Limits...
Tumor necrosis factor-dependent segmental control of MIG expression by high endothelial venules in inflamed lymph nodes regulates monocyte recruitment.

Janatpour MJ, Hudak S, Sathe M, Sedgwick JD, McEvoy LM - J. Exp. Med. (2001)

Bottom Line: Quantitative PCR analyses revealed the upregulation of many chemokines in the inflamed lymph node, including MCP-1 and MIG.HEVs did not express detectable levels of MCP-1; however, a subset of HEVs in inflamed lymph nodes in wild-type (but not tumor necrosis factor [TNF] mice) expressed MIG and this subset of HEVs preferentially supported monocyte binding.Together, these results suggest that the lymph node microenvironment can dictate the nature of molecules expressed on HEV subsets in a TNF-dependent fashion and that inflammation-induced MIG expression by HEVs can mediate monocyte recruitment.

View Article: PubMed Central - PubMed

Affiliation: DNAX Research Institute, Inc., Palo Alto, CA 94304, USA.

ABSTRACT
Monocytes recruited from the blood are key contributors to the nature of an immune response. While monocyte recruitment in a subset of immunopathologies has been well studied and largely attributed to the chemokine monocyte chemoattractant protein (MCP)-1, mechanisms mediating such recruitment to other sites of inflammation remain elusive. Here, we showed that localized inflammation resulted in an increased binding of monocytes to perifollicular high endothelial venules (HEVs) of lymph nodes draining a local inflammatory site. Quantitative PCR analyses revealed the upregulation of many chemokines in the inflamed lymph node, including MCP-1 and MIG. HEVs did not express detectable levels of MCP-1; however, a subset of HEVs in inflamed lymph nodes in wild-type (but not tumor necrosis factor [TNF] mice) expressed MIG and this subset of HEVs preferentially supported monocyte binding. Expression of CXCR3, the receptor for MIG, was detected on a small subset of peripheral blood monocytes and on a significant percentage of recruited monocytes. Most importantly, in both ex vivo and in vivo assays, neutralizing anti-MIG antibodies blocked monocyte binding to inflamed lymph node HEVs. Together, these results suggest that the lymph node microenvironment can dictate the nature of molecules expressed on HEV subsets in a TNF-dependent fashion and that inflammation-induced MIG expression by HEVs can mediate monocyte recruitment.

Show MeSH

Related in: MedlinePlus

MIG expression on HEVs is correlated with increased monocyte-selective recruitment. (A) Serial sections of lymph nodes draining inflamed footpads were stained with antibodies against PNAd (top left panel, green) to identify HEVs, IP10 (top right panel, green), MCP-1 (bottom left panel, green), or an isotype control (bottom right panel, green). On all sections, an antibody against the B-cell marker B220 (red) identified the follicles and was used for orientation. MCP-1 and IP10 was not displayed on HEVs, but rather in macrophage-rich areas. (B) Serial sections (left and middle panels) were stained with either antibodies against PNAd (left panel, green) and B220 (left panel, red) or MIG (middle panel, green) and 6CKine (middle panel, red). MIG was expressed on a subset of 6CKine+ HEVs. White arrows pair vessels between left and middle panels; yellow arrow indicates a vessel that is absent in the section represented in the middle panel. The right panel depicts vessels that were stained in vitro for 6CKine (red) and MIG (green). The outline depicts the B cell follicle border. Arrows depict vessels in which expression is completely colocalized (white) or partially colocalized (yellow). (C) Serial sections of lymph nodes draining inflamed footpads were stained for PNAd (left panels, green) and CD11b (red) or MIG (right panels, green) and CD11b (red). MIG+ HEVs showed a higher association with monocytes than MIG− HEVs (arrows indicate MIG− vessels). (D) Serial sections were stained with antibodies against PNAd and MIG. The number of PNAd+ and MIG+ vessels was then counted across 10 lymph nodes. Data is presented as the percentage of total HEVs that were MIG+. Lymph nodes that were not inflamed (white bar) had <2% MIG+ HEVs. Upon inflammation (black bar), the number of MIG+ HEVs increased to ∼12%. This experiment was repeated twice. (E) Immunohistochemistry was performed as shown in C. All PNAd+ (total HEVs) and MIG+ vessels were counted and scored for whether a monocyte was bound. Data is presented as percentage of HEV+ for at least one bound monocyte. Upon inflammation (black bars) this increased from ∼6% in lymph nodes that were not inflamed (white bar) to ∼20%. The percentage of the subpopulation of total inflamed HEVs that were MIG+ and had a bound monocyte was >60%; whereas, the percentage of the MIG− HEV subpopulation that had a bound monocyte was only 13%. This experiment was repeated twice.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2195975&req=5

fig3: MIG expression on HEVs is correlated with increased monocyte-selective recruitment. (A) Serial sections of lymph nodes draining inflamed footpads were stained with antibodies against PNAd (top left panel, green) to identify HEVs, IP10 (top right panel, green), MCP-1 (bottom left panel, green), or an isotype control (bottom right panel, green). On all sections, an antibody against the B-cell marker B220 (red) identified the follicles and was used for orientation. MCP-1 and IP10 was not displayed on HEVs, but rather in macrophage-rich areas. (B) Serial sections (left and middle panels) were stained with either antibodies against PNAd (left panel, green) and B220 (left panel, red) or MIG (middle panel, green) and 6CKine (middle panel, red). MIG was expressed on a subset of 6CKine+ HEVs. White arrows pair vessels between left and middle panels; yellow arrow indicates a vessel that is absent in the section represented in the middle panel. The right panel depicts vessels that were stained in vitro for 6CKine (red) and MIG (green). The outline depicts the B cell follicle border. Arrows depict vessels in which expression is completely colocalized (white) or partially colocalized (yellow). (C) Serial sections of lymph nodes draining inflamed footpads were stained for PNAd (left panels, green) and CD11b (red) or MIG (right panels, green) and CD11b (red). MIG+ HEVs showed a higher association with monocytes than MIG− HEVs (arrows indicate MIG− vessels). (D) Serial sections were stained with antibodies against PNAd and MIG. The number of PNAd+ and MIG+ vessels was then counted across 10 lymph nodes. Data is presented as the percentage of total HEVs that were MIG+. Lymph nodes that were not inflamed (white bar) had <2% MIG+ HEVs. Upon inflammation (black bar), the number of MIG+ HEVs increased to ∼12%. This experiment was repeated twice. (E) Immunohistochemistry was performed as shown in C. All PNAd+ (total HEVs) and MIG+ vessels were counted and scored for whether a monocyte was bound. Data is presented as percentage of HEV+ for at least one bound monocyte. Upon inflammation (black bars) this increased from ∼6% in lymph nodes that were not inflamed (white bar) to ∼20%. The percentage of the subpopulation of total inflamed HEVs that were MIG+ and had a bound monocyte was >60%; whereas, the percentage of the MIG− HEV subpopulation that had a bound monocyte was only 13%. This experiment was repeated twice.

Mentions: Serial sections of inflamed lymph nodes were subjected to double-indirect immunohistochemistry (Fig. 3 A). On all sections, an antibody against the B cell marker B220 was used to illuminate the B cell follicles for orientation. Serial sections were stained with antibodies against PNAd to identify HEVs, IP10, MCP-1 or an isotype control. While both MCP-1 and IP10 were highly expressed in macrophage-rich areas, to our surprise neither was displayed on HEVs (Fig. 3 A).


Tumor necrosis factor-dependent segmental control of MIG expression by high endothelial venules in inflamed lymph nodes regulates monocyte recruitment.

Janatpour MJ, Hudak S, Sathe M, Sedgwick JD, McEvoy LM - J. Exp. Med. (2001)

MIG expression on HEVs is correlated with increased monocyte-selective recruitment. (A) Serial sections of lymph nodes draining inflamed footpads were stained with antibodies against PNAd (top left panel, green) to identify HEVs, IP10 (top right panel, green), MCP-1 (bottom left panel, green), or an isotype control (bottom right panel, green). On all sections, an antibody against the B-cell marker B220 (red) identified the follicles and was used for orientation. MCP-1 and IP10 was not displayed on HEVs, but rather in macrophage-rich areas. (B) Serial sections (left and middle panels) were stained with either antibodies against PNAd (left panel, green) and B220 (left panel, red) or MIG (middle panel, green) and 6CKine (middle panel, red). MIG was expressed on a subset of 6CKine+ HEVs. White arrows pair vessels between left and middle panels; yellow arrow indicates a vessel that is absent in the section represented in the middle panel. The right panel depicts vessels that were stained in vitro for 6CKine (red) and MIG (green). The outline depicts the B cell follicle border. Arrows depict vessels in which expression is completely colocalized (white) or partially colocalized (yellow). (C) Serial sections of lymph nodes draining inflamed footpads were stained for PNAd (left panels, green) and CD11b (red) or MIG (right panels, green) and CD11b (red). MIG+ HEVs showed a higher association with monocytes than MIG− HEVs (arrows indicate MIG− vessels). (D) Serial sections were stained with antibodies against PNAd and MIG. The number of PNAd+ and MIG+ vessels was then counted across 10 lymph nodes. Data is presented as the percentage of total HEVs that were MIG+. Lymph nodes that were not inflamed (white bar) had <2% MIG+ HEVs. Upon inflammation (black bar), the number of MIG+ HEVs increased to ∼12%. This experiment was repeated twice. (E) Immunohistochemistry was performed as shown in C. All PNAd+ (total HEVs) and MIG+ vessels were counted and scored for whether a monocyte was bound. Data is presented as percentage of HEV+ for at least one bound monocyte. Upon inflammation (black bars) this increased from ∼6% in lymph nodes that were not inflamed (white bar) to ∼20%. The percentage of the subpopulation of total inflamed HEVs that were MIG+ and had a bound monocyte was >60%; whereas, the percentage of the MIG− HEV subpopulation that had a bound monocyte was only 13%. This experiment was repeated twice.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2195975&req=5

fig3: MIG expression on HEVs is correlated with increased monocyte-selective recruitment. (A) Serial sections of lymph nodes draining inflamed footpads were stained with antibodies against PNAd (top left panel, green) to identify HEVs, IP10 (top right panel, green), MCP-1 (bottom left panel, green), or an isotype control (bottom right panel, green). On all sections, an antibody against the B-cell marker B220 (red) identified the follicles and was used for orientation. MCP-1 and IP10 was not displayed on HEVs, but rather in macrophage-rich areas. (B) Serial sections (left and middle panels) were stained with either antibodies against PNAd (left panel, green) and B220 (left panel, red) or MIG (middle panel, green) and 6CKine (middle panel, red). MIG was expressed on a subset of 6CKine+ HEVs. White arrows pair vessels between left and middle panels; yellow arrow indicates a vessel that is absent in the section represented in the middle panel. The right panel depicts vessels that were stained in vitro for 6CKine (red) and MIG (green). The outline depicts the B cell follicle border. Arrows depict vessels in which expression is completely colocalized (white) or partially colocalized (yellow). (C) Serial sections of lymph nodes draining inflamed footpads were stained for PNAd (left panels, green) and CD11b (red) or MIG (right panels, green) and CD11b (red). MIG+ HEVs showed a higher association with monocytes than MIG− HEVs (arrows indicate MIG− vessels). (D) Serial sections were stained with antibodies against PNAd and MIG. The number of PNAd+ and MIG+ vessels was then counted across 10 lymph nodes. Data is presented as the percentage of total HEVs that were MIG+. Lymph nodes that were not inflamed (white bar) had <2% MIG+ HEVs. Upon inflammation (black bar), the number of MIG+ HEVs increased to ∼12%. This experiment was repeated twice. (E) Immunohistochemistry was performed as shown in C. All PNAd+ (total HEVs) and MIG+ vessels were counted and scored for whether a monocyte was bound. Data is presented as percentage of HEV+ for at least one bound monocyte. Upon inflammation (black bars) this increased from ∼6% in lymph nodes that were not inflamed (white bar) to ∼20%. The percentage of the subpopulation of total inflamed HEVs that were MIG+ and had a bound monocyte was >60%; whereas, the percentage of the MIG− HEV subpopulation that had a bound monocyte was only 13%. This experiment was repeated twice.
Mentions: Serial sections of inflamed lymph nodes were subjected to double-indirect immunohistochemistry (Fig. 3 A). On all sections, an antibody against the B cell marker B220 was used to illuminate the B cell follicles for orientation. Serial sections were stained with antibodies against PNAd to identify HEVs, IP10, MCP-1 or an isotype control. While both MCP-1 and IP10 were highly expressed in macrophage-rich areas, to our surprise neither was displayed on HEVs (Fig. 3 A).

Bottom Line: Quantitative PCR analyses revealed the upregulation of many chemokines in the inflamed lymph node, including MCP-1 and MIG.HEVs did not express detectable levels of MCP-1; however, a subset of HEVs in inflamed lymph nodes in wild-type (but not tumor necrosis factor [TNF] mice) expressed MIG and this subset of HEVs preferentially supported monocyte binding.Together, these results suggest that the lymph node microenvironment can dictate the nature of molecules expressed on HEV subsets in a TNF-dependent fashion and that inflammation-induced MIG expression by HEVs can mediate monocyte recruitment.

View Article: PubMed Central - PubMed

Affiliation: DNAX Research Institute, Inc., Palo Alto, CA 94304, USA.

ABSTRACT
Monocytes recruited from the blood are key contributors to the nature of an immune response. While monocyte recruitment in a subset of immunopathologies has been well studied and largely attributed to the chemokine monocyte chemoattractant protein (MCP)-1, mechanisms mediating such recruitment to other sites of inflammation remain elusive. Here, we showed that localized inflammation resulted in an increased binding of monocytes to perifollicular high endothelial venules (HEVs) of lymph nodes draining a local inflammatory site. Quantitative PCR analyses revealed the upregulation of many chemokines in the inflamed lymph node, including MCP-1 and MIG. HEVs did not express detectable levels of MCP-1; however, a subset of HEVs in inflamed lymph nodes in wild-type (but not tumor necrosis factor [TNF] mice) expressed MIG and this subset of HEVs preferentially supported monocyte binding. Expression of CXCR3, the receptor for MIG, was detected on a small subset of peripheral blood monocytes and on a significant percentage of recruited monocytes. Most importantly, in both ex vivo and in vivo assays, neutralizing anti-MIG antibodies blocked monocyte binding to inflamed lymph node HEVs. Together, these results suggest that the lymph node microenvironment can dictate the nature of molecules expressed on HEV subsets in a TNF-dependent fashion and that inflammation-induced MIG expression by HEVs can mediate monocyte recruitment.

Show MeSH
Related in: MedlinePlus