8ae2eca0be1e4fac57f2ac90cd5e7f71 pmc_9101217.pdf 448d0b4afba268a3c41eac5c8994f5ee5ef3b0a0 pmc_9101217.pdf f0239c196f8bbd7bdb2e519996ff6712c3d39e8d9a1bab500f31b1e0bc4357a6 pmc_9101217.pdf Title: Potential of Oral Nanoparticles Containing Cytokines as Intestinal Mucosal Immunostimulants in Pigs: A Pilot Study Subject: Antimicrobial resistance is a global threat that is worryingly rising in the livestock sector. Among the proposed strategies, immunostimulant development appears an interesting approach to increase animal resilience at critical production points. The use of nanoparticles based on cytokine aggregates, called inclusion bodies (IBs), has been demonstrated as a new source of immunostimulants in aquaculture. Aiming to go a step further, the objective of this study was to produce cytokine nanoparticles using a food-grade microorganism and to test their applicability to stimulate intestinal mucosa in swine. Four cytokines (IL-1, IL-6, IL-8, and TNF-) involved in inflammatory response were produced recombinantly in Lactococcus lactis in the form of protein nanoparticles (IBs). They were able to stimulate inflammatory responses in a porcine enterocyte cell line (IPEC-J2) and alveolar macrophages, maintaining high stability at low pH and high temperature. In addition, an in vivo assay was conducted involving 20 piglets housed individually as a preliminary exploration of the potential effects of IL-1 nanoparticles in piglet intestinal mucosa after a 7 d oral administration. The treated animals tended to have greater levels of TNF- in the blood, indicating that the tested dose of nanoparticles tended to generate an inflammatory response in the animals. Whether this response is sufficient to increase animal resilience needs further evaluation. Keywords: immunostimulant; cytokines; nanoparticles; piglets; antimicrobial resistance Author: Adrià López-Cano, Alex Bach, Sergi López-Serrano, Virginia Aragon, Marta Blanch, Jose J. Pastor, Gemma Tedó, Sofia Morais, Elena Garcia-Fruitós and Anna Arís Creator: LaTeX with hyperref Producer: pdfTeX-1.40.21 CreationDate: Thu Apr 21 10:52:25 2022 CEST ModDate: Thu Apr 21 10:59:12 2022 CEST Tagged: no UserProperties: no Suspects: no Form: none JavaScript: no Pages: 14 Encrypted: no Page size: 595.276 x 841.89 pts (A4) Page rot: 0 File size: 1326526 bytes Optimized: no PDF version: 1.7 name type encoding emb sub uni object ID ------------------------------------ ----------------- ---------------- --- --- --- --------- OHKNFJ+VnURWPalladioL Type 1 Custom yes yes no 10 0 LEVXPH+EURM10 Type 1 Builtin yes yes no 15 0 FBXDNR+URWPalladioL-Bold Type 1 Custom yes yes no 19 0 PSMMWL+URWPalladioL-Roma Type 1 Custom yes yes no 24 0 GETDJI+URWPalladioL-Ital Type 1 Custom yes yes no 28 0 RSYDPD+VnURWPalladioL-Bold Type 1 Custom yes yes no 32 0 SZMRXJ+TimesNewRomanPS-BoldItalicMT Type 1C WinAnsi yes yes no 50 0 IFEDSM+CMSY10 Type 1 Builtin yes yes no 72 0 SPXLNH+PazoMath Type 1 Builtin yes yes no 79 0 GIZSCG+EURM7 Type 1 Builtin yes yes no 86 0 FHBIAZ+EURB7 Type 1 Builtin yes yes no 90 0 DUJUUF+PazoMath-Italic Type 1 Builtin yes yes no 94 0 MUGGGS+EURB10 Type 1 Builtin yes yes no 107 0 WBVKDJ+URWPalladioL-BoldItal Type 1 Custom yes yes no 115 0 BCDEEE+PalatinoLinotype-Bold TrueType WinAnsi yes yes no 123 0 BCDFEE+PalatinoLinotype-Roman TrueType WinAnsi yes yes no 127 0 BCDGEE+PalatinoLinotype-Italic TrueType WinAnsi yes yes no 131 0 BCDIEE+PalatinoLinotype-Roman CID TrueType Identity-H yes yes yes 135 0 BCDKEE+PalatinoLinotype-Italic CID TrueType Identity-H yes yes yes 142 0 FPTLHW+PalatinoLinotype,Italic CID TrueType Identity-H yes yes yes 159 0 FPTLHW+PalatinoLinotype,Bold CID TrueType Identity-H yes yes yes 165 0 FPWNMN+PalatinoLinotype CID TrueType Identity-H yes yes yes 171 0 BCDEEE+PalatinoLinotype-Bold TrueType WinAnsi yes yes no 195 0 BCDFEE+PalatinoLinotype-Roman TrueType WinAnsi yes yes no 199 0 BCDGEE+PalatinoLinotype-Italic TrueType WinAnsi yes yes no 203 0 BCDIEE+PalatinoLinotype-Roman CID TrueType Identity-H yes yes yes 207 0 Jhove (Rel. 1.22.1, 2019-04-17) Date: 2023-01-17 02:35:15 CET RepresentationInformation: pmc_9101217.pdf ReportingModule: PDF-hul, Rel. 1.12.1 (2019-04-17) LastModified: 2022-12-14 02:18:47 CET Size: 1326526 Format: PDF Version: 1.7 Status: Well-Formed and valid SignatureMatches: PDF-hul MIMEtype: application/pdf PDFMetadata: Objects: 372 FreeObjects: 1 IncrementalUpdates: 0 DocumentCatalog: PageLayout: SinglePage PageMode: UseNone Outlines: Item: Title: Introduction Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@4923ab24 Item: Title: Materials and Methods Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@44c8afef Children: Item: Title: Bacterial and Culture Strains Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@7b69c6ba Item: Title: Genetic Construct Design Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@46daef40 Item: Title: Cytokine Nanoparticle Production and Purification Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@12f41634 Item: Title: Immunoassays Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@13c27452 Item: Title: Gene Expression Analyses Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@262b2c86 Item: Title: Enzyme-Linked Immunosorbent Assay (ELISA) Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@371a67ec Item: Title: Temperature and pH IB Stability Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@5ed828d Item: Title: In Vivo Assay Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@50d0686 Item: Title: Statistical Analysis Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@7a3d45bd Item: Title: Results Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@1e7c7811 Children: Item: Title: Production and Characterization of Cytokine-Based Nanoparticles Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@77ec78b9 Item: Title: Immunostimulation of Swine Intestinal Cells and Macrophages Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@1a3869f4 Item: Title: Temperature and pH Stability of Cytokine Nanoparticles Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@a38d7a3 Item: Title: Swine In Vivo Experiments Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@77f99a05 Item: Title: Discussion Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@63440df3 Item: Title: Conclusions Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@3aeaafa6 Item: Title: References Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@76a3e297 Info: Title: Potential of Oral Nanoparticles Containing Cytokines as Intestinal Mucosal Immunostimulants in Pigs: A Pilot Study Author: Adrià López-Cano, Alex Bach, Sergi López-Serrano, Virginia Aragon, Marta Blanch, Jose J. Pastor, Gemma Tedó, Sofia Morais, Elena Garcia-Fruitós and Anna Arís Subject: Antimicrobial resistance is a global threat that is worryingly rising in the livestock sector. Among the proposed strategies, immunostimulant development appears an interesting approach to increase animal resilience at critical production points. The use of nanoparticles based on cytokine aggregates, called inclusion bodies (IBs), has been demonstrated as a new source of immunostimulants in aquaculture. Aiming to go a step further, the objective of this study was to produce cytokine nanoparticles using a food-grade microorganism and to test their applicability to stimulate intestinal mucosa in swine. Four cytokines (IL-1, IL-6, IL-8, and TNF-) involved in inflammatory response were produced recombinantly in Lactococcus lactis in the form of protein nanoparticles (IBs). They were able to stimulate inflammatory responses in a porcine enterocyte cell line (IPEC-J2) and alveolar macrophages, maintaining high stability at low pH and high temperature. In addition, an in vivo assay was conducted involving 20 piglets housed individually as a preliminary exploration of the potential effects of IL-1 nanoparticles in piglet intestinal mucosa after a 7 d oral administration. The treated animals tended to have greater levels of TNF- in the blood, indicating that the tested dose of nanoparticles tended to generate an inflammatory response in the animals. Whether this response is sufficient to increase animal resilience needs further evaluation. 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