EFFECTS OF HUSBANDRY AND MOUSE STRAINS ON MONONUCLEAR PHAGOCYTES
Monte S. Meltzer
Mouse peritoneal macrophages are widely used as a standard for study of mononuclear phagocyte function. To be useful as a standard, however, morphologic, functional, and biochemical properties of these cells should be reproducible: reproducible from day to day within one's own laboratory and between dif- ferent laboratories. Two considerations that can profoundly affect reproducibility are the strains of mice selected for study and conditions under which the animals are bred and housed. Despite the importance of these factors, however, there have been very few systematic studies on macrophage func- tion and the general health of laboratory mice. For example, male mice from certain strains housed 5 - 8 cage establish a pecking order by tail and ear biting. Once this pecking order is established, tail biting ceases. However, transfer of a new male mouse into that cage restarts the entire process.
Macrophage function in these mice, animals with chronic super- ficial skin lesions, may be very different from "normal" mice.
For the most part, general information on the care and use of laboratory mice can be found in any of several standard text-
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books and manuals and will not be discussed here (1 - 4). Ani- mals with overt clinical signs of infection or abnormal develop- ment are usually recognized, and the causative event corrected or the animals excluded from study. More serious situations, however, arise from breakdown of proper laboratory animal man- agement (housing, sanitation, and husbandry) and inapparent or subclinical infections. More often than not, these two problems are coincident.
Difficulties described for quantitation of macrophage- induced tumor cytotoxicity illustrate the complexity of this problem. There is general agreement that with normal mice, resident peritoneal macrophages or inflammatory macrophages in- duced in the peritoneal cavity by injection of sterile irritants are not cytopathic to tumor cells in vitro. Peritoneal macro- phages, however, can develop tumoricidal activity after treat- ment in vivo or in vitro with any of a wide variety of activa- tion agents. There have been instances when macrophages from apparently normal mice exerted tumoricidial activity. Boyle and Omerod (5) showed that macrophages from healthy mice housed in a conventional but "dirty" animal room were cytopathic to tu- mor cells in vitro; cells from matched mice kept isolated in a relatively "clean" animal room had no effect on tumor cell growth. Similar observations were made with nude mice: macro- phages from nude mice housed in a clean conventional environ- ment were tumoricidal; cells from germ-free nude mice had no cytotoxic activity (6). Both reports illustrate situations in which specific-pathogen-free animals were affected by their en- vironment to develop alterations in macrophage function. In both cases, mice had no overt symptoms of infection.
It should be noted at this point that certain procedures intended to limit subclinical infections in an animal colony can also affect macrophage activity. The suppression of macro- phage function in mice that receive hyperchlorinated (25 ppm) drinking water is an example of such procedures (7). Moreover, cells from germ-free animals are not comparable to cells from conventionally housed animals and, in fact, have been shown to have several functional defects (8).
Subclinical viral infections are a major cause of abnormal immune responses in apparently healthy mice. There are many rodent viruses which can be carried in an animal colony without overt symptoms of i-lness or even gross pathologic changes.
Alterations in normal immune function have been reported for inapparent infections with murine leukemia virus (9), lympho- cytic choriomeningitis virus (10), murine cytomegalovirus (11), reovirus 3 (12), ectromelia virus (13), murine hepatitis virus
(14), minute virus of mice (15), lactic dehydrogenase-elevating virus (16), and Sendai virus (17). Many of these viruses are carried in transplantable tumors. One should, therefore, be especially vigilant in examination of macrophage function in
tumor-bearing animals. Sérodiagnostic services such as that offered by Microbiological Associates, Inc., Rockville, Mary- land should be used to monitor periodically animals for a wide spectrum of murine viruses.
Genetic differences in macrophage function between strains of mice must also be considered in the study of mononuclear phagocytes. In attempts to establish a new assay, it is, of course, most prudent to use the strain of mice for which the particular assay of macrophage function was initially defined.
This is not always possible. One should therefore be aware of possible genetic differences in macrophage response. Charac- terization of these genetic differences, however, is very in- complete. There are few systematic studies in the current literature which address this problem. One example of differ- ences in macrophage function between mouse strains can be shown for development of nonspecific tumoricidal activity following in vivo or in vitro treatments (18) (Table I). It is likely that similar observations can be made with other mononuclear phagocyte functions.
TABLE J. Genetic Variation in Development of Macrophage Tumoricidal Activity among Mouse Strains
Responsive C3H/HeN AKR/N CBA/CaHN CBA/N C57BL/6N C57BL/10J C57L/N DBA/UN DBA/2N NZW/N NZB/N
NIH Swiss (outbred) BlO.A/SgSnJ
Variable BALB/cAnN A/WySnJ RII/AnN
Nonresponsi v<
A/J A/HeJ A/HeN AL/N C3H/HeJ C57BL/10SCCR C57BL/10SCN P/J
P/JN
aPeritoneal exudate macrophages from mice inoculated intrâ- per itoneally 7 days previously with viable Mycobacterium bovis, strain BCG were adjusted to an equal macrophage concentration and cultured with prelabeled tumor target cells for 48 hr.
Cytotoxicity was estimated by measurement of radiolabel release and confirmed by observation of cultures under phage microscopy.
REFERENCES
1. "Biology of the Laboratory Mouse" (E. L. Green, e d . ) , Second edition. Dover Publications, Inc., New York, 1975.
2. "Guide for the Care and Use of Laboratory Animals." U.S.
Department of Health, Education and Welfare. Public Health Service, National Institutes of Health. DHEW Publication No. (NIH) 78-23, 1978.
3. "Long-Term Holding of Laboratory Rodents." ILAR News.
Volume XIX, Number 4, 2076. (Institute of Laboratory Ani- mal Resources, Assembly of Life Sciences, National Research Counci..)
4. Laboratory animal management: Rodents. ILAR News, Vol.
XX, Number 3, 1977.
5. M. D. P. Boyle and M. G. Ormerod. Destruction of alloge- neic tumor cells by peritoneal macrophages: Production of lytic effectors by immune mice. Transplantation 21: 242- 246, 1976.
6. M. S. Meltzer. Tumoricidal responses in vitro of peri- toneal macrophages from conventionally housed and germ-free nude mice. Cell. Immunol. 22: 176-181, 1976.
7. Fidler, I. J. Depression of macrophages in mice by drink- ing hyperchlorinated water. Nature 270: 735-736, 1977.
8. T. T. MacDonald and P. B. Carter. Requirement for a bac- terial flora before mice generate cells capable of mediat- ing the delayed hypersensitivity reaction to sheep red blood cells. J. Immunol. 122: 2624-2629, 1979.
9. W. S. Ceglowski and H. Friedman. Immunosuppression by leukemia viruses. III. Adoptive transfer of antibody- forming cells to Friend disease virus-infected mice.
J. Immunol. 103: 460-466, 1970.
10. C. A. Mims and S. Wainwright. The immunodepressive action of lymphocytic choriomeningitis virus in mice. J. Immunol.
101: 7171-7172, 1968.
11. J. E. Osborn and J. Medearis. Studies of relationship between mouse cytomegalovirus and interferon. Proc. Soc.
Exp. Biol. Med. 121: 819-824, 1966.
12. J. 0. Klein, G. M. Green, J. K. C. Tilles, E. H. Kass, and M. Finland. Effect of intranasal reovirus infection on antibacterial activity of mouse lung. J. Infect. Dis. 119:
43-50, 1969.
13. A. W. Gledhill, D. L. J. Bilbey, and J. S. F. Niven. Ef- fect of certain murine pathogens on pathocytic activity.
Br. J. Exp. Pathol. 46: 433-442, 1965.
14. A. L. Notkins. Lactic dehydrogenase virus. Bacteriol.
Rev. 29: 143-160, 1965.
15. G. D. Bonnard, E. K. Manders, D. A. Campbell, R. B. Herber- raan, and M. J. Collins. Immunosuppressive activity of a
subline of the mouse EL-4 lymphoma. Evidence for minute virus of mice causing the inhibition. J. Exp. Med. 143:
187-199, 1976.
16. V. Riley. The LDH-virus: An interfering biological con- taminant. Science 200z 124-126, 1978.
17. G. L. Jakak and G. M. Green. The effect of sendai virus infection on bacteriocidal and transport mechanisms of the mouse lung. J. Clin. Invest. 51: 1989-1998, 1972.
18. D. Boraschi and M. S. Meltzer. Macrophage activation for tumor cytotoxicity: Genetic variation in macrophage tu- mor ici dal capacity among mouse strains. Cell. Immunol.
45: 188-194, 1979.
