DTH Model Introduction

Delayed-type hypersensitivity (DTH) is an inflammatory response characterized by mononuclear cell infiltration and tissue cell damage after the action of effector T cells with corresponding antigens. It usually occurs during bacterial and viral infections, oncological diseases, transplant rejection, and contact dermatitis.

Mouse contact hypersensitivity response is a widely used model to study T cell-mediated immune responses to contact allergens. Oxazolone (OXA) is a hapten, which can combine with skin proteins to form complete antigens when applied to the skin, stimulating the proliferation of T lymphocytes into sensitized lymphocytes. When applied again with OXA after 4-7 days, it causes a delayed allergic reaction locally.

Keyhole limpet hemocyanin (KLH) is a highly immunogenic protein macromolecule that enables dendritic cells to recognize Th1 cells. It is an ideal antigen for inducing late DTH. KLH is introduced systemically in mice. And then locally reintroduced several weeks later, DTH response was assessed by relative swelling at the injection site.

OXA and KLH-induced DTH mouse models were established by Biocytogen, which can be used for immunosuppressant efficacy evaluation.

Results

OXA induced DTH Mouse Model based on BALB/c mice

Establishment of DTH Mouse Model- OXA-induced DTH mouse model

Efficacy Evaluation of Dexamethasone in OXA-induced DTH Model of BALB/c Mice

Efficacy evaluation of Dexamethasone in DTH model of BALB/c mice. A. Ear thickness (mm). B. Change of ear thickness (mm). (A. Two-way ANOVA; B. One-way ANOVA; ** p< 0.01 , **** p< 0.0001).

Histologic Assessment of DTH in BALB/c Mice. Ear-swelling responses induced by 1% OXA were assessed by measurement of ear thickness (H&E staining). And assessment the efficacy of anti-TNFα Ab or Dexamethasone (Dex) in DTH model.

KLH Induced DTH Model

Establishment of DTH Mouse Model

• Experimental Animals: C57BL/6, BALB/c, 7-8 weeks old, female
• Modeling reagent: Keyhole Limpet Hemocyanin (KLH)
• Modeling sites: subcutaneous injection (s.c.); intradermal injection (i.d.)
• Modeling method: KLH injection on neck or ear, Sensitization: day 0, s.c. neck; Challenge: day 7, i.d. ear

Readouts
Included tests	Clinical scores	Body weight
		Ear thickness
	Ear histopathology (H&E)	Epidermal thickness
Optional tests	Molecular levels	Protein level (Elisa or Luminex)

Establishment of DTH Mouse Model based on C57BL/6 Mice

Detection indicators
Clinical symptoms	Body weight
	Ear thickness

Establishment of DTH Mouse Model based on C57BL/6 Mice
On Day 0, mice are immunized by an intraperitoneal injection with a KLH/CFA/IFA emulsion. On Day 7 , following baseline ear thickness measurements, mice are challenged with KLH by intradermal injecting (i.d.) into the ear. Additionally, dexamethasone 3 mg/kg in 10ml/kg) were administered subcutaneously for 4 consecutive days，and ear swelling is then measured on Day 8, Day 9 and Day 10.

Efficacy Evaluation of Dexamethasone on KLH-induced DTH model

Dexamethasone alleviates KLH-induced an DTH phenotype in C57BL/6 mice. Body weight (A), (B) Ear Thickness. Values are expressed as mean ± SEM.

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Detection indicators
Included detection	Clinical symptoms	Body weight
		Ear thickness
Optional detection	Histopathological scores	Immune infiltration
	Flow cytometry	Lymph nodes or spleen
	ELISA	Cytokine/chemokine analysis