“Proinflammatory T-cell responses to gut microbiota promote experimental autoimmune encephalomyelitis”
Lee YK, Menezes JS, Umesaki Y, & Mazmanian SK
PNAS. 2011 Mar; 108 (1): 4615-4622.

• Autoimmune disease
• T-cells enter CNS, attack myelin sheath
• Genetic precursors
• High rates of discordance in MZ twins (≥ 70%) http://www.ncbi.nlm.nih.gov/pubmedhealth/
PMH0001747/

• 100 trillion cells
• Primarily in gastrointestinal (GI) tract
• Confirmed role in GI immune system development and modulation

• MS associated with microbial contact
• More interaction with commensals
• Noninfectious symbionts modulate CD4 + T-cell responses in the intestine
– Reduced intestinal Th17 cells in GF mice
– Microbiota directs Th17 differentiation
– Th1 and Th17 response to infectious agents

The commensal gut microbiota modulates extraintestinal immune function in a model of multiple sclerosis.

Experimental Design
Results
Interpretation(s)

Experimental autoimmune encephalomyelitis (EAE)
• Mouse model of MS
• Induced by immunization with CNS antigens
– Myelin oligodendrocyte glycoprotein (MOG)
– Adjuvants: Freund’s adjuvant, pertussis toxin
• Immune cells enter CNS and destroy myelin sheath
• Th1 and Th17 cells highly associated with EAE development

Is EAE development affected in germ-free mice?
• Germ-free (GF) mice vs. specific pathogen-free (SPF) mice
• Induce EAE via inoculation with MOG/CFA
• Score EAE development

Why do GF mice display reduced EAE incidence and symptoms?

H&E stain
Myelin basic protein

• Reduced EAE in GF mice is due to lack of inflammation in the CNS
• Microbiota plays a role in the induction of EAE

• GF mice have developmental deficits for some inflammatory T-cell subsets
• Harvested CD4+ cells 8-10 days after immunization
• Reinoculated with MOG peptide
• Transferred to Rag1-/- SPF mice

T-cells from GF mice can be activated to induce EAE

• CD4+ T-cells from GF mice are not inherently unresponsive
• Microbes actively control the inflammatory response of T-cells in the CNS

Does the gut microbiota affect the proinflammatory profile of T-cells?
• Drained lymph nodes to harvest Th1 and Th17 cells 8d post-immunization
• Stained cells for IL-17A and IFNγ
– Cell-sorting
• Harvested cytokines
– ELISA

• Th1 and Th17 proinflammatory responses are reduced in the absence of the microbiota

Do certain microbes regulate extraintestinal immune response?
• Segmented filamentous bacteria (SFBs)
– “Uniquely able to induce Th17 cell differentiation in the small intestine” (p.4618)
• Inoculated GF mice with SFBs

Intermediate EAE development in
GF-SFB mice

• The microbiota influences the extraintestinal development of EAE, a mouse model of MS, through regulation of proinflammatory responses of Th1 and Th17
• SFBs in particular regulate EAE development

• The microbiota influences the extraintestinal development of EAE, a mouse model of MS, through regulation of proinflammatory responses of Th1 and Th17
• SFBs in particular regulate EAE development
• Reasonable, considering other autoimmune diseases in GF mice
• Relatively novel paradigm

• Investigate differences in T-cell activation after transfer to Rag1-/- mice, inoculation with SFB
– Role of microbiota in early immune system development
– Effect of additional microbial species
• Reduce EAE development in SPF mice via regulation of SFBs