Mice deficient in MIM expression are predisposed to lymphomagenesis

Abstract

Missing in metastasis (MIM) is a member of newly emerged inverse Bin-Amphiphysin-Rvs (BAR) domain protein family and a putative metastasis suppressor. Although reduced MIM expression has been associated with bladder, breast and gastric cancers, evidence for the role of MIM in tumor progression remains scarce and controversial. Herein we characterized a MIM knockout mouse strain and observed that MIM-deficient mice often developed enlarged spleens. Autopsy and histological analysis revealed that nearly 78% of MIM(-/-) mice developed tumors with features similar to diffuse large B lymphoma during a period from 1 to 2 years. MIM(-/-) mice also exhibited abnormal distribution of B cells in lymphoid organs with decrease in the spleen but increase in the bone marrow and the peripheral blood. Furthermore, the bone marrow of MIM(-/-) mice contained a higher percentage of pre-B2 cells but fewer immature B-cells than wild-type mice. In response to CXCL13, a B-cell chemokine released from splenic stromal cells, MIM-deficient B-cells did not undergo chemotaxis or morphological changes in response to the chemokine and also did not internalize CXCR5, the receptor of CXCL13. Microarray analyses demonstrated that MIM is the only member of the I-BAR domain family that was highly expressed in human B cells. However, low or absent MIM expression was common in either primary B-cell malignancies or established B-cell acute lymphocytic leukemia or lymphomas. Thus, our data demonstrate for the first time an important role for MIM in B-cell development and suggest that predisposition of MIM-null mice to lymphomagenesis may involve aberrant interactions between B lineage cells and the lymphoid microenvironment.

Fig. 1. Phenotypes of MIM KO mice

(A) MIM(−/−) mice have a shorter life span as compared to MIM(+/−) and MIM(+/+) mice as observed within 24 months. The p value (t test) refers to the difference between MIM(−/−) and MIM(+/+) mice. (B) Comparison of the spleens from a two-month old MIM(−/−) mouse and a MIM(+/+) mouse at the same age based on the weight (right, n=12) and the size (right). (C) Comparison of the whole body weight of MIM(−/−) and MIM(+/+) mice at 1 and 2 month old (n=6), respectively.

Fig. 2. Histological analysis of lymphoma developed in MIM KO mice

(A) HE staining of a lymphoma section. Several mitotic cells were indicated by arrows. (B) A lymphoma section was stained with CD20 antibody. (C) HE staining of a normal lymph node. (D) HE staining of a normal spleen. Original magnifications: 600 x.

Fig. 3. Aberrant distribution of MIM KO B lineage cells in the bone marrow and the spleen

(A) MIM is abundantly expressed in splenic B cells but weakly in splenic T cells. (B) Flow cytometry analysis of CD19⁺ B cells isolated from the spleen (SP), the bone marrow (BM) and the peripheral blood (PB). (C) Quantification of CD19⁺ B cells in lymphoid organs (n=3). P values (t test) refer to the statistical difference between WT and KO samples.

Fig. 4. MIM KO mice were partially impaired in B-cell development and the motility response to CXCL13

(A) The bone marrow of MIM KO mice contained an increased population of pre-B2 cells but a decreased population of immature B cells. (B) MIM KO mice developed significantly less sIgM+ B cells in both spleen and bone marrow. (C) MIM KO mice developed less sIgD+ B-cells in the spleen. (D) Splenic MIM KO B cells were unable to migrate efficiently into Transwell chambers containing 1 μg/mL CXCL13. (E) Motility of B cells was measured at difference doses of CXCL13. (F) MIM heterozygous B cells displayed a normal response to CXCL13. All the data represent three independent experiments (mean ± SEM).

Fig. 5. MIM is abundantly expressed in normal human B cells but poorly expressed in neoplastic B cells

(A) Microarray analysis of the transcripts of the I-BAR domain family members in human primary B cells (n=4), T-cells (n=8), granulocytes (n=4) and monocytes (n=2). (B) Differential expression of MIM in B acute lymphocytic leukemia (BALL) and normal primary peripheral blood B cells. MIM expression was measured by microarray in acute BALL cell lines (n = 9 cell lines, in triplicates, total 27 arrays), precursor B-ALL (P-BALL) patient samples (n=11) and normal peripheral blood CD19+ B cells (PRB) (n=4). (C) MIM is poorly expressed in lymphoma cell lines as measured by Western blot. a, CMK (a MIM expressing human megakaryoblastic cell line); b, MCF-7 breast cancer cells; c, WEHI231 (murine); d, SU-DHL-6; e, SU-DHL-4; f, Ramos; g, RAJI; h, Daudi; i, Karpas-k422; and j, Farage. *, indicating a weak MIM expression.