Detection of the bcl-2 t(14;18) translocation and proto-oncogene expression in primary intraocular lymphoma

Abstract

Purpose: Primary intraocular lymphoma (PIOL) is a diffuse large B cell lymphoma that initially infiltrates the retina, vitreous, or optic nerve head, with or without central nervous system involvement. This study examined the expression of the bcl-2 t(14;18) translocation, the bcl-10 gene, and high expression of bcl-6 mRNA in PIOL cells.

Methods: Microdissection and PCR analysis were used to examine vitreous specimens in patients with PIOL for the presence of bcl-2 t(14;18) translocations, the bcl-10 gene, and expression of bcl-6 mRNA. A medical record review was also conducted to determine whether the bcl-2 t(14;18) translocation correlated with prognosis.

Results: Forty of 72 (55%) PIOL patients expressed the bcl-2 t(14;18) translocation at the major breakpoint region. Fifteen of 68 (22%) patients expressed the translocation at the minor cluster region. The bcl-10 gene was detected in 6 of 26 (23%) patients, whereas 4 of 4 (100%) PIOL patients expressed higher levels of bcl-6 mRNA compared with inflammatory lymphocytes. An analysis of clinical outcome in 23 PIOL patients revealed no significant association between bcl-2 t(14;18) translocations and survival or relapse. However, patients with the translocation were significantly younger.

Conclusions: PIOL has unique molecular patterns of bcl-2, bcl-10, and bcl-6 when compared with other systemic lymphomas. This study lays the foundation for future studies aimed at exploring the genotypic classification of PIOL based on the quantitative molecular framework of gene expression profiling, with the goal of providing useful adjuncts to the pathologic diagnosis of this complex disease.

FIGURE 1. Typical cytology of a PIOL cell

A characteristic PIOL cell (arrow) from the vitreous with basophilic cytoplasm and large prominent irregular nuclei.

FIGURE 2

(A) Sample gel of PCR analysis for the bcl-2 t(14;18) translocation at the Mbr. PCR amplification of DNA from microdissected cells from vitreous specimens of PIOL patients using primers for the bcl-2 t(14;18) translocation at the Mbr. Lanes 1 to 5, PIOL samples; +, positive control; −, negative control. Seventy-two vitreous specimens were examined; 40 (55%) were positive. (B) Sample gel of PCR analysis for the bcl-2 t(14;18) translocation at the mcr. PCR amplification of DNA from microdissected cells from vitreous specimens of PIOL patients using primers for the bcl-2 t(14;18) translocation at the mcr. Lanes 1 to 3, PIOL samples; +, positive control; −, negative control. Sixty-eight vitreous specimens were examined; 15 (22%) were positive. In both (A) and (B), differences in migration through the gel were caused by variability where the translocation could occur in the J_H region of the IgH gene, resulting in different sizes of amplified product.

FIGURE 3. Sample gel of PCR analysis for the bcl-10 gene

PCR amplification of DNA from microdissected cells from vitreous specimens of PIOL patients using primers for the bcl-10 gene. Lanes 1 to 5, PIOL samples; −, negative control. Twenty-six vitreous specimens were examined; 6 (23%) were positive.

FIGURE 4

Analysis of expression of bcl-6 mRNA in PIOL cells compared with non-PIOL cells. Expression of bcl-6 was examined by PCR amplification of the cDNA complement of mRNA from four PIOL samples, four uveitis samples, and one healthy control sample. Autoradiography was used to compare relative amounts of bcl-6 and 18S mRNA. All four PIOL patients had higher expression of bcl-6, but none of the non-PIOL patients had high bcl-6 expression.