Functional Cellular Responses and Cytokine Profiles

The objective of this section is to provide a description of several immune parameters and methodologies that are used in the preclinical setting to characterize immune hazards for human risk assessments. The utility of a standalone functional evaluation, specifically the rat T cell-dependent antibody response (TDAR) model developed for regulated immunotoxicity evaluations (Gore et al, 2004), is mainly used for hazard identification, i.e., unintended immunosuppression. This assay is based on an end point parameter that is the 

Immunotoxicology Strategies for Pharmaceutical Safety Assessment, edited by Danuta X Herzyk 

Additional parameters that are readily incorporated into a stand-alone immune function test such as the KLH-TDAR model include ex vivo lymphocyte proliferation, cytokine protein expression, and immunophenotype analysis any or all of which can enhance hazard identification and characterization of a potential immunotoxicant. While the KLH-TDAR is an example of a combined immune function screen and mechanistic study, the ex vivo methodologies described herein are generally applicable to toxicology studies that do not include an immunization protocol. Moreover, the methodologies are not species-specific however, responsiveness to various stimulants to induce ex vivo lymphocyte proliferation and cytokine production may differ across species and strain, requiring procedural optimization for a given species and ex vivo test. 

While ex vivo proliferation may or may not afford the level of sensitivity as the more predictive functional tests (i.e., TDAR and NK cytotoxicity) (Luster et al., 1992 Dean et al., 1998 The ICICIS Investigators Group, 1998), concomitant alterations in complementary parameters, albeit not over the entire in vivo dose range, would provide a potential mechanism of the manifestation of an immune perturbation. This notion would support a single dose level (e.g., maximum exposure) to test ex vivo parameters, while the in vivo functional parameter with higher sensitivity would be performed over the entire dose range. 

Characteristic of immune function evaluations, there is considerable diversity in the approaches to proliferation assays, from the experimental design to the analytical method. Experimental procedures necessitate sterile technique and cell culture expertise to ensure accurate assessment of the cellular response to a particular stimulant. Culture conditions vary depending on the cell source and type of stimulant, but generally are conducted at 5% CO2,37 °C for 48 to 96 hours. The source of lymphocytes may be from peripheral blood, spleen. 

---

Chapter 4.1 Functional Cellular Responses and Cytokine Profiles... 

---