J. F. Loutit

Induction of hypoxia in normal and malignant tissues by changing the oxygen affinity of hemoglobin—Implications for theraphy

The drug BW12C, which increases the oxygen affinity of hemoglobin, reduces oxygen availability to tissues. This results in protection against radiation damage to the hemopoietic system and epidermal Langerhans cells in CBA mice. The drug also protects against beta-irradiation damage in pig epidermis. BW12C increases the hypoxic cell fraction in tumors and histological examination of an experimental T cell lymphoma shows that the induced hypoxia leads to tumor necrosis.

Osteopetrosis of microphthalmic mice—A defect of the hematopoietic stem cell?

The recessive genesmi andgl in the homozygous state determine, among other phenotypic effects, osteopetrosis in the house mouse. From a stock carryingmi derived from Grüneberg (1963) themi gene was bred into the standard CBA/H inbred strain. Microphthalmic mice of these two stocks and their hybrids were treated as newborn by intraperitoneal injection and at weaning or maturity by intravenous injection of cell suspensions containing hematopoietic stem cells from phenotypically normal mice. Resolution of much of the osteopetrosis but not the other phenotypic effects occurred within a few months in the majority of cases, provided syngeneic or H-2 compatible allogeneic cells were given: it did not occur spontaneously or on giving H-2 incompatible cells or on giving compatible material by an inappropriate route. The results accord with hypotheses that (1) osteoclasis of scaffold-type woven bone is impaired inmi mi, (2) that osteoclastic cells are derived through circulating monocytes from hematopoietic stem cells, and (3) inmi mi this defect can be overcome by a transplant of normal hemopoietic stem cells.

Histocompatible Cells for the Resolution of Osteopetrosis in Microphthalmic Mice

Harwell microphthalmic mice from a small closed line-bred colony, when separated after 2 weeks parabiosis with normal siblings, showed later resolution of their osteopetrosis (except for jaws and tail). The former parabionts were tolerant of each other’s skin. This mutual tolerance was shown by skin grafts to be due to reduced heterozygosity of the stock, not to parabiosis. Such tolerance would allow acceptance by the osteopetrotic mice of precursors of normal bone resorbing cells from the sibling’s blood. Despite skeletal improvement the affected mice remained unthrifty.

Diurnal variation in urinary excretion of calcium and strontium

A marked variation in the 4-hourly urinary excretion of calcium and strontium was demonstrated in two male subjects on normal diet and activity. The general pattern of excretion was not, however, the same in the two who differed also in their pattern of response to a regular dietary intake of milk 4-hourly. Further experiment on one subject showed that on a regular 4-hourly diet of his normal calcium content a regular rhythm for calcium excretion was manifest only when activity was also controlled by his confinement to bed. Excretion of calcium was correlated, but not undissociably, with that of sodium. A similar daily rhythm was elicited by a diet low in calcium ( < 100 mg/day). Renal clearance of calcium and strontium ran parallel with a variation of up to sevenfold within a 24 h period. The results indicate a natural nycthemeral* variation in renal excretion of calcium and strontium, but this natural rhythm may be obscured by extrinsic factors, such as muscular exercise and loading of an internal reservoir such as the ‘exchangeable pool’.

Immunological Status and Longevity of Radiation-Chimaeras

(1) Homologous radiation-chimaeras, produced by irradiating CBAH mice with 950 rads of X-rays and resuscitating with AH myeloid tissue, usually die within 4 months of ‘secondary disease’. Bone marrow from adult AH mice is more strongly productive of ‘secondary disease’ than spleen from 5 to 10-day-old AH infants. (2) Secondary disease was not seen when C3HH infant spleen was given to restore C3H x 101. F1 irradiated mice or the F1 infant spleen to irradiated C3HH. (3) When AH, CBAH and CBA x A. F1 were each used as irradiated recipients and donors of adult bone marrow, the F1 approximated much more closely to a universal donor than universal recipient for the avoidance of secondary disease. (4) Those animals from the preceding group which survived 6 months or more were skin grafted with the three types of skin. The responses of the chimaeras in terms of ‘take’ of these skin grafts were the sums of responses of host and donor elements of the chimaeras. The evidence from these and other experiments is reviewed with the conclusions (1) that in homologous chimaeras the host is tolerant of the donated grafts of myeloid tissue, (2) that if the histoincompatibility between graft and host is marked the graft may react against the host to produce ‘secondary disease’; (3) that, if the histoincompatibility is slight or if immature myeloid tissue is given, ‘secondary disease’ is attenuated-the roles of ‘partial tolerance’, ‘immunological paralysis’ and ‘enhancement’ of the graft versus host reaction are discussed.

Histochemical phosphatases and metachromasia in murine tumours induced by bone-seeking radionuclides

Tumours induced in mice, either CBA normal and chimaerical, or C3H, by 90Sr or 226Ra or plutonium have been examined histochemically with (1) diazotate fast red violet LB salt in naphthol AS-MX phosphate buffer at pH 8·6 and 5·2, (2) 1: 9 dimethyl methylene blue (Taylor).It is concluded:(a) The diagnosis of osteosarcoma is facilitated with Taylors Blue which stains osteoid metachromatically. Cells of osteosarcoma, like normal osteoblasts, contain alkaline phosphatase but this may be lost by mutation either in the original tumour or subsequently on passage of the tumour serially to compatible hosts.(b) Osteosarcomata may contain giant-cells of two forms, bizarre tumour cells and osteoclasts; the latter contain acid phosphatase. Osteosarcomata which retain their osteoid on serial passage have few cells containing acid phosphatases.(c) Primitive mesenchymal cell tumours of angiomatous form may occur, if the bone marrow is irradiated, e.g. by 90Sr-90Y and Pu. These tumours lack osteoid and cells interpretable as osteoblasts or osteoclasts (though they destroy bone).(d) Tumours classifiable as fibrosarcomata occur rarely, and may be truly of fibroblastic origin or be mutated osteosarcomata.(e) Lymphomata also occur when the marrow is irradiated (90Sr-90Y and Pu). They may be generalized, when their cells may contain alkaline phosphatase or lack it. They may be localized to abdominal viscera, the reticulo-sarcomatous form, in which case the cells lack alkaline phosphatase.

BIOCYCLES IN THE RETICULOENDOTHELIAL SYSTEM

The reticuloendothelial system (RES) is difficult to define with any degree of accuracy, and I shall not attempt to do so. There appears, however, to be a general consensus that the network of reticulum cells in certain tissues, such as spleen, bone marrow, and lymphoid tissues, together with the littoral cells of the sinuses in those same tissues and in the liver, constitute a large part of it. These cells are derived from the primitive mesenchyme, with the possible exception of those in that peculiar lymphoid mass, the thymus, the epithelial elements of which represent an outgrowth of primitive endoderm; also, some of its reticular cells are credited with a similar origin. In addition, descendent cells of the primitive mesenchyme, particularly in the neighborhood of blood vessels, can be included in the RES. This results in the anatomical representation of the RES virtually all over the body. Functionally, the system is best identified by its capacity for scavenging debris, hence its alternative name of the macrophage system (Maximov and Bloom, 1952). It is thus intimately concerned with the physiological mechanisms of the body for the removal of effete cells. It is an interesting thought that the free-living individual relying on his own resources has a certain amount of tissue as capital equipment built up in embryonic life from nutrients supplied by a maternal host and an additional quota of cells that are expendable and renewable. Almost all of these consumable stores are epithelia, that is, lining surfaces that are shed externally. The skin is an obvious example. The detritus of the mucosae of the respiratory tract is passed up from the bronchi or down from the nasopharynx by ciliary action to be swallowed. It then joins the cells shed from the alimentary tract. Some of this debris from the alimentary tract (which must be considerable if the time for complete turnover is about 2 days, according to Leblond and Walker, 1956) is presumably available for digestion with reabsorption of the breakdown products; some is merely discharged in the fecal waste. Epithelia shed from the genitourinary tract are similarly lost to the outside world. Even the products of the germinal epithelium finally leave the body proper. Thus all this valuable material is not available for re-use or, if it is, only after digestion to its simplest conslituents. The cells as capital equipment, be they of ectodermal origin such as the nervous system, of endodermal origin such as liver parenchyma, or of mesoblastic origin, are tucked away in internal sites. Then there is either no further turnover of cells, as alleged for nervous tissue, or replication only following abnormal loss of cells as in the hepatic parenchyma following necrosis (in fact, repair of most lesions is effected by fibroblastic proliferation). * Bicycle, that commonly used term for an acrobatic means of locomotion on two wheels, is a misbegotten bastard of Latin (bi-) and Greek (KUKAOS). Furthermore, it cannot be used for a cycle pertaining to life (B-m).

ROLE OF THE THYMUS IN THE RADIATION CHIMERA

Burnet (1962) postulated: ( I ) that the thymus may be the site where differentiation of lymphocytes into clones with definable immunological function occurs, and (2) that forbidden clones reactive against self might be eliminated there. If the latter were the case, it would be expected that thymectomized mice, recovering from lethal doses of X-rays under the therapeutic influence of intravenously injected hemopoietic cells, would be more likely, than intact mice, to show effects attributable to such forbidden clones. Like graft-versus-host reactions in radiation chimeras, this should lead to a “secondary syndrome” with dysplasia or aplasia of the lymphatic system. We (Barnes et al . , 1962) have already shown that CBA/ H mice may manifest a n aplasia of the lymphatic tissue when the restorative hemopoietic inoculum consists of syngeneic cells from either: ( 1 ) fetal liver or ( 2 ) adult bone marrow in suboptimal amounts or ( 3 ) adult bone marrow repeatedly passaged within lethally X-rayed mice. The present experiments were set up to see if thymectomized CBA/H mice were more prone than intact mice to this form of secondary syndrome. This is most readily scored in living animals as a secondary loss of weight, which is associated clinically with a decline in their general well-being and histologically, with dysplasia of the lymphatic tissues. While the work was in progress, Miller, Doak. and Cross (1963) reported comparable experiments, which indicated that their thymectomized CBA mice under these circumstances, like neonatally thymectomized mice, failed to dcvelop an adequate structure and function of the lymphatic tissues.

Safe Limits of Occupational and General Exposure to Radiation

The biological effects of radiation are reviewed from the standpoint of safe limits of occupational and general exposure to radiation. lt is pointed out that many people are exposed to small doses of radiation through their occupations. Those occupationally exposed are all over age 18 and under the supervision of competent medical authorities. The principal limiting factor as far as exposure of the general population is concerned is the genetic problem of inheritance, and this must be considered carefully in establishing limits for radiation exposure. (C.H.)