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Incunabular immunological events in prion trafficking.

Michel B, Meyerett-Reid C, Johnson T, Ferguson A, Wyckoff C, Pulford B, Bender H, Avery A, Telling G, Dow S, Zabel MD - Sci Rep (2012)

Bottom Line: Monocytes and dendritic cells (DCs) required Complement for optimal prion delivery to lymph nodes hours later in a second wave of prion trafficking.B cells constituted the majority of prion-bearing cells in the mediastinal lymph node by six hours, indicating intranodal prion reception from resident DCs or subcapsulary sinus macrophages or directly from follicular conduits.These data reveal novel, cell autonomous prion lymphotropism, and a prominent role for B cells in intranodal prion movement.

View Article: PubMed Central - PubMed

Affiliation: College of Veterinary Medicine and Biomedical Sciences, Department of Microbiology, Immunology and Pathology, Prion Research Program at Colorado State University, Fort Collins, Colorado 80521, USA.

ABSTRACT
While prions probably interact with the innate immune system immediately following infection, little is known about this initial confrontation. Here we investigated incunabular events in lymphotropic and intranodal prion trafficking by following highly enriched, fluorescent prions from infection sites to draining lymph nodes. We detected biphasic lymphotropic transport of prions from the initial entry site upon peripheral prion inoculation. Prions arrived in draining lymph nodes cell autonomously within two hours of intraperitoneal administration. Monocytes and dendritic cells (DCs) required Complement for optimal prion delivery to lymph nodes hours later in a second wave of prion trafficking. B cells constituted the majority of prion-bearing cells in the mediastinal lymph node by six hours, indicating intranodal prion reception from resident DCs or subcapsulary sinus macrophages or directly from follicular conduits. These data reveal novel, cell autonomous prion lymphotropism, and a prominent role for B cells in intranodal prion movement.

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Related in: MedlinePlus

Flow cytometric analysis of immune cells trafficking prions from the PC to MedLN 2 HPI.PBS (rows a–g, v-ab, ac-ai, and ax-bd), fluorescent beads (h–n, v-ab, aj-ap, and ax-bd) or prion rods (o–u, v-ab, aq-aw, ax-bd) were injected into the PC of mice and cells harvested from peritoneal lavage fluid (a-ab) or mediastinal lymph nodes (ac-bd) two hours later. Graphs in the first column show cells from mice treated with PBS (panels a, v ac, and ax), fluorescent beads (h, v, aj, and ax) and prion rods (panels o, v, aq, and ax). Fluorescent cells (red or green dots) and total cells (grey dots) are plotted to show relative size (forward scatter, linear scale), granularity (side scatter, log scale) and proportion of total live cells that fluoresce. Cells were also stained with antibodies against immune cell surface markers and gated for SSChiLy6G- Ly6C+CD11b+ monocytes (panels b, i, p, w, ad, ak, ar, and ay), SSChiLy6G+ Ly6C-CD11c- CD11b+ neutrophils (c, j, q, x, ae, al, as, and az), SSChiLy6G- Ly6C-CD11b+CD11c+ DCs (d, k, r, y, af, am, at, ba), SSChiLy6G-Ly6C-CD11c-CD11b+ MΦs, (e, l, s, z, ag, an, au, and b), SSCloB220+CD21+CD3- B cells (f, m, t, aa, ah, ao, av, and bc) and SSCloCD21-B220- CD3+ T cells (g, n, u, ab, ai, ap, aw, and bd). Cell counts in graphs are shown as log10 per 105 total cells. Horizontal bars below the PC and MedLN panels represent relative proportions of prion-bearing monocytes (red stripe), neutrophils (solid red), DCs (dotted), MΦs (checkered), B cells (white) and T cells (black).
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f3: Flow cytometric analysis of immune cells trafficking prions from the PC to MedLN 2 HPI.PBS (rows a–g, v-ab, ac-ai, and ax-bd), fluorescent beads (h–n, v-ab, aj-ap, and ax-bd) or prion rods (o–u, v-ab, aq-aw, ax-bd) were injected into the PC of mice and cells harvested from peritoneal lavage fluid (a-ab) or mediastinal lymph nodes (ac-bd) two hours later. Graphs in the first column show cells from mice treated with PBS (panels a, v ac, and ax), fluorescent beads (h, v, aj, and ax) and prion rods (panels o, v, aq, and ax). Fluorescent cells (red or green dots) and total cells (grey dots) are plotted to show relative size (forward scatter, linear scale), granularity (side scatter, log scale) and proportion of total live cells that fluoresce. Cells were also stained with antibodies against immune cell surface markers and gated for SSChiLy6G- Ly6C+CD11b+ monocytes (panels b, i, p, w, ad, ak, ar, and ay), SSChiLy6G+ Ly6C-CD11c- CD11b+ neutrophils (c, j, q, x, ae, al, as, and az), SSChiLy6G- Ly6C-CD11b+CD11c+ DCs (d, k, r, y, af, am, at, ba), SSChiLy6G-Ly6C-CD11c-CD11b+ MΦs, (e, l, s, z, ag, an, au, and b), SSCloB220+CD21+CD3- B cells (f, m, t, aa, ah, ao, av, and bc) and SSCloCD21-B220- CD3+ T cells (g, n, u, ab, ai, ap, aw, and bd). Cell counts in graphs are shown as log10 per 105 total cells. Horizontal bars below the PC and MedLN panels represent relative proportions of prion-bearing monocytes (red stripe), neutrophils (solid red), DCs (dotted), MΦs (checkered), B cells (white) and T cells (black).

Mentions: Particulate antigen can be transported to SLOs via cell dependent and independent mechanisms1011. The presence of prions, but not beads, in various locations consistent with lymph nodes that drain the injection site suggested active uptake and trafficking by migratory immune cells. To investigate this possibility, we first assessed whether immune cells associated with fluorescent prions hours after injection. Isolation of adequate cell numbers from subcutaneous tissue and the oral cavity proved difficult for characterizing innate immune cells at these injection sites. We therefore chose the peritoneal cavity (PC) as an alternative inoculation route because it provides an optimal site for collecting immune cells. We inoculated mice intraperitoneally (IP) with 5 µg of fluorescent prions, collected cells by peritoneal lavage 2 HPI and analyzed them by flow cytometry (Figures 2 and 3).


Incunabular immunological events in prion trafficking.

Michel B, Meyerett-Reid C, Johnson T, Ferguson A, Wyckoff C, Pulford B, Bender H, Avery A, Telling G, Dow S, Zabel MD - Sci Rep (2012)

Flow cytometric analysis of immune cells trafficking prions from the PC to MedLN 2 HPI.PBS (rows a–g, v-ab, ac-ai, and ax-bd), fluorescent beads (h–n, v-ab, aj-ap, and ax-bd) or prion rods (o–u, v-ab, aq-aw, ax-bd) were injected into the PC of mice and cells harvested from peritoneal lavage fluid (a-ab) or mediastinal lymph nodes (ac-bd) two hours later. Graphs in the first column show cells from mice treated with PBS (panels a, v ac, and ax), fluorescent beads (h, v, aj, and ax) and prion rods (panels o, v, aq, and ax). Fluorescent cells (red or green dots) and total cells (grey dots) are plotted to show relative size (forward scatter, linear scale), granularity (side scatter, log scale) and proportion of total live cells that fluoresce. Cells were also stained with antibodies against immune cell surface markers and gated for SSChiLy6G- Ly6C+CD11b+ monocytes (panels b, i, p, w, ad, ak, ar, and ay), SSChiLy6G+ Ly6C-CD11c- CD11b+ neutrophils (c, j, q, x, ae, al, as, and az), SSChiLy6G- Ly6C-CD11b+CD11c+ DCs (d, k, r, y, af, am, at, ba), SSChiLy6G-Ly6C-CD11c-CD11b+ MΦs, (e, l, s, z, ag, an, au, and b), SSCloB220+CD21+CD3- B cells (f, m, t, aa, ah, ao, av, and bc) and SSCloCD21-B220- CD3+ T cells (g, n, u, ab, ai, ap, aw, and bd). Cell counts in graphs are shown as log10 per 105 total cells. Horizontal bars below the PC and MedLN panels represent relative proportions of prion-bearing monocytes (red stripe), neutrophils (solid red), DCs (dotted), MΦs (checkered), B cells (white) and T cells (black).
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC3368226&req=5

f3: Flow cytometric analysis of immune cells trafficking prions from the PC to MedLN 2 HPI.PBS (rows a–g, v-ab, ac-ai, and ax-bd), fluorescent beads (h–n, v-ab, aj-ap, and ax-bd) or prion rods (o–u, v-ab, aq-aw, ax-bd) were injected into the PC of mice and cells harvested from peritoneal lavage fluid (a-ab) or mediastinal lymph nodes (ac-bd) two hours later. Graphs in the first column show cells from mice treated with PBS (panels a, v ac, and ax), fluorescent beads (h, v, aj, and ax) and prion rods (panels o, v, aq, and ax). Fluorescent cells (red or green dots) and total cells (grey dots) are plotted to show relative size (forward scatter, linear scale), granularity (side scatter, log scale) and proportion of total live cells that fluoresce. Cells were also stained with antibodies against immune cell surface markers and gated for SSChiLy6G- Ly6C+CD11b+ monocytes (panels b, i, p, w, ad, ak, ar, and ay), SSChiLy6G+ Ly6C-CD11c- CD11b+ neutrophils (c, j, q, x, ae, al, as, and az), SSChiLy6G- Ly6C-CD11b+CD11c+ DCs (d, k, r, y, af, am, at, ba), SSChiLy6G-Ly6C-CD11c-CD11b+ MΦs, (e, l, s, z, ag, an, au, and b), SSCloB220+CD21+CD3- B cells (f, m, t, aa, ah, ao, av, and bc) and SSCloCD21-B220- CD3+ T cells (g, n, u, ab, ai, ap, aw, and bd). Cell counts in graphs are shown as log10 per 105 total cells. Horizontal bars below the PC and MedLN panels represent relative proportions of prion-bearing monocytes (red stripe), neutrophils (solid red), DCs (dotted), MΦs (checkered), B cells (white) and T cells (black).
Mentions: Particulate antigen can be transported to SLOs via cell dependent and independent mechanisms1011. The presence of prions, but not beads, in various locations consistent with lymph nodes that drain the injection site suggested active uptake and trafficking by migratory immune cells. To investigate this possibility, we first assessed whether immune cells associated with fluorescent prions hours after injection. Isolation of adequate cell numbers from subcutaneous tissue and the oral cavity proved difficult for characterizing innate immune cells at these injection sites. We therefore chose the peritoneal cavity (PC) as an alternative inoculation route because it provides an optimal site for collecting immune cells. We inoculated mice intraperitoneally (IP) with 5 µg of fluorescent prions, collected cells by peritoneal lavage 2 HPI and analyzed them by flow cytometry (Figures 2 and 3).

Bottom Line: Monocytes and dendritic cells (DCs) required Complement for optimal prion delivery to lymph nodes hours later in a second wave of prion trafficking.B cells constituted the majority of prion-bearing cells in the mediastinal lymph node by six hours, indicating intranodal prion reception from resident DCs or subcapsulary sinus macrophages or directly from follicular conduits.These data reveal novel, cell autonomous prion lymphotropism, and a prominent role for B cells in intranodal prion movement.

View Article: PubMed Central - PubMed

Affiliation: College of Veterinary Medicine and Biomedical Sciences, Department of Microbiology, Immunology and Pathology, Prion Research Program at Colorado State University, Fort Collins, Colorado 80521, USA.

ABSTRACT
While prions probably interact with the innate immune system immediately following infection, little is known about this initial confrontation. Here we investigated incunabular events in lymphotropic and intranodal prion trafficking by following highly enriched, fluorescent prions from infection sites to draining lymph nodes. We detected biphasic lymphotropic transport of prions from the initial entry site upon peripheral prion inoculation. Prions arrived in draining lymph nodes cell autonomously within two hours of intraperitoneal administration. Monocytes and dendritic cells (DCs) required Complement for optimal prion delivery to lymph nodes hours later in a second wave of prion trafficking. B cells constituted the majority of prion-bearing cells in the mediastinal lymph node by six hours, indicating intranodal prion reception from resident DCs or subcapsulary sinus macrophages or directly from follicular conduits. These data reveal novel, cell autonomous prion lymphotropism, and a prominent role for B cells in intranodal prion movement.

Show MeSH
Related in: MedlinePlus