Omentum and milky spots

• the omentum is formed by a double layer of

mesothelial cells

• connect the stomach, pancreas, spleen and

colon

• has immunological and wound-healing properties

• embedded within the omentum are structures

which are clusters of leukocytes, called milky

spots

Omentum and milky spots• milky spots are mainly composed of macrophages and

B1 cells

• B1cells forma unique subset of B cells

• can be distinguished from conventional B (B2) cells by

expression of distinct cell-surface markers and antigen

receptors that can bind common bacterial epitopes

• have a recognized potential to produce natural

antibodies that provide a first protection to bacterial

infections

• B1 cells are localized in distinct locations, such as the

peritoneal cavitiy and the spleen

• MS described to lack dendritic cells as well as follicular

dendritic cells

Are milky spots secondary lymphoid organs?

Mouse system

• want to address the immunological potential of milky spots in the

absence of lymph nodes, spleen, and Peyer’s patches

• used splenectomized lymphotoxin-alpha (LTa)-deficient mice (Lta-/-),

which as a result of their deficiency already lacked lymph nodes

and Peyer’s patches

• animals, devoid of secondary lymphoid organs, were reconstituted

with wild-type bone marrow (SLP mice)

• compared to irradiated C57BL/6 mice that were similarly

reconstituted with wild-type bone marrow

Antibody responses to peritoneal antigens in the absence of conventional lymphoid organs

• WT and SLP mice produce similar titers of NP-specific IgM, IgG1, IgG2a+b and IgG3• after immunization with TNP-KLH SLP mice showed slower generation of IgG titer• concentration to see how much secific IgG was produced

conventional lymphoid organs are not needed for B cell response

NP-OVA i.p. TNP-KLH i.p. NP-OVA i.p.

The milky spots of the omentum collect peritoneal cells and antigens

• activation of peritoneal cells promotes their migration to the omentum • at least some particulates can be passively collected by the omentum in addition to being captured by phagocytic cells

HEV + B cell

The milky spots of the omentum collect peritoneal cells and antigens

• peritoneal cells use PTX-sensitive mechanisms to actively migrate to the omentum • can also accumulate in the omentum by other mechanisms • the segregation of B and T cells and the formation of follicular structures in the MSs are controlled by PTX-sensitive mechanisms

Antigen-specific B cell responses occur in the omentum

GC B cells

specific non-specific

NP-OVA i.p. SRBC i.p boost d14

d8 d5

GC B cells Isotype-switchedplasmacells

SRBC specific IgG secreting cells

Antigen-specific B cell responses occur in the omentum

• MSs support local germinal center B cell responses to peritoneal antigens

GC B cellsSRBC i.p d14 NP-OVA i.p d14

Antigen-specific T cell responses in the omentum

• T cell responses can also occur

in the MSs of the omentum,

even in mice that lack spleen,

LNs, and Peyer’s patches

T cell activation homing receptor OTII CD4 i.p.4 hr later immunized

Antigen-specific T cell responses in the omentum

• antigen-experienced CD4+ and CD8+ T cells recirculate through the peritoneal cavity and omentum• even when these cells were primed outside of the peritoneal cavity

influenza i.n.21d p.i.

4-get mice helminth larvae oral28 d p.i.

Milky spot development requires LTα and CXCL13, but not LTi cells inactive genes

for CCL19, CCL21

• MSs were much smaller or even absent in Lta-/- and Cxcl13-/- mice• MSs were much smaller or even absent in Lta-/- and Cxcl13-/- mice• defects in theMSs of Lta-/- mice seem not be due to an absolute blockade in development

Milky spot development requires LTα and CXCL13, but not LTi cells

• even though CXCL13 is very important for the development of the MSs, its expression is not controlled by Lta• Lti cells are not needed

lymphoid tissue inducer cells

CXCL13 is expressed surrounding the B cell areas

• CXCL13 is expressed around the B cell area• absence of conventional FDC networks

spleen

omentum

CXCL13 is expressed surrounding the B cell areas

• network of ERTR7+ stromal cells throughout the MSs• the cellular architecture and pattern of chemokine expression in the MSs are different than in conventional lymphoid organs

Somatic hypermutation and affinity selection occurs in the absence of conventional secondary

lymphoid organs

• somatic hypermutation is still evident in SLP mice• but the process of clonal selection is unusual• the MSs of the omentum support some aspects of T cell dependent B cell responses

NP-OVA i.p.boost d14

W to L position 33 higher affinity

YYYG motif at V-Djunction NP-binding sequence

Summary

• the omentum functions much more broadly as a secondary

lymphoid organ

• it is structurally, developmentally, and functionally unique

• intersection of recirculating lymphocytes and peritoneal drainage makes the

MSs ideal sites for the initiation of local immune responses

• the structure of the MSs seems inside-out relative to that of conventional

lymphoid organs or even ectopic lymphoid follicles

the MSs of the omentum are unique secondary lymphoid organs that

sample antigens from the peritoneal cavity and promote local, albeit

unusual, immune responses