Patricia A. John

Preferential homing of tumor-infiltrating lymphocytes in tumor-bearing mice

SummaryIn view of the current interest in the use of lymphoid cells in adoptive immunotherapy of patients with advanced cancer, we have studied the homing patterns of various lymphoid effector cells in mammary-tumor-bearing mice. Single-cell suspensions of total splenocytes, natural killer (NK) cells, and lymphokine-activated killer (LAK) cells were prepared from the spleens of C3H/OuJ mice. Tumor-infiltrating lymphocytes (TIL) were isolated from mammary adenocarcinomas excised from retired breeder females of the same substrain. Effector cells were labeled with indium-111 and injected via a tail vein into female C3H/OuJ mice bearing one or more mammary tumors. Twenty-four hours after administration, total splenocytes, NK cells, and LAK cells distributed themselves evenly between normal mammary tissue and mammary adenocarcinomas. Only TIL had a higher concentration in tumors than in corresponding normal mammary tissue. The ability of the different lymphocyte preparations to lyse YAC-1 cells was determined by means of a 4-h 51Cr-release cytotoxicity assay. Cells harvested from LAK cell cultures and further enriched by centrifugation through a discontinuous Percoll gradient and interleukin-2 (IL-2)-stimulated TIL demonstrated the highest levels of cytotoxicity, while total splenocytes and fresh TIL were characterized by the lowest levels. Since IL-2-stimulated TIL were highly cytotoxic and exhibited better tumor localization than both NK cells and LAK cells in this system, they may be the lymphoid effectors of choice for adoptive immunotherapy of advanced cancer.

Response to and production of prostaglandin by murine thymus, spleen, bone marrow, and lymph node cells

Abstract Prostaglandins influence cellular immune responses of mice and men. We demonstrated that the proliferation of cells from the spleen, lymph node, bone marrow, and thymus of young adult C57BL/6 mice may be inhibited by exogenous and endogenous prostaglandin E 2 . Thymocytes produced very little prostaglandin in the presence or absence of mitogen; lymph node cells showed no increase in prostaglandin (PG) production with mitogen; bone marrow cells and splenocytes demonstrated fourfold and tenfold increases in production of prostaglandin in the presence of concanavalin A. Bone marrow and thymus cells were more sensitive to PG suppression. The sensitivity of the various cell populations, the quantities of supernate IPG, and augmentation by indomethacin suggest that PG may modulate cell function in the bone marrow, thymus, and spleen, but not in unchallenged lymph node.

Decreased natural cytotoxicity in mice with high incidence of mammary adenocarcinoma

In an attempt to gather evidence relevant to the question of whether natural killer (NK) cells play a role in resisting the development of primary tumors, we compared natural cell-mediated cytotoxicity in two substrains of C3H mice. Animals of the C3H/OuJ substrain are at high risk for the formation of mammary adenocarcinomas, while C3Heb/FeJ mice have a low incidence of such tumors. Natural cytotoxicity of splenic mononuclear cells was lower in the high-risk substrain, suggesting that a lesion in NK cell activity may be involved in murine mammary tumorigenesis. This difference was observed in animals between 5 and 37 weeks of age. There was no significant difference in the number of splenic large granular lymphocytes between the substrains. A significant difference in the ability of splenic lymphocytes from the two substrains to bind to the target cells was noted. Since the binding capacity of lymphocytes was greater in mice with reduced NK cell activity, the lesion in cytotoxicity may exist at a postbinding step in the lytic sequence. It is felt that the C3H mouse may provide a useful model for studying the role of NK cells in controlling primary tumors.

Lysis of fresh murine mammary tumor cells by syngeneic natural killer cells and lymphokine-activated killer cells

SummaryWe have compared the ability of natural killer (NK) cells from two substrains of C3H mice that differ with respect to their susceptibility to the development of mammary adenocarcinomas to lyse fresh syngeneic mammary tumor cells. Single cell suspensions of mammary tumors from retired breeder females were used as targets in 22-h 51Cr-release cytotoxicity assays with syngeneic NK cells. Tumor cell suspensions were prepared by enzymatic digestion of finely minced tissue followed by centrifugation through a discontinuous Percoll gradient. Effector cells were prepared by passing spleen cells over nylon wool followed by centrifugation through Percoll fraction 7. Syngeneic NK cells had significant levels of lysis against 5/8 tumors studied. NK cells from low risk animals (C3Heb/FeJ) consistently demonstrated greater cytotoxicity against tumor cell preparations than did effectors from the high tumor substrain (C3H/OuJ). Study of cytocentrifuge preparations stained with Wright-Giemsa revealed that the two substrains were identical with respect to the number of azurophilic granules present in the cytoplasm of their NK cells. We have also shown that lymphokine-activated killer (LAK) cells can be generated from splenocytes in C3H mice. While LAK cells from both substrains were capable of lysing fresh syngeneic mammary tumor cells in vitro, LAK cells from the animals at high risk for the formation of mammary adenocarcinomas had greater cytotoxicity against tumor cell suspensions than LAK cells from the low tumor substrain.

Shorter turn-around-time and improved patient care in peripheral blood progenitor cell collection procedures.

The collection of peripheral blood progenitor cells (PBPC) requires the combined efforts of the Transfusion Medicine/Hemapheresis and Hematology/Oncology services and HLA/Progenitor Cell and Immunology laboratories. Coordination and communication among these different services and laboratories are key to attaining an optimal collection in a timely manner for the patient undergoing PBPC collection. In an effort to improve patient care by same‐day decision to cease or continue collections avoiding unnecessary collections, needless patient trips to the hospital and ultimately increasing patient satisfaction, a flow chart was used to capture the sequence of events. The flow chart served as a powerful tracking tool that defined system process and steps to attain enumeration of CD34+ cells the same day of collection. It provided documentation of work flow from each of the independent operations involved in progenitor cell collection and enumeration turn‐around‐time including attending and staff time involvement. By using the flow chart, potential and actual problem areas were demonstrated and this allowed for creative thinking and problem solving by individual sections rather than recriminations. Finally, it focused all the staff involved in the common goal of a shorter turn‐around‐time for CD34+ cell enumeration the same day of collection. This allowed a prompt decision for subsequent leukapheresis as improved service to oncology patients and their physicians. J. Clin. Apheresis 14:35–41, 1999.