YBX has functional roles in CpG-ODN against cold stress and bacterial infection of Misgurnus anguillicaudatus.

Misgurnus anguillicaudatus (M. anguillicaudatus) is a widely cultivated fish. However, in M. anguillicaudatus breeding, the frequent cold stress during daily breeding could induce immune suppression and increase the risk of infection, causing serious economic loss. Based on existing findings, CpG Oligonucleotides (CpG-ODNs) may be an ideal protective agent for low temperature fish breeding, performing anti-infective when faced with cold stress with cold shock proteins Y box binding proteins (YBX). Although YBX has pleiotropic functions, its roles in CpG-ODNs-mediated immunity (especially under cold situations) remain largely unexplored. To clarify the relationship among them, we identified the YBX1/YBX2 in M. anguillicaudatus and analyzed using a series of bioinformatics methods. After that, we immunized the fish with 3 types of CpG-ODNs and challenged with Aeromonas hydrophila (A. hydrophila). Here we showed that the best anti-bacterial effect of CpG-B was accompanied by the significant upregulation of YBX1. And the detection of the YBX1 downstream effectors confirmed that CpG-B induced the YBX1-mediated Th1 oriented responses to A. hydrophila by regulation of the NLRP3 (Caspase-A/-B), IL-1ß, IL-12 and IFN-γ. Afterwards, we found that under cold stress, CpG-B can activate the NLRP3 and NF-κB pathways through YBX1, a key mediator of anti-A. hydrophila in CpG-B immunization. In this study, we demonstrated CpG-B protection against infection in low temperature, and its interaction with YBX1, expanded the research of CpG-ODN under cold stress, and provided a new CpG-ODN application for low temperature fish farming.

Overexpression of pEGF improved the gut protective function of Clostridium butyricum partly through STAT3 signal pathway.

Clostridium butyricum (C. butyricum) is a probiotic that could promote animal growth and protect gut health. So far, current studies mainly keep up with the basic biological functions of C. butyricum, missing the effective strategy to further improve its protective efficiency. A recent report about C. butyricum alleviating intestinal injury through epidermal growth factor receptor (EGFR) inspired us to bridge this gap by porcine epidermal growth factor (EGF) overexpression. Lacking a secretory overexpression system, we constructed the recombinant strains overexpressing pEGF in C. butyricum for the first time and obtained 4 recombinant strains for highly efficient secretion of pEGF (BC/pPD1, BC/pSPP, BC/pGHF, and BC/pDBD). Compared to the wild-type strain, we confirmed that the expression level ranges of the intestinal development-related genes (Claudin-1, GLUT-2, SUC, GLP2R, and EGFR) and anti-inflammation-related gene (IL-10) in IPECs were upregulated under recombinant strain stimulation, and the growth of Staphylococcus aureus and Salmonella typhimurium was significantly inhibited as well. Furthermore, a particular inhibitor (stattic) was used to block STAT3 tyrosine phosphorylation, resulting in the downregulation on antibacterial effect of recombinant strains. This study demonstrated that the secretory overexpression of pEGF in C. butyricum could upregulate the expression level of EGFR, consequently improving the intestinal protective functions of C. butyricum partly following STAT3 signal activation in IPECs and making it a positive loop. These findings on the overexpression strains pointed out a new direction for further development and utilization of C. butyricum. KEY POINTS: • By 12 signal peptide screening in silico, 4 pEGF overexpression strains of C. butyricum/pMTL82151-pEGF for highly efficient secretion of pEGF were generated for the first time. • The secretory overexpression of pEGF promoted the intestinal development, antimicrobial action, and anti-inflammatory function of C. butyricum. • The overexpressed pEGF upregulated the expression level of EGFR and further magnified the gut protective function of recombinant strains which in turn partly depended on STAT3 signal pathway in IPECs.

Intestinal guard: Human CXCL17 modulates protective response against mycotoxins and CXCL17-mimetic peptides development.

Mycotoxin contamination is an ongoing and growing issue that can create health risks and even cause death. Unfortunately, there is currently a lack of specific therapy against mycotoxins with few side effects. On the other hand, the strategic expression of CXCL17 in mucosal tissues suggests that it may be involved in immune response when exposed to mycotoxins, but the exact role of CXCL17 remains largely unknown. Using Caco-2 as a cell model of the intestinal epithelial barrier (the first line of defense against mycotoxins), we showed that a strong production of ROS-dependent CXCL17 was triggered by mycotoxins via p38 and JNK pathways. Under the mycotoxins stress, CXCL17 modulated enhanced immuno-protective response with a remission of inflammation and apoptosis through PI3K/AKT/mTOR. Based on our observed feedback of CXCL17 to the mycotoxins, we developed the CXCL17-mimetic peptides in silico (CX1 and CX2) that possessed the safety and the capability to ameliorate mycotoxins-inducible inflammation and apoptosis. In this study, the identification of detoxifying feature of CXCL17 is a prominent addition to the chemokine field, pointing out a new direction for curing the mycotoxins-caused damage.

Protective effects of Clostridium butyricum against oxidative stress induced by food processing and lipid-derived aldehydes in Caco-2 cells.

The human body is almost always facing the oxidative stress caused by foodborne aldehydes such as glyoxal (GO) and methylglyoxal (MGO), 4-hydroxyhexenal (HHE), and 4-hydroxynonenal (HNE). When these aldehydes build up, they can cause a range of harm. However, a probiotic, Clostridium butyricum, can increase nuclear factor erythroid-2 related factor 2 (Nrf2) and may have the potential to relieve oxidative stress. If C. butyricum is indeed resistant to aldehydes, the advantages (accessibility, convenience, and safety) will be of great significance compared with drugs. Unfortunately, whether C. butyricum can play a role in alleviating toxic effects of foodborne aldehydes in the intestine (the first line of defense against food-derived toxin) was unclear. To investigate these, we measured the viability, ROS, autophagy, and inflammatory cytokine expression of Caco-2 which were co-cultured with C. butyricum and stimulated by the four aldehydes via Nrf2 pathway (Staphylococcus aureus and Enterococcus faecium as controls). Then, we explored the link among C. butyricum, NLRP6, and Nrf2 signaling pathways when facing the stimuli. In the present study, we demonstrated that Clostridium butyricum relieved the oxidative stress induced by the aldehydes in Caco-2. Most interestingly, we found a "complementary" relationship between NLRP6 and Nrf2 in C. butyricum treatment under aldehyde stress. Our research not only makes a contribution to the popularization of C. butyricum as a probiotic-rich food instead of medicines but also sheds new light on the application of subsequent microecological formulation of C. butyricum. KEY POINTS: • The adverse effects are caused in a dose-dependent manner by foodborne aldehydes. • Clostridium butyricum can significantly ameliorate oxidative stress. • There is a "complementary" relationship between the NLRP6 and Nrf2 signaling pathways. • Using Clostridium butyricum foods to alleviate oxidative stress shows great prospects.

Identification of RBP4 from bighead carp (Hypophthalmichthys nobilis) / silver carp (Hypophthalmichthys molitrix) and effects of CpG ODN on RBP4 expression under A. hydrophila challenge.

Retinol-binding protein 4 (RBP4) is known as a highly conserved adipokine for immune activation. Aeromonas hydrophila (A. hydrophila) is the most common zoonotic pathogen in aquaculture, which causes serious economic losses to aquaculture, especially to bighead carp (Hypophthalmichthys nobilis, H. nobilis) and silver carp (Hypophthalmichthys molitrix, H. molitrix). Recent studies along with our previous findings have shown that synthetic oligodeoxynucleotides containing CpG motifs (CpG ODN) can play a good role in aquatic animals against infection. In order to clarify the relationship between CpG ODN and RBP4 under A. hydrophila infection, firstly, full-length RBP4 cDNAs from H. nobilis and H. molitrix were cloned. And characteristics of RBP4, including sequence and structure, tissue distribution and genetic evolution were analyzed. In addition, mRNA expression levels of RBP4, cytokine, toll-like receptors (TLRs), morbidity and survival rates of H. nobilis and H. molitrix were observed post CpG ODN immunization or following challenge. The results indicated that hn/hm_RBP4 (RBP4 genes obtained from H. nobilis and H. molitrix) had the highest homology with Megalobrama amblycephala. Distribution data showed that the expression level of hn_RBP4 mRNA was higher than that of hm_RBP4. After CpG ODN immunization followed by A.hydrophila challenge, significantly higher survival was observed in both carps, together with up-regulated RBP4 expression. Meanwhile, hn/hm_IL-1ß level was relatively flat (and decreased), hn/hm_IFN-γ, hn/hm_TLR4 and hn/hm_TLR9 levels increased significantly, but hn/hm_STRA6 showed no significant change, compared with control. Moreover, CpG ODN immunization could induce stronger immune protective responses (higher IFN-γ/gentle IL-1ß level and lower morbidity/higher survival rate) against A. hydrophila in H. nobilis, along with higher RBP4 level, when compared with that in H. molitrix. These results demonstrated that RBP4 was well involved in the immune protection of CpG ODN. Based on the results, we speculated that in the case of A. hydrophila infection, TLR9 signaling pathway was activated by CpG ODN. Subsequently, CpG ODN up-regulated RBP4, and RBP4 activated TLR4 signaling pathway. Then TLR4 and TLR9 synergistically improved the anti-infection responses. Our findings have good significance for improving resistance to pathogen infection in freshwater fish.

Co-expressing GroEL-GroES, Ssa1-Sis1 and Bip-PDI chaperones for enhanced intracellular production and partial-wall breaking improved stability of porcine growth hormone.

Porcine growth hormone (pGH) is a class of peptide hormones secreted from the pituitary gland, which can significantly improve growth and feed utilization of pigs. However, it is unstable and volatile in vitro. It needs to be encapsulated in liposomes when feeding livestock, whose high cost greatly limits its application in pig industry. Therefore we attempted to express pGH as intracellular soluble protein in Pichia pastoris and feed these yeasts with partial wall-breaking for swine, which could release directly pGH in intestine tract in case of being degraded in intestinal tract with low cost. In order to improve the intracellular soluble expression of pGH protein in Pichia pastoris and stability in vitro, we optimized the pGH gene, and screened molecular chaperones from E. coli and Pichia pastoris respectively for co-expressing with pGH. In addition, we had also explored conditions of mechanical crushing and fermentation. The results showed that the expression of intracellular soluble pGH protein was significantly increased after gene optimized and co-expressed with Ssa1-Sis1 chaperone from Pichia pastoris. Meanwhile, the optimal conditions of partial wall-breaking and fermentation of Pichia pastoris were confirmed, the data showed that the intracellular expression of the optimized pGH protein co-expressed with Ssa1-Sis1 could reach 340 mg/L with optimal conditions of partial wall-breaking and fermentation. Animal experiments verified that the optimized pGH protein co-expression with Ssa1-Sis1 had the best promoting effects on the growth of piglets. Our study demonstrated that Ssa1-Sis1 could enhance the intracellular soluble expression of pGH protein in Pichia pastoris and that partial wall-breaking of yeast could prevent pGH from degradation in vitro, release targetedly in the intestine and play its biological function effectively. Our study could provide a new idea to cut the cost effectively, establishing a theoretical basis for the clinic application of unstable substances in vitro.

Molecular cloning, characterization and expression profiles of Annexin family (ANXA1~A6) in yellow catfish (Pelteobagrus fulvidraco) and ANX regulation by CpG ODN responding to bacterial infection.

Up to now, many previous reports have emphasized that Annexins (ANX) family played an important role in immune responses. Aeromonas hydrophila (A. hydrophila), the most common zoonotic pathogenic bacteria of yellow catfish (Pelteobagrus fulvidraco), can cause serious economic loss, especially to yellow catfish with high economic value. In our previous work, we demonstrated that synthetic oligodeoxynucleotides containing CpG motifs (CpG ODN) owned powerful immunostimulatory activity. However, the relationship among Pelteobagrus fulvidraco Annexins (Pf_ANX), CpG ODN and A. hydrophila is unknown. Therefore, we cloned Pf_ANX1-6 genes and analyzed its sequences, structures, genetic evolution, post-translation modifications (PTMs), Ca2+ ion binding sites and tissue distribution to reveal the relevance. In addition, we investigated the responses of ANXA1-6 and cytokines in intestine and spleen as well as morbidity/survival rate of fish post CpG ODN immunization and/or A. hydrophila infection. The results showed that compared with challenge alone (challenge-CK) group, the CpG immunization following challenge (CpG-challenge) group displayed relatively flat IL-1ß level throughout in both organs. Meanwhile, the expression of IFN-γ and morbidity/survival rate of fish in CpG-challenge group showed a great improvement compared with the challenge-CK group. Our results indicated that CpG ODN could improve morbidity/survival by up-regulating Pf_ANXA 1, 2 and 5 in the intestine and spleen to ameliorate inflammatory responses and promote anti-infective responses. Our findings offer some important insights into ANX related to the immunity of fish infection and lay a theoretical basis for the prevention and treatment of fish infections.

Effects of porcine IL-17B and IL-17E against intestinal pathogenic microorganism.

The interleukin-17 (IL-17) family plays a critical role in host defense, allergic reactions, and even tumorigenesis on different mucous membranes. IL-17 family has been cloned in human and mouse, as well IL-17A, IL-17 F in swine. So far, current knowledge on the cloning and biological functions of porcine IL-17B (poIL-17B) and porcine IL-17E (poIL-17E) is limited. In this study, poIL-17B and poIL-17E, mainly expressed in intestine, were cloned and characterized. Expression of poIL-17B and poIL-17E was upregulated after pathogenic microorganism infection. Moreover, the significant enhanced expression of antibacterial peptides PR-39 and pBD-1 was observed when poIL-17B and poIL-17E were over-expressed in the small intestinal epithelial cell line IPEC-J2. This demonstrated that poIL-17B and poIL-17E might have anti-infective capability. Pathogens infection data showed that pathogens could up-regulate poIL-17B/E expression levels. After stimulating the cells with the pathogen, continued with probiotics, the expression of poIL-17B/E was down-regulated. Meanwhile, the induced expression of poIL-17E was greater than that of poIL-17B. Invasion data indicated that poIL-17B and poIL-17E both could inhibit effectively pathogenic microorganism, while inhibitory capability of poIL-17B was stronger than that of poIL-17E. Therefore, poIL-17B and poIL-17E both could be important members against intestinal infection in the porcine IL-17 family. This study provided a theoretical basis for the prevention of intestinal diseases in pigs and thus achieved healthy farming.

Recombinant plasmids containing CpG with porcine host defense peptides (PR-39/pBD-1) modulates the innate and adaptive intestinal immune responses (including maternal-derived) in piglets.

CpG oligodeoxynucleotides (CpG-ODN) is an immunoenhancer, which is composed of unmethylated cytosine and guanine. Host Defense Peptides (HDPs) are small molecule polypeptides with various immunological activities that have been shown to induce a stronger innate immune response in piglets with synthetic CpG-ODN. Therefore, combination of CpG-ODN and HDPs was expected to be a novel immunoadjuvant with high efficiency, low toxicity and great potential. However, cost of synthetic HDPs or CpG-ODN is too high to be advantageous for animal farming. In this study, in order to improve the immune function of vaccine and reduce cost, a series of recombinant plasmids (containing HDPs gene (PR-39/pBD-1) and different numbers of CpG motifs) were constructed. In vitro, porcine lymphocytes were stimulated by recombinant plasmids to verify the immunostimulatory function of recombinant plasmids. In vivo, recombinant plasmids were used to immunize piglets with Enterotoxigenic Escherichia coli (ETEC) vaccine to analyze effects of recombinant plasmids on the mucosal immune responses. In addition, dosage screening and capability of maternal antibody responses were also investigated. Our results showed that recombinant plasmids had strong adjuvant effects especially the plasmid pVAX49-PR-39 and pVAX49-pBD-1. Moreover, there was no diarrhea in piglets using pVAX49-PR-39 or pVAX49-pBD-1 as adjuvants. These findings suggested that recombinant plasmids (containing PR-39/pBD-1 and CpG) as adjuvants of vaccines could enhance immune stimulation better than HDPs or CpG alone. It has a good protective effect on maintaining health of newborn piglets. Among them, both plasmids pVAX49-PR-39 and pVAX49-pBD-1 could be used as effective vaccine adjuvants for piglets.

Modulating the 3' end-DNA and the fermentation process for enhanced production and biological activity of porcine interferon-gamma.

Porcine gamma interferon is a cytokine produced by activated T cells and NK cells with broad-spectrum antiviral activity and immunomodulatory function. However, pIFN-γ is a secretory protein that has a short half-life in organisms and is easily inactivated, making it difficult to apply widely in clinics. Therefore, we tried to optimize the expression of pIFN-γ in Pichia pastoris to obtain a large amount of highly active, easily purified pIFN-γ protein in vitro. Through C-terminal sequence analysis, we found a signal sequence (EKREAEAE) that was easily enzymolysed by a signal peptide enzyme, resulting in degradation and inactivation of the pIFN-γ protein. In this study, we optimized the pIFN-γ gene recombination sequence and mutated the 3' end of the pIFN-γ gene, resulting in a higher expression level and stronger biological activity, as well as a significant upregulation in the expression of the interferon-stimulated genes Mx1 and OAS1 in IPEC-J2 jejunal epithelial cells. Our data also showed that the fermentation process could significantly improve productivity. A recombinant Pichia pastoris strain with the optimized pIFN-γ gene could obtain a high yield of pIFN-γ protein, up to 9536 mg/L, after staged incubation for 0-24 h at 28°C, pH 6.0, and 50% dissolved oxygen (DO), followed by incubation for 24-72 h at 25°C, pH 6.0 and 30% DO. These data demonstrated, for the first time, that the expression level of pIFN-γ in Pichia pastoris was improved significantly by gene optimization with 3' end mutation and a fermentation process that maintained good biological activity, which is beneficial to the application of pIFN-γ in animal husbandry.

CRIP1, a novel immune-related protein, activated by Enterococcus faecalis in porcine gastrointestinal epithelial cells.

Cysteine-rich intestinal protein 1 (CRIP1) is an important transcriptional regulation factor during the tumor development. Although it was largely studied in the human or mouse, no report has provided functional evidence for it in the swine. To date, the real sequence of porcine CRIP1 (poCRIP1) was also still unknown. In this study, clear characteristics for the poCRIP1 were represented. A 552bp poCRIP1 cDNA was obtained from porcine brain tissue using real time reverse transcriptase PCR. The poCRIP1 showed 89% and 93% homologous with human and cattle, respectively. And it also contained one conserved domain, LIM-CRIP domain. Meanwhile, the genomic structure and promoter map was done and several conserved transcriptional regulatory sites were also predicted in this study. The expression pattern of poCRIP1 indicated that poCRIP1 is expressed in mucosal tissue. An infection experiment about the gut was designed to analyze whether or not poCRIP1 was functional in gut immunity, and an interesting result was that poCRIP1 was only activated by an opportunistic pathogen, Enterococcus faecalis FA2-2. It was the first report to identify the full-length sequence of poCRIP1 gene, represent a clear characteristic and immunologic role of CRIP1 in domestic animal until now.

Effect of cooperation of chaperones and gene dosage on the expression of porcine PGLYRP-1 in Pichia pastoris.

Mammalian peptidoglycan recognition proteins (PGLYRPs) are highly conserved pattern-recognition molecules of the innate immune system with considerable bactericidal activity, which manifest their potential values for the application to food and pharmaceutical industry. However, the effective expression of porcine PGLYRP-1 in Pichia pastoris has not been reported so far. In this study, expression in P. pastoris was explored as an efficient way to produce functional porcine PGLYRP-1. Cooperation of chaperones co-expression and gene dosage (including protein disulfide isomerase (PDI)/binding protein (BiP) and pglyrp-1) were used to enhance functional expression of antimicrobial protein in P. pastoris. Overexpression of PDI was certainly able to increase secretion level of PGLYRP-1 protein because the increase in secreted PGLYRP-1 secretion was correlated with the copy numbers of PDI in high copy pglyrp-1 clones. However, co-expression of BiP was proved to be detrimental to PGLYRP-1 secretion. In addition, we also found that excessive expression of PDI and/or BiP could decrease the mRNA expression of pglyrp-1 gene. This showed that PDI and BiP as the target genes of unfolded protein response (UPR) might regulate the transcription of the target protein. These data demonstrated for the first time that the combination of chaperones and gene dosages could improve the yield of PGLYRP-1, which could facilitate the application to food and pharmaceutical industry.

CpG oligodeoxynucleotide protect neonatal piglets from challenge with the enterotoxigenic E. coli.

CpG motifs activates mammalian lymphocytes and macrophages to produce cytokines and polyclonal Ig. These include IFN-γ, IL-12, TNF-a, which are important in the control of bacterial infection. But thus far, the innate immunostimulatory effects of CpG ODN against pathogen have been established mainly in mouse, monkey, sheep, chicken, but not in neonatal piglets. The purpose of this study is to determine the potential protection of CpG ODN against enterotoxigenic Escherichia coli (ETEC) (with which neonatal piglets were susceptible to infection in our lab) in neonatal piglets. Here, we show intranasal (IN)-mucosal and intramuscularly (IM) systemic administration of CpG ODN could enhance innate cellular (cytokine) immunity in the sera and intestine mucosa post challenge, and thereafter the development of antigen-specific antibodies in piglets. IN and IM immunizations of neonatal piglets without antigen both reduced the ETEC excretion and alleviated diarrhoea symptoms upon challenge, and IN route had better protection effects than IM route. Protection in this study was linked to induction of a Th1 response which induced by CpG ODN. Co-delivery with Emulsigen (EM), could improve protection mediated by CpG ODN. These observations indicate that IN administration of 100 µg/kg CpG ODN with 20% EM codelivery may represent a valuable strategy for induction of innate immunity against ETEC infection in neonatal piglets.

Immunization of aged pigs with attenuated pseudorabies virus vaccine combined with CpG oligodeoxynucleotide restores defective Th1 immune responses.

BACKGROUND AND AIMS: Attempts to immunize aged subjects often result in the failure to elicit a protective immune response. Murine model studies have shown that oligonucleotides containing CpG motifs (CpG ODN) can stimulate immune system in aged mice as effectively as in young mice. Since many physiological and pathophysiological data of pigs can be transferred to humans, research in pigs is important to confirm murine data. Here we investigated whether immunization of aged pig model with attenuated pseudorabies virus vaccine (PRV vaccine) formulated with CpG ODN could promote a successful development of immune responses that were comparable to those induced in young pigs in a similar manner. METHODOLOGY: Young and aged pigs were immunized IM with PRV vaccine alone, or in combination with CpG ODN respectively. At days 3, 7, 14 post immunization sera were assayed by ELISA for IgG titres, at day 7 for IgG1 and IgG2 subtypes titres. All blood samples collected in evacuated test tubes with K-EDTA at day 7 were analyzed for flow cytometer assay. Blood samples at day 7 collected in evacuated test tubes with heparin were analysed for antigen-specific cytokines production and peripheral blood mononuclear cells (PBMCs) proliferative responses. RESULTS: CpG ODN could enhance Th1 responses (PRV-specific IgG2/IgG1 ratio, proliferative responses, Th1 cytokines production) when used as an adjuvant for the vaccination of aged pigs, which were correlated with enhanced CD4+ T cells percentage, decreased CD4+CD8+CD45RO+ T cells percentage and improved PRV-specific CD4+ T cells activation. CONCLUSIONS: Our results demonstrate a utility for CpG ODN, as a safe vaccine adjuvant for promoting effective systemic immune responses in aged pig model. This agent could have important clinical uses in overcoming some of age-associated depressions in immune function that occur in response to vaccination.

[Protective effects of micro-encapsulated Bifidobacteria on gut barrier after hemorrhagic shock and resuscitation: experiment with rats].

OBJECTIVE: To investigate the effects of micro-encapsulated bifidobacteria on gut barrier and bacterial translocation after hemorrhagic shock and resuscitation. METHODS: Sprague-Dawley rats were divided into 6 groups: PBS+sham shock group fed with PBS for 7 days and then undergoing sham shock, bifidobacteria+sham shock group fed with bifidobacteria (10(9) cfu/d) for 7 days and then undergoing sham shock, micro-encapsulated bifidobacteria+sham shock group, fed with micro-encapsulated bifidobacteria (10(9) cfu/d) for 7 days and then undergoing sham shock, PBS+hemorrhagic shock group fed with PBS for 7 days and then undergoing hemorrhagic shock, bifidobacteria+shock group fed with bifidobacteria for 7 days and then undergoing hemorrhagic shock, and micro-encapsulated bifidobacteria+shock group, fed with micro-encapsulated bifidobacteria for 7 days and then undergoing hemorrhagic shock. Three hours after resuscitation laparotomy was performed, distal cecum was resected to undergo bacteriological analysis of the cecal content, mesenteric lymph nodes (MLNs), a liver lobe, and the middle part of spleen were resected to undergo bacterial culture for bacterial translocation, and the terminal ileum was resected to observe the villous damage. RESULTS: There was no significant difference in the amount of blood loss among the 3 hemorrhagic shock groups. The amounts of aerobes in cecum of the bifidobacteria+shock and micro-encapsulated bifidobacteria+shock groups, especially that of the latter group, were significantly lower than that of the PBS+shock group. The amounts of anaerobes and the amounts of bifidobacteria in cecum of the bifidobacteria+shock group and micro-encapsulated bifidobacteria+shock group, especially those of the latter group, were significantly higher than those of the PBS+shock group. No bacterial translocation to liver was observed in all groups. The magnitudes of total aerobes translocation in spleen of the bifidobacteria+shock and encapsulated bifidobacteria+shock groups were significantly lower than that of the PBS+shock group, however, there were not significant differences in the translocation in the MLN of total aerobes ad bifidobacteria among different groups. The percentage of ileal villous damage of the bifidobacteria+shock and encapsulated bifidobacteria+shock groups were significantly lower than that of the PBS+shock group. CONCLUSION: Bifidobacteria effectively protects the gut barrier, reduces bacterial translocation from the gut after hemorrhagic shock and resuscitation. And micro-encapsulated Bifidobacteria can enhance those effects further.

Encapsulated Bifidobacteria reduced bacterial translocation in rats following hemorrhagic shock and resuscitation.

OBJECTIVE: The aim of the present study was to determine the effects of peroral encapsulated Bifidobacteria on intestinal microflora, bacterial translocation (BT), plasma endotoxin, and ileal villi injury in a rat model of hemorrhagic shock. METHODS: Sprague-Dawley rats were fed daily with three different diet supplements: phosphate buffered saline, Bifidobacteria (10(9) colon-forming units/day), or microencapsulated Bifidobacteria (10(9) colony-forming units/day). After 7 d of treatment, rats were anesthetized for hemorrhagic or sham shock. Then a laparotomy was performed to determine microbiological analysis of cecal content, BT to mesenteric lymph nodes, plasma endotoxin, and terminal ileal villous damage. RESULTS: In the hemorrhagic-shock model, rats pretreated with Bifidobacteria showed decreases in total aerobes in cecum, magnitude of total aerobes to BT, levels of plasma endotoxin, and percentage of ileal villous damage when compared with rats treated with phosphate buffered saline. Encapsulated Bifidobacteria induced greater decreases than intact Bifidobacteria in this model, except for no difference in percentage of ileal villous damage between the two groups. In addition, the incidence of BT was decreased in hemorrhagic rats pretreated with Bifidobacteria compared with control. However, the magnitude of total anaerobes and Bifidobacteria BT were similar among hemorrhagic-shocked rats receiving three different supplements. CONCLUSION: Bifidobacteria can be useful in preventing BT in hemorrhagic-shocked rats, and encapsulated Bifidobacteria can augment this effect further. Peroral administration of Bifidobacteria may be a favorable strategy to prevent sepsis and multiple organ dysfunction syndrome in hemorrhagic shock.