Matrine Targets Intestinal Lactobacillus acidophilus to Inhibit Porcine Circovirus Type 2 Infection in Mice.

Porcine circovirus type 2 (PCV2) has caused huge economic losses to the pig industry across the world. Matrine is a natural compound that has been shown to regulate intestinal flora and has anti-PCV2 activity in mouse models. PCV2 infection can lead to changes in intestinal flora. The intestinal flora has proved to be one of the important pharmacological targets of the active components of Traditional Chinese Medicine. This study aimed to determine whether matrine exerts anti-PCV2 effects by regulating intestinal flora. In this study, fecal microbiota transplantation (FMT) was used to evaluate the effect of matrine on the intestinal flora of PCV2-infected Kunming (KM) mice. The expression of the Cap gene in the liver and the ileum, the relative expression of IL-1ß mRNA, and the Lactobacillus acidophilus (L. acidophilus) gene in the ileum of mice were determined by real-time quantitative polymerase chain reaction (qPCR). ELISA was used to analyze the content of secretory immunoglobulin A (SIgA) in small intestinal fluid. L. acidophilus was isolated and identified from the feces of KM mice in order to study its anti-PCV2 effect in vivo. The expression of the Cap gene in the liver and the ileum and the relative expression of L. acidophilus and IL-1ß mRNA in the ileum were determined by qPCR. The results showed that matrine could reduce the relative expression of IL-1ß mRNA by regulating intestinal flora, and that its pharmacological anti-PCV2 and effect may be related to L. acidophilus. L. acidophilus was successfully isolated and identified from the feces of KM mice. The in vivo experiment revealed that administration of L. acidophilus also reduced the relative expression of IL-1ß mRNA, and that it had anti-PCV2 effects in PCV2-infected mice. It was found that matrine could regulate the abundance of L. acidophilus in the gut of mice to exert an anti-PCV2 effect and inhibit PCV2-induced inflammatory response.

Probiotic supplementation containing Bacillus velezensis enhances expression of immune regulatory genes against pigeon circovirus in pigeons (Columba livia).

AIMS: In this study, we aimed to isolate and evaluate the efficacy of Bacillus velezensis as a probiotic and to assess its activity towards pigeons infected with pigeon circovirus (PiCV). METHODS AND RESULTS: Bacillus velezensis, isolated from pigeon faeces, was orally administered to pigeons for 60 days. After pigeons were challenged with PiCV, the PiCV viral load and expression of indicator genes for innate immunity were detected in spleen tissue and faeces of pigeons. Bacillus velezensis significantly reduced the PiCV viral load in the faeces and spleen of pigeons 5 days post-challenge (dpc). The mRNA expression levels of treated pigeons showed that interferon-gamma (IFN-γ), myxovirus resistance 1 (Mx1), and signal transducers and activators of transcription 1 (STAT1) genes were upregulated, whereas no expression of interleukin-4 (IL-4) was detected. Moreover, toll-like receptor 2 (TLR2) and 4 (TLR4) were significantly upregulated in probiotic-treated pigeons (P < 0·05). CONCLUSIONS: This is the first report showing that probiotic supplementation can effectively enhance the T-helper type 1 immune response and decrease the PiCV viral loads in pigeons. SIGNIFICANCE AND IMPACT OF THE STUDY: This study proposes that the administration of a probiotic strain, B. velezensis, to pigeons can protect against PiCV infection.

Cepharanthine and Curcumin inhibited mitochondrial apoptosis induced by PCV2.

BACKGROUND: Porcine circovirus type 2 (PCV2) is an immunosuppressive pathogen with high prevalence rate in pig farms. It has caused serious economic losses to the global pig industry. Due to the rapid mutation of PCV2 strain and co-infection of different genotypes, vaccination could not eradicate the infection of PCV2. It is necessary to screen and develop effective new compounds and explore their anti-apoptotic mechanism. The 13 natural compounds were purchased, with a clear plant origin, chemical structure and content and specific biological activities. RESULTS: The maximum no-cytotoxic concentration (MNTC) and 50% cytotoxic concentration (CC50) of 13 tested compounds were obtained by the cytopathologic effect (CPE) assay and (3-(4,5-dimethyithiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) method in PK-15 cells. The results of qPCR and Western blot showed that, compared with the PCV2 infected group, the expression of Cap in Paeonol (0.4 mg/mL and 0.2 mg/mL), Cepharanthine (0.003 mg/mL, 0.0015 mg/mL and 0.00075 mg/mL) and Curcumin (0.02 mg/mL, 0.001 mg/mL and 0.005 mg/mL) treated groups were significantly lowered in a dose-dependent manner. The results of Annexin V-FITC/PI, JC-1, Western blot and ROS analysis showed that the expression of cleaved caspase-3 and Bax were up-regulated Bcl-2 was down-regulated in Cepharanthine or Curcumin treated groups, while ROS and MMP value were decreased at different degrees and the apoptosis rate was reduced. In this study, Ribavirin was used as a positive control. CONCLUSIONS: Paeonol, Cepharanthine and Curcumin have significant antiviral effect. And the PCV2-induced Mitochondrial apoptosis was mainly remitted by Cepharanthine and Curcumin.

Matrine exhibits antiviral activity in a PRRSV/PCV2 co-infected mouse model.

BACKGROUND: PRRSV and PCV2 co-infection is very common in swine industry which results in huge economic losses worldwide. Although vaccination is used to prevent viral diseases, immunosuppression induced by PRRSV and PCV2 leads to vaccine failure. PURPOSE: Our previous results have demonstrated that Matrine possess antiviral activities against PRRSV/PCV2 co-infection in vitro. This study aims to establish a PRRSV/PCV2 co-infected KM mouse model and evaluate the antiviral activities of Matrine against PRRSV/PCV2 co-infection. STUDY DESIGN: A total of 144 KM mice were randomly divided into six groups with 24 mice in each group, named as: normal control, PRRSV/PCV2 co-infected group (PRRSV/PCV2 group), Ribavirin treatment positive control (Ribavirin control) and Matrine treatment groups (Matrine 40 mg/kg, Matrine 20 mg/kg and Matrine 10 mg/kg). METHODS: Except normal control group, all mice in other five groups were inoculated with PRRSV, followed by PCV2 at 2 h later. At 7 days post-infection (dpi), mice in the treatment groups were intraperitoneally administered with various doses of Matrine and Ribavirin, twice a day for 5 consecutive days. RESULTS: PRRSV N and PCV2 CAP genes were detected by PCR in multiple tissues including heart, liver, spleen, lungs, kidneys, thymus and inguinal lymph nodes. The viral load of PCV2 was the highest in liver followed by thymus and spleen. Although PRRSV were detected in most of tissues, but the replication of PRRSV was not significantly increased, as shown by qPCR analysis. Comparing with PCV2 infection alone, PRRSV infection significantly elevated PCV2 replication and exacerbated PCV2 induced interstitial pneumonia. qPCR analysis demonstrated 40 mg/kg Matrine significantly attenuated PCV2 replication in liver and alleviated virus induced interstitial pneumonia, suggesting Matrine could directly inhibit virus replication. In addition, Matrine treatment enhanced peritoneal macrophages phagocytosis at 13 and 16 dpi, and 40 mg/kg of Matrine increased the proliferation activity of lymphocytes. Body weight gain was continuously promoted by administrating Matrine at 10 mg/kg. CONCLUSION: Matrine possessed antiviral activities via inhibiting virus replication and regulating immune functions in mice co-infected by PRRSV/PCV2. These data provide new insight into controlling PRRSV and PCV2 infection and support further research for developing Matrine as a new possible veterinary medicine.

Sophorasubprosrate polysaccharide suppress the inflammatory reaction of RAW264.7 cells infected with PCV2 via regulation NF-κB/MAPKs/c-Jun signal pathway and histone acetylation modification.

The purpose of this study was to investigate the regulation of Sophorasubprosrate polysaccharide (SSP) on inflammatory response and histone acetylation modification of RAW264.7 cells (mouse mononuclear macrophage cell line) infected with porcine circovirus type 2 (PCV2). We further explored the role of inflammatory response and histone acetylation modification on the basis of the original study. The results showed that SSP decreased the secretion levels of TNF-α and IL-6 and the intracellular iNOS, COX-2 enzyme activities and their mRNA expression levels in PCV2 infected RAW264.7 cells, but increased the level of IL-10 secretion and its mRNA expression. SSP inhibited the phosphorylation levels of proteins of p65, ERK1/2, p38 and c-Jun in RAW264.7 cells infected with PCV2. The activities of HAT and HDAC enzymes and the mRNA expression levels of HAT1 and HDAC1 were increased when the PCV2-infected RAW264.7 cells were treated by SSP. Meanwhile, the expression of acetylation modification of histones both H3 and H4 was obviously inhibited. In conclusion, SSP may reduce the acetylation levels of both H3 and H4 and activate NF-κB/MAPKs/c-Jun signaling pathway by increasing the activity of HADC enzyme and the expression of HDAC mRNA, further inhibiting inflammatory response by regulating the gene expression levels of inflammatory factors. The findings indicated that the molecular mechanism of how traditional Chinese medicine polysaccharide regulates inflammatory signal pathways and inflammatory factors by regulating histone acetylation.

Antiviral Effect of Epigallocatechin Gallate via Impairing Porcine Circovirus Type 2 Attachment to Host Cell Receptor.

The green tea catechin epigallocatechin gallate (EGCG) exhibits antiviral activity against various viruses. Whether EGCG also inhibits the infectivity of circovirus remains unclear. In this study, we demonstrated the antiviral effect of EGCG on porcine circovirus type 2 (PCV2). EGCG targets PCV2 virions directly and blocks the attachment of virions to host cells. The microscale thermophoresis assay showed EGCG could interact with PCV2 capsid protein in vitro with considerable affinity (Kd = 98.03 ± 4.76 µM), thereby interfering with the binding of the capsid to the cell surface receptor heparan sulfate. The molecular docking analysis of capsid-EGCG interaction identified the key amino acids which formed the binding pocket accommodating EGCG. Amino acids ARG51, ASP70, ARG73 and ASP78 of capsid were found to be critical for maintaining the binding, and the arginine residues were also essential for the electrostatic interaction with heparan sulfate. The rescued mutant viruses also confirm the importance of the key amino acids of the capsid to the antiviral effect of EGCG. Our findings suggest that catechins could act as anti-infective agents against circovirus invasion, as well as provide the basic information for the development and synthesis of structure-based anti-circovirus drugs.

Lipid peroxidation impairs growth and viability of nursery pigs reared under commercial conditions1.

The objective of this study was to investigate the impact of lipid peroxidation in a dose-dependent manner on growth, health, and oxidative stress status of nursery pigs. A total of 2,200 weaned pigs (5.95 ± 0.20 kg BW) were housed in 100 pens (22 pigs per pen) in a randomized complete block design based on initial BW and sex. Pigs were randomly assigned within blocks to 5 dietary treatments, consisting of a corn-soybean meal-based diet supplemented with 5% of either control corn oil (iodine value = 118, FFA = 0.06%, anisidine value = 3, peroxide value = 3 mEq/kg oil) or peroxidized corn oil (iodine value = 120, FFA = 0.35%, anisidine value = 30, peroxide value = 163 mEq/kg oil). These 2 diets were blended to obtain 5 levels of peroxidation with final treatments designated as 0 (diet with 5% control oil), 25%, 50%, 75%, and 100% (diet with peroxidized corn oil) peroxidation. Diets were fed ad libitum for 43 d. Blood samples were collected on d 33 from 20 pigs per treatment to determine serum oxidative stress markers and vitamin E concentrations and again on d 43 (14 d after vaccination) to determine immune response to porcine circovirus type 2 (PCV2) and Mycoplasma hyopneumoniae (Mhyo). Gain:feed ratio decreased linearly (P = 0.023) with increasing peroxidation, but pen ADG and ADFI were not affected. Number of pigs removed for medical treatment, total number medically treated, pigs culled for low end weight, and mortality increased, and full-value pigs linearly decreased (P < 0.04) with increasing peroxidation. Consequently, total pen gain (weight of viable pigs that remained in test pens at the end of the study minus weight of pigs placed) decreased linearly (P < 0.01) with increasing peroxidation. Antibody titers to Mhyo and PCV2 increased postvaccination (P < 0.001), but did not differ due to dietary treatment. Serum concentrations of malondialdehyde, 8-hydroxy-2'-deoxyguanosine, and protein carbonyl were not affected by peroxidation. Total antioxidant capacity and serum vitamin E concentrations decreased (P = 0.01) linearly with increasing peroxidation. Data show a dose-dependent negative impact of lipid peroxidation on pig productivity when determined under field population conditions, being primarily manifested by increased mortality, number of pigs medically treated, and number of culled pigs (≤13.6 kg BW). Results underscore the importance of proper assessment of lipid peroxidation as part of quality control to prevent oxidative stress and performance losses in weaned pigs.

Taurine attenuates OTA-promoted PCV2 replication through blocking ROS-dependent autophagy via inhibiting AMPK/mTOR signaling pathway.

Previous research found that ochratoxin A (OTA) could promote PCV2 replication by inducing autophagy. The aim of this study is to evaluate the effect of dietary amino acid derivative taurine on OTA-promoted PCV2 replication and explore the underlying mechanism. The results showed that taurine could inhibit OTA-promoted PCV2 replication in PK-15 cells. The effect of taurine could be mediated by its ability to attenuate ROS level and block OTA-promoted autophagy. Indeed, induction of autophagy by rapamycin could suppress the inhibitory effect of taurine on OTA-promoted PCV2 replication. Furthermore, taurine supplementation inhibited 5'AMP-activated protein kinase (AMPK) and activated mammalian target of rapamycin (mTOR). Activation of AMPK by acadesine (AICAR) could suppress the effect of taurine. In conclusion, taurine treatment suppresses autophagy by regulating the ROS/AMPK/mTOR signaling axis, thereby inhibiting OTA-promoted PCV2 replication. These findings provide the rationale for the use of taurine as an intervention against PCV2 infection.

Porcine circovirus type 3 (PCV3) shedding in sow colostrum.

The major objective of this work was to investigate the shedding of porcine circovirus type 3 (PCV3) in sow colostrum. PCV3 titers in the serum and colostrum samples of 38 sows were determined using qPCR. Interestingly, this is the first report regarding the identification of PCV3 from the colostrum samples. In the studied farm, the prevalence of PCV3 in the colostrum samples was 44.74% (17/38). When sows were grouped based on the PCV3 titers in the serum into the "High-viremic", "Low-viremic" and "Non-viremic" sows, it was shown that the High-viremic sows showed significantly higher PCV3 colostrum prevalence (100%; 9/9) with the PCV3 titers ranging from 4.01 to 7.33 genomic copies/mL. The results indicated that PCV3 in the colostrum might be partly influenced by the viremic stage of the infection. However, the results also showed that approximately 41% of sows shedding PCV3 with low titers in the colostrum (7/17) were non-viremic sows. In conclusion, this study identified the presence of PCV3 in sow colostrum. Clinical impacts and mechanisms of colostrum shedding of PCV3 should be further investigated.

SeMet attenuates OTA-induced PCV2 replication promotion by inhibiting autophagy by activating the AKT/mTOR signaling pathway.

Porcine circovirus type 2 (PCV2) is recognized as the causative agent of porcine circovirus-associated diseases. PCV2 replication could be promoted by low doses of ochratoxin A (OTA) as in our previous study and selenium has been shown to attenuate PCV2 replication. However, the underlying mechanism remains unclear. The aim of the study was to investigate the effects of selenomethionine (SeMet), the major component of organic selenium, on OTA-induced PCV2 replication promotion and its potential mechanism. The present study demonstrates that OTA could promote PCV2 replication as measured by cap protein expression, viral titer, viral DNA copies and the number of infected cells. In addition, OTA could activate autophagy as indicated by up-regulated light chain 3 (LC3)-II and autophagy-related protein 5 expressions and autophagosome formation. Further, OTA could down-regulate p-AKT and p-mTOR expressions and OTA-induced autophagy was inhibited when insulin was applied. SeMet at 2, 4 and 6 µM had significant inhibiting effects against OTA-induced PCV2 replication promotion. Furthermore, SeMet could attenuate OTA-induced autophagy and up-regulate OTA-induced p-AKT and p-mTOR expression inhibition. Rapamycin, an inhibitor of AKT/mTOR, could reverse the effects of SeMet on OTA-induced autophagy and the PCV2 replication promotion. In conclusion, SeMet could block OTA-induced PCV2 replication promotion by inhibiting autophagy by activating the AKT/mTOR pathway. Therefore, SeMet supplementation could be an effective prophylactic strategy against PCV2 infections and autophagy may be a potential marker to develop novel anti-PCV2 drugs.

Comparison of cytokine profiles in peripheral blood mononuclear cells between piglets born from Porcine circovirus 2 vaccinated and non-vaccinated sows.

This study was aimed to evaluate the effect of Porcine circovirus 2 (PCV2) sow vaccination on cell-mediated immune responses in sows and their progeny. At 7 weeks before farrowing, fifteen PCV2 PCR negative pregnant sows with medium-low antibody values were selected and randomly distributed in two groups according to the antibody levels. Seven sows were vaccinated with a commercial PCV2 vaccine and eight were injected with phosphate-buffered saline at 6 and 3 weeks before farrowing. Blood samples were taken from sows and their piglets (n = 90) during the study duration. PCV2 DNA and antibodies were tested in sera, and cytokine (IFN-α, IFN-γ, IL-12p40, TNF-α, IL-1ß, IL-8, IL-4, IL-6 and IL-10) levels were assessed in supernatant from cultured peripheral blood mononuclear cells. All sows and piglets were negative by PCV2 PCR throughout the study. Significantly higher PCV2 antibody levels were detected in vaccinated sows after vaccination and in their offspring after colostrum ingestion compared to the non-vaccinated counterparts. Vaccinated sows did not show significant differences in cytokine secretion levels at farrowing compared to unvaccinated dams. In contrast, piglets from vaccinated sows had significantly higher levels of cytokines linked to Th1 memory cells (IFN-γ and TNF-α) in comparison to the ones from non-vaccinated dams. In conclusion, PCV2 sow vaccination, apart from triggering a humoral immunity response in sows and their progeny, might be associated to an increased transfer of cell-mediated immunity from the dam to the piglet.

Selenizing astragalus polysaccharide attenuates PCV2 replication promotion caused by oxidative stress through autophagy inhibition via PI3K/AKT activation.

Our previous studies have shown that oxidative stress could promote the porcine circovirus type 2 (PCV2) replication, and astragalus polysaccharide (APS)/selenium could suppress PCV2 replication. However, whether selenizing astragalus polysaccharide (sAPS) provides protection against oxidative stress-induced PCV2 replication promotion and the mechanism involved remain unclear. The present study aimed to explore the mechanism of the PCV2 replication promotion induced by oxidative stress and a novel pharmacotherapeutic approach involving the regulation of autophagy of sAPS. Our results showed that H2O2 promoted PCV2 replication via enhancing autophagy by using 3-methyladenine (3-MA) and autophagy-related gene 5 (ATG5) knockdown. Sodium selenite, APS, the mixture of sodium selenite and APS, and sAPS significantly inhibited H2O2-induced PCV2 replication promotion, respectively. Among these, sAPS exerted maximal inhibitory effect. sAPS could also significantly inhibit autophagy activated by H2O2 and increase the Akt and mTOR phosphorylation. Moreover, LY294002, the specific phosphoinositide 3-kinase/protein kinase B (PI3K/AKT) inhibitor, significantly alleviated the effects of sAPS on autophagy and PCV2 replication. Taken together, we conclude that H2O2 promotes PCV2 replication by inducing autophagy and sAPS attenuates the PCV2 replication promotion through autophagy inhibition via PI3K/AKT activation.

Effect of Maternal Antibody Transfer on Antibody Dynamics and Control of Porcine Circovirus Type 2 Infection in Offspring.

Sow immunity plays an important role in preventing viral infection and disease in newborn piglets. Vertical transmission of porcine circovirus type 2 (PCV2) may perpetuate porcine circovirus associated disease (PCVAD) in newborn and growing pigs. Hence, the immunological effects of maternal immunoglobulin transfer of PCV2-specific antibodies on PCV2 viremia and immune response in piglets in commercial swine herds were evaluated. Sow vaccination has been shown to reduce viral shedding and viremia, and increases the neutralizing antibody (NA) titers. Since NAs are important for control of PCVAD and mammary secretions may contain high anti-PCV2 NA levels, we examined the PCV2 NA levels in colostrum, milk, sow serum, and piglet serum over time to investigate an association between NA levels and protection against infection. NA titers were remarkably high (up to 10-6 50% neutralizing titer) in sow serum and colostrum on all farms regardless of viremia levels. In piglets vaccinated at 3 weeks of age, NA titers peaked at 10 weeks of age and continued to maintain high viral neutralizing titers to slaughter. The impact of maternally derived neutralizing activity was most evident during the suckling period. Although PCV2 was transmitted from sows to piglets in colostrum, piglets were largely nonviremic at weaning. Thus, NAs appear to control or suppress initial infection even though they are unable to clear or prevent infection later in life.

Immunomodulatory and antioxidant effects of total flavonoids of Spatholobus suberectus Dunn on PCV2 infected mice.

Oxidative stress plays an important role in the pathogenesis of virus infection and antioxidants are becoming promising candidates as therapeutic agents. This study is designed to investigate the effect of total flavonoids of Spatholobus suberectus Dunn (TFSD) on oxidative stress in mice induced by porcine circovirus type 2 (PCV2) infection. The PCV2 infection leads to significant decrease in thymus and spleen indices, elevation of xanthine oxidase (XOD) and myeloperoxidase (MPO) activities, reduction in GSH level and GSH to GSSG ratio and decline of superoxide dismutase (SOD) activity, indicating the formation of immunosuppression and oxidative stress. TFSD treatment recovered the alteration of viscera index, antioxidant content and activities of oxidative-associated enzymes to a level similar to control. Our findings suggested that PCV2 induced immunosuppression and oxidative stress in mice and TFSD might be able to protect animals from virus infection via regulation of immune function and inhibition of oxidative stress.

Effect of total flavonoids of Spatholobus suberectus Dunn on PCV2 induced oxidative stress in RAW264.7 cells.

BACKGROUND: This study was carried out to investigate the effect of total flavonoids of Spatholobus suberectus Dunn (TFSD) on PCV2 induced oxidative stress in RAW264.7 cells. METHODS: Oxidative stress model was established in RAW264.7 cells by infecting with PCV2. Virus infected cells were then treated with various concentrations (25 mg/ml, 50 mg/ml and 100 mg/ml) of TFSD. The levels of oxidative stress related molecules (NO, ROS, GSH and GSSG) and activities of associated enzymes (SOD, MPO and XOD were analyzed using ultraviolet spectrophotometry, fluorescence method and commercialized detection kits. RESULTS: PCV2 infection induced significant increase of NO secretion, ROS generation, GSSG content, activities of both XOD and MPO, and dramatically decrease of GSH content and SOD activity in RAW264.7 cells (P < 0.05). After treating with TFSD, PCV2 induced alteration of oxidative stress related molecule levels and enzyme activities were recovered to a level similar to control. CONCLUSION: Our findings indicated that TFSD was able to regulate oxidative stress induced by PCV2 infection in RAW264.7 cells, which supports the ethnomedicinal use of this herb as an alternative or complementary therapeutic drug for reactive oxygen-associated pathologies.

Astragalus polysaccharides attenuate PCV2 infection by inhibiting endoplasmic reticulum stress in vivo and in vitro.

This study explored the effects of Astragalus polysaccharide (APS) on porcine circovirus type 2 (PCV2) infections and its mechanism in vivo and vitro. First, fifty 2-week-old mice were randomly divided into five groups: a group without PCV2 infection and groups with PCV2 infections at 0, 100, 200 or 400 mg/kg APS treatments. The trial lasted for 28 days. The results showed that APS treatments at 200 and 400 mg/kg reduced the pathological injury of tissues, inhibited PCV2 infection and decreased glucose-regulated protein 78 (GRP78) and GADD153/CHOP gene mRNA and protein expression significantly (P < 0.05). Second, a study on endoplasmic reticulum stress mechanism was carried out in PK15 cells. APS treatments at 15 and 45 µg/mL significantly reduced PCV2 infection and GRP78 mRNA and protein expression (P < 0.05). Tunicamycin supplementation increased GRP78 mRNA and protein expression and significantly attenuated the APS-induced inhibition of PCV2 infection (P < 0.05). Tauroursodeoxycholic acid supplementation decreased GRP78 mRNA and protein expression and significantly inhibited PCV2 infection (P < 0.05). In addition, fifty 2-week-old mice were randomly divided into five groups: Con, PCV2, APS + PCV2, TM + PCV2 and TM + APS + PCV2. The results were similar to those in PK15 cells. Taken together, it could be concluded that APS suppresses PCV2 infection by inhibiting endoplasmic reticulum stress.

The antiviral activity of arctigenin in traditional Chinese medicine on porcine circovirus type 2.

Arctigenin (ACT) is a phenylpropanoid dibenzylbutyrolactone lignan extracted from the traditional herb Arctium lappa L. (Compositae) with anti-viral and anti-inflammatory effects. Here, we investigated the antiviral activity of ACT found in traditional Chinese medicine on porcine circovirus type 2 (PCV2) in vitro and in vivo. Results showed that dosing of 15.6-62.5µg/mL ACT could significantly inhibit the PCV2 proliferation in PK-15 cells (P<0.01). Dosing of 62.5µg/mL ACT 0, 4 or 8h after challenge inoculation significantly inhibited the proliferation of 1MOI and 10MOI in PK-15 cells (P<0.01), and the inhibitory effect of ACT dosing 4h or 8h post-inoculation was greater than 0h after dosing (P<0.01). In vivo test with mice challenge against PCV2 infection demonstrated that intraperitoneal injection of 200µg/kg ACT significantly inhibited PCV2 proliferation in the lungs, spleens and inguinal lymph nodes, with an effect similar to ribavirin, demonstrating the effectiveness of ACT as an antiviral agent against PCV2 in vitro and in vivo. This compound, therefore, may have the potential to serve as a drug for protection of pigs against the infection of PCV2.

The immunological activity of Lycium barbarum polysaccharides liposome in vitro and adjuvanticity against PCV2 in vivo.

In previous researches, the results showed that Lycium barbarum polysaccharides (LBP) encapsulated with liposome could enhance the immune activity of LBP. Therefore, the present study was designed to investigate the effects of LBPL on spleen lymphocytes and macrophages of mice in vitro and evaluate the immunological adjuvant activity of PCV2 vaccine in vivo. The results showed that LBPL could significantly promote splenocyte proliferation synergistically with PHA or LPS, increase the ratio of CD4(+) to CD8(+) T cells and promote the cytokine secretion of macrophages; enhance PCV2-specific IgG antibody responses, promote Th1 cytokines (IFN-γ and TNF-a) and Th2 cytokine (IL-4) secretion. The histomorphological observation of spleen demonstrated that LBPL as a vaccine adjuvant also has good improvement and stimulating effect on the immune organ.

Role of Chinese herbal medicinal ingredients in secretion of cytokines by PCV2-induced endothelial cells.

While T-lymphocytes are the major cell type responsible for host responses to a virus (including induction of inflammatory responses to aid in ultimate removal of virus), other cells, including macrophages, epithelial and dendritic cells also have key roles. Endothelial cells also play important roles in physiologic/pathologic processes, like inflammation, during viral infections. As endothelial cells can be activated to release various endogenous compounds, including some cytokines, ex vivo measures of cytokine formation by the cells can be used to indirectly assess any potential endothelial dysfunction in situ. The research presented here sought to investigate potential immunolomodulatory effects of five saponins on endothelial cells: Saikosaponins A (SSA) and D (SSD), Panax Notoginseng Saponin (PNS) and Notoginsenoside R1 (SR1) and Anemoside B4 (AB4). For this, cells (porcine iliac artery endothelial line) were challenged with a virus isolate PCV2-AH for 24 h and then treated with the test saponin (at 1, 5 or 10 µg/ml) for an additional 24 h at 37 °C. The culture supernatants were then collected and analyzed for interleukin (IL)-2, -4 and -10, as well as interferon (IFN)-γ by ELISA. The results revealed that PNS and SR1 inhibited the production of IL-4; PNS, SR1 and AB4 inhibited the secretion of IL-10; SSA, SSD and PNS up-regulated IL-2 expression; SSA and SSD increased the level of IFNγ. All these changes were significant. Taken together, the data suggested these saponins might potentially have a capacity to regulate immune responses in vivo via changes in production of these select cytokines by infected endothelial cells. Nevertheless, the impact of these agents on other key cell types involved in anti-viral responses, including T-lymphocytes, remains to be determined.

Astragalus polysaccharides inhibits PCV2 replication by inhibiting oxidative stress and blocking NF-κB pathway.

Porcine circovirus type 2 (PCV2) is the primary causative agent of porcine circovirus-associated disease (PCVAD). Astragalus polysaccharide (APS), as one kind of biological macromolecule extracted from Astragalus, has antiviral activities. This study was undertaken to explore the effect of APS on PCV2 replication in vitro and the underlying mechanisms. Our results showed that adding APS before PCV2 infection decreased significantly PCV2 DNA copies, the number of infected cells, MDA level, ROS level and NF-κB activation in PK15 cells and increased significantly GSH contents and SOD activity compared to control without APS. Oxidative stress induced by BSO could eliminate the effect of PCV2 replication inhibition by APS. LPS, as a NF-κB activator, could attenuate the effect of PCV2 replication inhibition by APS. BAY 11-7082, as a NF-κB inhibitor, could increase the effect of PCV2 replication inhibition by APS. In conclusion, APS inhibits PCV2 replication by decreasing oxidative stress and the activation of NF-κB signaling pathway, which suggests that APS might be employed for the prevention of PCV2 infection.