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Updated: Aug 17, 2026
Establishment of Epstein-Barr Virus Growth-transformed Lymphoblastoid Cell Lines
Published on: November 8, 2011
Maintenance of the CD40-related immunodeficient response in hyper-IgM B cells immortalized with a LMP1-regulated
Kristina T Lu1, Rebecca L Dryer, Charles Song
1Nelson Biological Laboratories, Rutgers, The State University of New Jersey, 604 Allison Road, Piscataway, NJ 08854, USA.
Insights
This study investigates a CD40 signaling defect in a patient with hyper-immunoglobulin M syndrome using Epstein-Barr virus lymphoblastoid cell lines. Results show B cells retain the defect, offering a model to study CD40 and LMP1 signaling independently.
Area of Science:
- Immunology
- Cell Biology
Background:
- Patient pt1 with hyper-immunoglobulin M syndrome exhibits a CD40-mediated B cell activation defect.
- This defect leads to low CD23 expression, absent germ-line transcription, and impaired class-switch recombination.
- In vitro studies showed these deficiencies could be corrected by sustained CD40 signaling.
Purpose of the Study:
- To further analyze the CD40 signaling defect in pt1 B cells.
- To compare downstream functions in response to constitutive versus regulated CD40 and latent membrane protein-1 (LMP1) signals.
- To establish a model for studying the independent effects of LMP1 and CD40 signaling.
Main Methods:
- Generated two types of Epstein-Barr virus lymphoblastoid cell lines (LCLs) from pt1: pt1-LCL (constitutive LMP1 expression) and pt1-LCL(tet) (tet-inducible LMP1 expression).
- Compared B cell phenotypes (CD23, CD38 expression) and responses to CD40 and LMP1 signals in both LCL types.
- Analyzed mitogenic activation in response to CD40 and LMP1 signals.
Main Results:
- Immortalized pt1-LCLs initially showed low CD23 and high CD38, reverting to high CD23/low CD38 with culture, suggesting CD40 defect reversal.
- pt1-LCL(tet) cells maintained the CD23(lo)/CD38(hi) phenotype and failed to up-regulate CD23 upon CD40 stimulation.
- Mitogenic activation in pt1-LCL(tet) cells was dependent on LMP1, not CD40, indicating distinct signaling pathway responses.
Conclusions:
- The pt1-LCL(tet) cell line maintains the CD40-related signaling defect.
- This model allows for the independent study of LMP1 and CD40 signaling pathways.
- The findings highlight differences in B cell responses to CD40 and LMP1 signals in the context of the pt1 defect.
Abstract:
Our previous investigation of a patient (pt1) with non-X-linked hyper-immunoglobulin M syndrome revealed a CD40-mediated defect in B cell activation that resulted in low CD23 expression and absence of germ-line transcription and class-switch recombination. These deficiencies were complemented in vitro by a high threshold of sustained signaling through CD40. To further analyze the signaling defect in pt1 B cells, two types of Epstein-Barr virus lymphoblastoid cell lines (LCLs) were generated that either constitutively expressed the viral transforming protein latent membrane protein-1 (LMP1; pt1-LCL) or expressed it under the control of a tet-inducible promoter (pt1-LCL(tet)). Because LMP1 signals through the CD40 pathway, the pt1-LCL and pt1-LCL(tet) lines allow comparison of downstream functions in response to either constitutive LMP1 signals or regulated LMP1 and CD40 signals. Immortalized pt1-LCLs were initially CD23(lo)/CD38(hi) and reverted to a CD23(hi)/CD38(lo) phenotype upon extended growth in culture, suggesting that the CD40 defect was reversed by selection and/or constitutive expression of LMP1. In contrast, pt1-LCL(tet) cells retained the CD23(lo)/CD38(hi) phenotype after extended periods of culture and failed to up-regulate CD23 in response to CD40 signals. Analysis of pt1-LCL(tet) cells in response to the CD40 signals in the presence or absence of LMP1 revealed that mitogenic activation resulted only from LMP1 and not CD40, indicating a difference in the response of pt1 B cells to these two distinct signals. Together, these data demonstrate that the pt1-LCL(tet) cells maintain the CD40-related defect and provide a unique approach to study the independent effects of LMP1- and CD40-directed signals.
