Weekly Injections

Figure 5.14. Loss of splenic B cells following repetitive injection of anti-CD137. BALB/c mice were injected weekly with 200 |g I.P. the indicated number of times. One week after the final injection the mice were euthanized and the frequency and absolute numbers (data not shown) of CD19+ B cells determined by flow cytometry. Control mice were injected with a non-crossreactive rat anti-human CD137 mAb.

analysis revealed other anomalies or pathologies in the treated animals. Included among these were splenomegaly, lymphadenopathy, hepatomegaly, anemia, extramedullar hematopoiesis, abnormal white blood cell counts, loss of CD19+ cells in the spleen and bone marrow, a marked increase in B cells in the lymph nodes, and marked elevation of Lin- Sca-1 + c-kit+ bone marrow cells (Niu et al., manuscript submitted). We further noted marked accumulation of CD8+ T cells in the lungs, liver, and bone marrow. Figure 5.14 illustrates the reduced frequency of CD19+B cells in the spleens of naive C57BL/6 given two or five weekly intraperitoneal injections of anti-CD137. The reduced frequency of splenic B cells was mirrored by a loss in absolute numbers of B cells and not due to an increase in other cell elements.

Accompanying the loss of CD19+ B cells from the spleen, anti-CD137 injected mice lost almost all antibody secreting cell function following five weekly injections as indicated by a precipitous drop in serum IgG levels (Figure 5.15).

This was particularly evident in the bone marrow where greater than 98% of antibody secreting plasma cells were lost (Figure 5.16). The reason for this loss is not clear, but it is important to note that we observed a tenfold increase in the number of CD8 T cells in the bone marrow of these mice, and this suggests to us that their presence may be a contributing factor.

The disruption of normal homeostasis of the hematopoietic system together with hepatomegaly and multi-focal hepatitis in anti-CD137 treated mice is T cell dependent. Repeated injections of anti-CD137 mAbs in nude or Rag-deficient mice have no observable ill effect. However, following T and B cell reconstitution, i

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Figure 5.15. Loss of serum IgG following injection of anti-CD137. BALB/c mice were injected I.P. with 200 |g of anti-CD137 or rat IgG weekly for 5 weeks. At week 6 the mice were bled and serum IgG levels were determined by ELISA.

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