracted from plasma samples of calving cows, and 542 proteins were identified by isobaric tags for relative and absolute quantitation (iTRAQ) proteomic analysis. Dietary supplementation with selenium yeast caused differential abundance of 48 proteins with a fold change of more than 1.2 or less than 0.83 (p < 0.05); 14 proteins were upregulated and 34 were downregulated. The top five gene ontology (GO) enrichment terms for the differentially expressed proteins were protein homotetramerization (or tetramerization), defense response to bacteria or fungus, acute-phase reactions, nucleotide catabolic process, and positive regulation of lipid metabolic process. All proteins involved in acute-phase reactions were downregulated, indicating that selenium ameliorates systemic inflammation. The vast majority of proteins involved in the defense response to microorganisms were downregulated, thereby affecting innate immunity. The decreased abundance of apolipoprotein A-I and apolipoprotein C-II, critical proteins for positive regulation of lipid metabolism, indicated that selenium may optimize lipid metabolism. The iTRAQ results showed that prenatal supplementation with yeast selenium can relieve systemic inflammation after parturition. Moreover, selenium may reduce the effects of metabolic diseases, which can improve glyconeogenesis and prevent ketosis and fatty liver.

Toxicity of DON on GPx1-Overexpressed or Knockdown Porcine Splenic Lymphocytes In Vitro and Protective Effects of Sodium Selenite.

Deoxynivalenol (DON) is a common contaminant of grain worldwide and is often detected in the human diet and animal feed. Selenium is an essential trace element in animals. It has many biological functions. The role of selenium in the body is mainly orchestrated by selenoprotein. Glutathione peroxidase (GPx) also exists widely in the body and has attracted much attention due to its high antioxidant capacity. In order to explore the effect of the GPx1 gene on toxicity of DON, in this study, we overexpressed or knockdown GPx1 in porcine splenic lymphocytes, then added different concentrations of DON (0.1025, 0.205, 0.41, and 0.82 µg/mL) and sodium selenite (2 µmol/L) to the culture system. Using various techniques, we detected antioxidant function, free radical content, cell apoptosis, and methylation-related gene expression to explore the effect of GPx1 expression on DON-induced cell damage. We also explored whether selenium can antagonize the toxicity of DON in these two cell models and revealed the protective effect of sodium selenite on DON-induced cell damage in GPx1-overexpressing or knockdown splenic lymphocytes. Finally, our findings revealed the following: (1) GPx1 can regulate the antioxidant capacity, apoptosis rate, and expression of DNA methylation-related genes in pig splenic lymphocytes. (2) Na2SeO3 (2 µmol/L) can regulate the antioxidant capacity, apoptosis rate, and expression of DNA methylation-related genes in pig splenic lymphocytes, and this effect is more significant in GPx1-overexpressing cells than in GPx1-knockdown cells. (3) DON can cause oxidative damage, apoptosis, and methylation injury in GPx1-overexpressing or knockdown pig splenic lymphocytes in a concentration-dependent manner. (4) Na2SeO3 (2 µmol/L) can antagonize the toxic effect of DON on GPx1-overexpressing or knockdown pig splenic lymphocytes. Our findings may have important implications for food/feed safety, human health, and environmental protection.

Research Progress on the Toxic Antagonism of Selenium Against Mycotoxins.

Animal feed is prone to becoming infected with molds during production and storage, resulting in secondary metabolite mycotoxins, such as aflatoxin B1 (AFB1), T-2 toxins, deoxynivalenol (DON), and ochratoxin A (OTA), which are harmful to humans and animals. Selenium is an essential trace element for humans and animals, and it is also an effective antioxidant. Many studies have shown that selenium can reduce the damage caused by mycotoxins in animals. This article reviews the current literature on the antagonistic effects of selenium on AFB1, T-2, DON, and OTA toxicity.

Protective Role of Selenium in Immune-Relevant Cytokine and Immunoglobulin Production by Piglet Splenic Lymphocytes Exposed to Deoxynivalenol.

Deoxynivalenol (DON) is a mycotoxin that causes immunosuppression, especially in swine. Selenium (Se) is essential for proper functioning of the immune system in animals. However, little is known about the effects of DON and Se on cytokine or immunoglobulin production in piglets. Here, we addressed this gap by examining piglet splenic lymphocyte responses in vitro. Cells were stimulated with concanavalin A, a T cell stimulatory lectin, in the absence or presence of DON (0.1, 0.2, 0.4, and 0.8 µg/mL), Se (Na2SeO3, 2 µM), or combinations of Se 2 µM and DON 0.1-0.8 µg/mL for 12, 24, or 48 h. At each time point, supernatants and cells were collected and the expression of cytokine and immunoglobulin protein and mRNA was examined. Compared with control and Se-alone treatments, DON exposure significantly and dose dependently decreased the expression levels of IL-2, IL-4, IL-6, IL-10, IFN-γ, IgG, and IgM mRNA and protein. By contrast, co-treatment with DON + Se significantly increased the mRNA and protein levels of all factors examined, except IL-4 and IL-6, compared with DON treatment alone. The results of this investigation demonstrate that Se has the potential to counteract DON-induced immunosuppression in piglets and is a promising treatment for DON-mediated toxicity.

The Molecular Mechanisms of Protective Role of Se on the G2/M Phase Arrest of Jejunum Caused by AFB1.

Aflatoxin B1 (AFB1) is the most toxic among the mycotoxins and causes detrimental health effects on human and animals. Selenium (Se) plays an important role in chemopreventive, antioxidant, anticarcinogen, and detoxification and involved in cell cycle regulation. The aim of this study was to explore the molecular mechanisms of selenium involved in inhibition of G2/M cell cycle arrest of broiler's jejunum. A total of 240 one-day-old healthy Cobb broilers were randomly divided into four groups and fed with basal diet (control group), 0.6 mg/kg AFB1 (AFB1 group), 0.4 mg/kg Se (+Se group), and 0.6 mg/kg AFB1 + 0.4 mg/kg Se (AFB1 + Se group) for 21 days, respectively. The histological observation and morphological analysis revealed that 0.4 mg/kg Se prevented the AFB1-associated lesions of jejunum including the shedding of the apical region of villi, the decreased villus height, and villus height/crypt ratio. The cell cycle analysis by flow cytometry showed that 0.4 mg/kg Se ameliorated the AFB1-induced G2/M phase arrest in jejunal cells. Moreover, the expressions of ATM, Chk2, p53, Mdm2, p21, PCNA, Cdc25, cyclin B, and Cdc2 analyzed by immunohistochemistry and qRT-PCR demonstrated that 0.4 mg/kg Se restored these parameters to be close to those in the control group. In conclusion, Se promoted cell cycle recovery from the AFB1-induced G2/M phase arrest by the molecular regulation of ATM pathway in the jejunum of broilers. The outcomes from the present study may lead to a better understanding of the nature of selenium's essentiality and its protective roles against AFB1.

Use of antimicrobial peptides as a feed additive for juvenile goats.

Although antimicrobial peptides (AMPs) have been used as feed additives, only a few studies have examined their use in ruminants. In this study, we evaluated the use of AMPs(recombinant swine defensin and a fly antibacterial peptide were mixed by 1:1) as a medicated feed additive for juvenile goats. Dietary treatments included control groups (group I: 300 g concentrate; group III: 600 g concentrate), and AMP-supplemented groups (group II: 300 g concentrate + 3.0 g AMPs; group IV: 600 g concentrate + 3.0 g AMPs). AMP-treated groups exhibited an increase in bacterial genera, including Fibrobacter, Anaerovibrio, and Succiniclasticum, and the ciliate genus Ophryoscolex; as well a reduction in bacterial genera, such as Selenomonas, Succinivibrio, and Treponema, and the ciliate genera Polyplastron, Entodinium, and Isotricha. The changes in Fibrobacter, Anaerovibrio, Ophryoscolex, Polyplastron, Entodinium, and Isotricha were related to the concentrate. AMP treatment led to increased body weight, average daily weight gain, enzymatic activity (pectinase, xylanase, and lipase), especially in the normal concentrate group, and influence on ruminal fermentation function. In addition, goats treated with AMPs had higher rumen microorganism diversity indices than the control groups. Our results demonstrate that AMPs can be utilized as feed additives for juvenile goats.

Sodium selenite prevents suppression of mucosal humoral response by AFB1 in broiler's cecal tonsil.

Aflatoxin B1 (AFB1), the most common mycotoxin in human food and animal feed, produces hepatotoxic, genotoxic and immunosuppressive effects in multiple species. Selenium (Se) has emerged as an important element in the dietary prevention of various toxic agents. The present study was designed to scrutinize the protective effects of sodium selenite on the histological lesions and suppression of mucosal humoral response in the cecal tonsil generated by AFB1. A total of 156 one-day-old broilers were divided into four groups and fed on basal diet (control group), 0.6 mg/kg AFB1 (AFB1 group), 0.4 mg/kg Se supplement (+Se group), and 0.6 mg/kg AFB1 + 0.4 mg/kg Se supplement (AFB1+Se group) respectively for 21 days. Our results showed that 0.4 mg/kg Se supplement in broiler's diets could improve the AFB1-induced histological lesions in the cecal tonsils including the depletion of lymphocytes in the lymphatic nodules as well as the shedding of microvilli in the absorptive cells. Moreover, Se could restore the decreased number of IgA+ cells and expression levels of pIgR, IgA, IgG, and IgM mRNA induced by AFB1 to be close to those in the control group. These results demonstrated that 0.4 mg/kg supplemented dietary Se in the form of sodium selenite could protect the cecal tonsils from the histological lesions and suppression of the mucosal humoral response provoked by 0.6 mg/kg AFB1. Our study may provide new experimental evidences for better understanding of AFB1-induced damage of mucosal immunity and protective effect of Se against this toxin.

Protective role of selenium in the activities of antioxidant enzymes in piglet splenic lymphocytes exposed to deoxynivalenol.

We evaluated the effects of selenium (Se) on antioxidant enzymes of piglet splenic lymphocytes exposed to deoxynivalenol (DON). We measured cell viability, the activities of several antioxidant enzymes, and lactate dehydrogenase (LDH), as well as total antioxidant capacity (T-AOC) and the levels of malonaldehyde (MDA) and hydrogen peroxide (H2O2). We found that DON exposure increased the concentrations of LDH, MDA, and H2O2 in all experimental groups in a dose-dependent manner, while the concentrations of other antioxidant enzymes were decreased. In Se-pretreated DON-exposed cells, damage to antioxidant enzymes was reduced, especially in the lower-dose DON groups over longer exposure times. These results may indicate that in piglet splenic lymphocytes, Se can alleviate DON-induced damage to antioxidant enzymes by improving glutathione peroxidase activity. Se may function as a potential antioxidative agent to alleviate DON-induced oxidative stress.

Dietary High Fluorine Alters Intestinal Microbiota in Broiler Chickens.

This study investigated the effects of dietary high fluorine on ileal and cecal microbiota in broiler chickens. Two hundred eighty 1-day-old broiler chickens were randomly assigned to four groups and raised for 42 days. The control group was fed a corn-soybean basal diet (fluorine 22.6 mg/kg). The other three groups were fed the same basal diet, but supplemented with 400, 800, and 1200 mg/kg fluorine (high fluorine groups I, II, and III), administered in the form of sodium fluoride. The microbiota of ileal and cecal digesta was assessed with plate counts and polymerase chain reaction-based denaturing gradient gel electrophoresis (PCR-DGGE). It was found that, compared with those in the control group, the counts of Lactobacillus spp. and Bifidobacterium spp. were markedly decreased (P < 0.01 or P < 0.05), whereas the counts of Escherichia coli and Enterococcus spp. were significantly increased (P < 0.01 or P < 0.05) in the high fluorine groups II and III. PCR-DGGE analysis showed that the number of DGGE bands, similarity, and Shannon index of ileal and cecal bacteria were markedly reduced in the high fluorine groups II and III from 21 to 42 days. Sequencing analysis revealed that the composition of the intestinal microbiota was altered in the high fluorine groups. In conclusion, dietary fluorine in the range of 800-1200 mg/kg obviously altered the bacterial counts, and the diversity and composition of intestinal microbiota in broiler chickens, a finding which implies that dietary high fluorine can disrupt the natural balance and structure of the intestinal microbiota.

Pathway underlying small intestine apoptosis by dietary nickel chloride in broiler chickens.

The aims of this study were to investigate the pathways which dietary nickel chloride (NiCl2) affects small intestine apoptosis in broiler chickens by observing the ultrastructure, and bcl-2, bax, and caspase-3 protein expression and mRNA expression, and cytochrome C, bak and caspase-9 mRNA expression of the small intestine. A total of 240 one-day-old avian broilers were divided into four groups and fed a corn-soybean basal diet as the control diet or three experimental diets supplemented with 300, 600, and 900 mg/kg of NiCl2 for 42 days. Ultrastructurally, the microvilli were apparently exfoliated, and the mitochondria were swollen and the number of lysosomes increased in the intestinal cells of three experimental groups. As measured by TUNEL and flow cytometry (FCM), the percentage of apoptotic cells in the small intestine and the lymphocytes in the ileum were significantly increased in three experimental groups when compared with those of the control group. Meanwhile, immunohistochemistry, quantitative real-time polymerase chain reaction (qRT-PCR) and enzyme-linked immuno-sorbent assay (ELISA) tests showed that the protein expression, mRNA expression levels were decreased in the bcl-2, whereas those of bax and caspase-3, and the cytochrome C, bak and caspase-9 mRNA expression levels were increased in three experimental groups. The abovementioned results show that pathway of dietary NiCl2-induced small intestine apoptosis is related to the mitochondrial damage and promotion of the cytochrome C release from mitochondria, which activates the mitochondrion-mediated apoptosis pathway.

E. adenophorum induces Cell Cycle Arrest and Apoptosis of Splenocytes through the Mitochondrial Pathway and Caspase Activation in Saanen Goats.

The precise cytotoxicity of E. Adenophorum in relation to the cell cycle and apoptosis of splenocytes in Saanen goats remains unclear. In the present study, 16 Saanen goats were randomly divided into four groups, which were fed on 0%, 40%, 60% and 80% E. adenophorum diets. The results of TUNEL, DAPI and AO/EB staining, flow cytometry analysis and DNA fragmentation assays showed that E. adenophorum induced typical apoptotic features in splenocytes, suppressed splenocyte viability, and caused cell cycle arrest in a dose-dependent manner. However, westernblot, ELISA, qRT-PCR and caspase activity analyses showed that E. adenophoruminhibited Bcl-2 expression, promoted Bax translocation to the mitochondria, triggered the release of Cytc from the mitochondria into the cytosol, and activated caspase-9 and -3 and the subsequent cleavage of PARP. Moreover, in E. adenophorum-induced apoptosis, the protein levels of Fas, Bid, FasL and caspase-8 showed no significant changes. E. adenophorum treatment induced the collapse of ΔΨm. Moreover, these data suggested that E. adenophorum induces splenocyte apoptosis via the activation of the mitochondrial apoptosis pathway in splenocytes. These findings provide new insights into the mechanisms underlying the effects of E. adenophorum cytotoxicity on splenocytes.

Induction and mechanism of HeLa cell apoptosis by 9-oxo­10, 11-dehydroageraphorone from Eupatorium adenophorum.

9-Oxo-10, 11-dehydroageraphorone (euptox A), a cadenine sesquiterpene, is the main toxin from Eupatorium adenophorum. The aim of the present study was to examine the induction and mechanism of HeLa cell apoptosis by euptox A. The apoptosis­inducing effect of the euptox A on HeLa cells was examined by MTT assay. The underlying mechanism was analyzed by flow cytometry and quantitative PCR. Flow cytometry results suggested that euptox A effectively inhibited the proliferation of HeLa cells, arrested the cell cycle transition from S to G2/M phase, did not continue to complete the cell cycle activity (mainly from 4 times and mitosis), and induced cell proliferation. The RT-qPCR detection results showed that euptox A induced apoptosis by improving the gene expression level of apoptotic proteases such as caspase-10 in HeLa cells. Its mechanism of action was associated with the upregulation of apoptotic gene expression and arresting of the cell cycle.