FACS Quantitation of Leucine Aminopeptidase and Acid Phosphatase on Tumor-Associated Macrophages From Metastatic and Nonmetastatic Mouse Mammary Tumors

Abstract

Macrophages were isolated by adherence from tumors produced by a number of murine mammary carcinoma lines and were examined by fluorescence-activated cell sorting for quantitation of leucine aminopeptidase and acid phosphatase. The tumors included three lines, 66, 67, and 168, which were originally derived from a single, spontaneously arising tumor in a BALB/cfC₃H mouse and two other lines, D2A1 and D2F2, which were derived from a single tumor arising from the transplantable hyperplastic alveolar nodule line, D2. These five lines differ from one another in a number of characteristics, including the ability to metastasize spontaneously to the lung from subcutaneous implants and to form experimental metastases in lungs following intravenous injection. Line 67 is nonmetastatic under both circumstances, whereas lines 66, D2A1, and D2F2 are metastatic under the same conditions. Intermediate to these is line 168, which is nonmetastatic from the subcutaneous site but capable of colonizing the lung with an efficiency similar to 66 when injected IV. Tumor-associated macrophages (TAM) from lines 66, D2A1, and D2F2 contained the greatest amounts of leucine aminopeptidase (LAP) and those from line 67 the least, with TAM from 168 being intermediate. Conversely, the TAM from line 67 had the greatest amounts of acid phosphatase (APTase) and those from line 168 the least. In addition to differences among tumors in enzyme levels of the adherent TAM, the precentages of TAM that were adherent were also different among the tumors. Only 12% of TAM from line 67 were recovered in the adherent fraction as opposed to 35–38% of TAM from lines 66 and 168.

These results confirm and extend our previous findings that TAM from metastatic tumors have increased levels of LAP compared to TAM from nonmetastatic tumors. They also demonstrate noncoordinate expression of LAP and APTase in TAM, and illustrate how a population of TAM can be homogeneous for one enzyme and heterogeneous for another. Furthermore, the difference in the percentage of macrophages that are adherent between metastatic and nonmetastatic tumors is another indication both of the heterogeneous nature of TAM and of the role of a tumor in determining the type of host infiltrate with which it is associated.

tumor-associated macrophage, heterogeneity, activation