Natural cytotoxicity in rats: Radiation-induced changes in the early killing of allogeneic cells

CELLULAR

78, 206-216 (1983)

IMMUNOLOGY

Natural Cytotoxicity in Rats: Radiation-Induced Changes in the Early Killing of Allogeneic Cells L. JANE MCNEILAGE’ AND BARBARA F. HESLOP Department of Surgery, University of Otago Medical School, Dunedin, New Zealand Received December 20. 1982; accepted February 10. 1983

In somestrain combinationsamong inbred rats intravenously injected s’Cr-labeled lymphocytes are rapidly destroyed in substantial numbers by unsensitized allogeneic hosts.This phenomenon has been referred to as natural cytotoxicity (NC) and is characterized by decreased lymph node radioactivity, increased kidney and urine radioactivity, and to a lesser extent increased liver radioactivity in allogeneic hosts, when compared with the distribution of label in syngeneic recipients of the same cell suspension.A single exposure to 800 rad either 1 or 7 days before the injection of 5’Cr-labeled lymphocytes effected a reduction in NC as defined by all the above parameters in a strain combination exhibiting high NC. The same dosage of radiation abolished NC in a strain combination exhibiting intermediate - low NC. BecauseNC was not always completely abolished, the phenomenon was held to be partially radiosensitive. An increasedaccumulation of 5’Cr-labeled lymphocytes wasseenin the lymph nodesof both syngeneic and allogeneic irradiated hostswhen compared with nonirradiated controls, although the increase was greater in allogeneic than in syngeneic hosts. This increased colonization in the lymph nodes of irradiated hosts seemedunlikely to be due to an increase in the available “space” in the lymph nodes following irradiation. INTRODUCTION

Examination of the distribution of intravenously injected radiolabeled lymphocytes in both syngeneic and allogeneic hosts has revealed a reduced accumulation of cells in the lymph nodes of allogeneic hosts (l-lo). Studies utilizing a rat strain combination (AS - HS) exhibiting a pronounced syngeneic-allogeneic difference with respect to lymph node localization of the injected lymphocytes showed that those cells which failed to reach the lymph nodes in allogeneic hosts were rapidly eliminated (within a few hours) and that destruction of these cells required an intact spleen (9). This phenomenon was referred to as natural cytotoxicity (NC). It was characterized by reduced lymph node radioactivity, increased kidney and urine radioactivity, and occasionally increased liver radioactivity in allogeneic hosts, when compared with the distribution of label in syngeneic recipients of the same cell suspension (11). Since NC involves the rapid killing of allogeneic lymphocytes by unsensitized hosts, it has features in common with other natural resistance systems,namely natural killer (NK) cell activity, F1 hybrid resistance to hemopoietic cells and in vitro F1 hybrid antiparent cell-mediated lympholysis (12). One of the unusual characteristics of these ’ Presentaddress:The Walter and Eliza Hall Institute of Medical Research,Post Office, Royal Melbourne Hospital, Victoria 3050, Australia. 206 0008~8749/83$3.00 Copyright All

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EFFECT

OF RADIATION

ON NATURAL

CYTOTOXICITY

207

reactions is their strongly radioresistant nature (13). The aim of the present study was to examine the radiosensitivity of the mechanism(s) controlling the early killing of “Cr-labeled lymph node cells in unsensitized allogeneic rats. MATERIALS

AND METHODS

Animals. Young adult rats (2-6 months) of the conventionally maintained AS, HS, DA, and AS2 inbred strains and (AS X HS)FI hybrids were used. The AS and HS strains are compatible at the major histocompatibility complex (RTl), expressing the RT 1’ haplotype, while DA and AS2 express the RTla and RTl’ haplotypes, respectively ( 14). Cell suspensions. Single-cell suspensions were prepared in the manner described previously (9). Briefly, pooled cervical, axillary, mesenteric, aortic, and iliac lymph nodes were disrupted by pressing gently with a gloved finger through a stainless steel sieve and were washed twice in cold (4°C) Hanks’ balanced salt solution (HBSS) supplied with 10% fetal calf serum (FCS). After filtration through stainless steel mesh the resulting cell suspension was adjusted to a final concentration of approximately 5 X 10’ cells/ml. Contaminating erythrocytes were removed on a Ficoll-Paque (Pharmacia Chemicals, Uppsala) gradient. The resultant lymphocyte layer was washed three times in HBSS + 10% FCS and resuspended at a concentration of 10’ cells/ ml. The cells were labeled with sodium [“Crlchromate in sterile isotonic saline (Radiochemical Centre, Amersham) for 30 min at 37°C using 40 &i/ml. Following incubation with the label the cells were washed a further three times and resuspended to a final concentration of 2 X lo* cells/ml for injection. Samples(50 ~1)were retained for determination of the total label injected. Experimental design. Labeled lymph node cells (2 X IO’) were injected into groups of allogeneic and syngeneic (control) hosts via an exposed lateral tail vein. Recipients were sacrificed 24 hr after injection and the radioactivity was counted in the pooled lymph nodes (described above), spleen, liver, kidneys, thymus, lungs, and a 500-~1 sample of blood. After subtraction of the background level, the radioactivity in a given organ was expressedas a percentage of the total dose injected. irradiation. Rats were exposed to whole-body irradiation in a 13.5-cm2 well, 35 cm from a 6oCosource (energy 1.251 MeV). The dose rate was 668 rad per minute, and rats received a total dose of 800 rad. Animals were kept either 24 hr or 7 days postirradiation before receiving ‘Cr-labeled lymph node cells intravenously. RESULTS (I) The Eflect of Whole-Body Irradiation on the Level of Natural Cytotoxicity Expressed by Allogeneic Hosts Recipient rats comprised four groups-(i) hosts syngeneic with the cell donors, (ii) semiallogeneic hosts, i.e., F, hybrids with one parent from the cell-donor strain and the other parent from a strain exhibiting a high level of NC toward the donor cells, (iii) allogeneic hosts exhibiting a high level of NC (11) against the donor-strain cells, and (iv) allogeneic hosts exhibiting an intermediate to low level of NC (11) toward the donor strain. All animals received 800 rad of whole-body irradiation 24 hr prior to the injection of “0-labeled lymph node cells. Table 1 summarizes the distribution of label in the irradiated hosts and in the appropriate nonirradiated controls. The salient points from the data depicted in Table

No. of rats SEM 2.95’ 1.60 2.34 2.80 3.03 4.06 2.33 3.98

R

119.40 100.61 116.48 51.23 90.08 122.94 89.09 133.48

800 0 800 0 800 800 0 800

SEM 1.53 2.18 1.85 3.55 2.14 4.13 5.51 3.07

R 66.58 102.12 81.41 91.37 55.39 84.77 107.47 75.45

Spleen

90.18 112.59 92.17 112.46 100.73 87.65 105.38 98.29

d

Liver J’? 90.20 131.97 77.82 172.22 133.33 79.27 100.91 73.33

SEM 1.09 3.36 3.17 2.32 1.90 3.26 3.06 3.43

9.57 19.14 12.59 15.93 11.29 1.46 3.67 1.63

SEM

Kidneys

DHost animals were given whole-body irradiation 24 hr prior to receiving %r-labeled lymph node cells. b Data are expressedas percentages of nonirradiated controls. ’ Total radioactivity recovered from the standard set of organs. d The level of lymph node radioactivity seen in the nonirradiated HS hosts was somewhat higher than usual (9, 1I).

AS - AS AS - (HS X AS)F, AS - (HS X AS)F, AS - HSd AS-HS AS2 -+ AS2 AS2 - DA AS2 - DA

Donor-host combination

Lymph nodes

Irradiation dose” (rad)

Radioactivity recovered” from

43.56 78.35 16.49 58.89 41.18 58.35 100.00 54.90

d

R 93.91 104.96 98.00 81.42 84.28 96.26 99.48 100.85

1.38 6.36 1.77 4.27 3.65 2.54 1.34 2.41

2.16

1.64 2.09 2.34 2.49

1.78 1.90

1.58

SEM

Total radioactivity recovered’ SEM

Blood (500~/.d sample)

Localization of “Cr-Labeled Lymph Node Cells in Irradiated Syngeneic, Allogeneic, and Semi-allogeneic Hosts

TABLE I

i2 p 8

ci

5 p % iij

8

EFFECT OF RADIATION

ON NATURAL

209

CYTOTOXICITY

1 are: (i) Lymph node radioactivity (LNR) levels were significantly increased (P < 0.001) in all irradiated hosts compared with their nonirradiated controls. (ii) Irradiation of the host reversed the direction of the syngeneic-allogeneic difference in LNR levels for the donor-host combination exhibiting an intermediate - low level of NC (AS - DA). (iii) While host irradiation significantly increased LNR in allogeneic hosts exhibiting a high level of NC toward donor cells (AS - HS), the syngeneic-allogeneic difference was reduced rather than eliminated. (iv) Blood radioactivity levels were significantly decreasedin all irradiated hosts. (v) Host irradiation produced a syngeneic-allogeneic difference in splenic radioactivity levels not seen in nonirradiated hosts. (vi) Irradiation produced a significant decreasein hepatic radioactivity in all donor-host combinations except AS2 - DA. (vii) Host irradiation significantly decreased kidney radioactivity levels in all irradiated hosts except the AS - AS donor-host combination. Some of the AS and HS hosts were housed in metabolic cagesfor the 24-hr period after the injection of “0-labeled AS lymph node cells. Table 2 shows the radioactivity recovered from the urine of these rats. Irradiation produced a significant decrease (amounting to 9% of the total dose injected) in the level of radioactivity excreted by the allogeneic (HS) hosts but did not alter the level of radioactivity present in the urine of the syngeneic (AS) hosts. Cell death is accompanied by the rapid excretion of label in the urine (1, 9). Therefore, the decrease in the excretion of label by irradiated allogeneic hosts suggestsa decreasein the number of cells killed by these hosts. This notion is consistent with the concomitant decreasein kidney and hepatic radioactivity observed in these animals. Redistribution of radioactivity (and hence cells) between various organs, for example, the spleen and the lymph nodes, has been observed in previous studies (2, 9, 15, 16) and was quantitated by Anderson et al. (17) as a “relocalization index” (RI). A relocalization index of unity indicates no movement of counts between the two organs considered. Table 3 gives relocalization indices calculated for irradiated and nonirradiated hosts from Table 1. The tables indicate that: (i) Irradiation produced significant movement of label from the blood to the lymph nodes accompanied by a small amount of relocalization of label from the spleen, liver, and kidneys to the lymph nodes in syngeneic hosts. (ii) Irradiation of semiallogeneic hosts resulted in considerable relocalization of label from the blood to the lymph nodes (RI = 5.52), along with smaller movements of label from the kidneys, spleen, and liver to the TABLE 2 Excretion of Label by Irradiated Hosts Total urine radioactivity for 24 hr” Donor strain

Host strain

No. of rats

/f

SEM

AS AS AS AS

AS ASh HS HSh

3 3 3 3

9.20 8.88 28.62 19.64

0.38 0.83 1.51 0.78

” Expressed as a percentage of the total dose injected. h Hosts received 800 rad whole-body irradiation 24 hr prior to cell transfer.

210

MC NEILAGE AND HESLOP TABLE 3

Changes in the Distribution of Cell Label Produced by Whole-Body Irradiation of Host Animals Relocalization indexh for detection of movement of label from Donor strain

Host strain pair”

Spleen to Blood to Kidneys to Liver to lymph nodes lymph nodes lymph nodes lymph nodes

Urine to lymph nodes

1.17

1.32

2.15

1.32

I .25

1.45

I .42

5.52

1.97

NDd

HS HS’ I

5.39

2.13

3.24

3.80

4.1-l

AS2

AS2 AS2’ I

1.46

1.40

2.15

1.57

ND

AS2

DA’ DA

2.12

1.63

2.65

2.04

ND

1

AS

AS’ AS

AS

(HS X AS)F, (HS X AS)F, ’

AS

1

1

” In each host pair, both received cells from the same cell suspension. ’ Relocalization indices were calculated using: [mean organ radioactivity (nonirradiated hosts)/mean lymph node radioactivity (nonirradiated hosts)]/[mean organ radioactivity (irradiated hosts)/mean lymph node radioactivity (irradiated hosts)]. ’ Hosts received 800 rad whole-body irradiation 24 hr prior to cell transfer. d ND Urine was not collected from these hosts.

lymph nodes. This is consistent with the large increase in LNR observed in these hosts (seeTable 1). (iii) Label relocalized predominantly from the spleen, urine, and kidneys to the lymph nodes with irradiation of allogeneic hosts expressing a high level of NC toward the donor cells (AS - HS). There was also significant movement of label from the liver and blood to the lymph nodes in these hosts. Relocalization of label from the urine, kidneys and liver suggesteda decreasein the proportion of donor cells killed in these irradiated allogeneic hosts. (iv) Host irradiation also produced a relocalization of label from the kidneys, spleen, and liver to the lymph nodes in the DA allogeneic hosts exhibiting a low - intermediate level of NC toward the AS2 donor cells, again suggesting a reduction in the death of injected cells in these hosts. (2) The Efect of Altering the Timing of Host Irradiation Relative to the Injection of Labeled Cells Rolstad (6) observed that the effect of irradiation on the localization of thoracic duct lymphocytes in allogeneic hosts was dependent upon the timing of host irradiation. Animals irradiated 24 hr prior to cell transfer showed an altered distribution pattern (compared with nonirradiated hosts) such that the usual syngeneic-allogeneic difference in LNR was decreased.Increasing the interval between irradiation and cell injection to 7 days almost returned the values to normal (6). The observation (Table 1) that irradiation of allogeneic hosts exhibiting a high level of NC toward donor lymphocytes (AS - HS) produced a decreasein the syngeneic-allogeneic LNR’ difference has been interpreted by the present authors as

EFFECT OF RADIATION

ON NATURAL

CYTOTOXICITY

211

indicating a decreasein the level of NC expressedby the allogeneic hosts toward the injected lymph node cells as a result of irradiation. This interpretation is supported by the reduction in kidney, urine, and liver radioactivity in the irradiated allogeneic hosts. Since these changesoccurred rapidly it seemedlikely that irradiation was acting on the effector arm of the reaction, therefore, it was of interest to determine the effect of increasing the interval between irradiation and cell transfer. The results depicted in Table 4 show that increasing the interval between host irradiation and cell transfer from 24 hr to 7 days did not alter the decrease in NC seenin the irradiated allogeneic hosts; nor did it alter the changes in cell distribution observed with irradiation of syngeneic (control) hosts. Animals injected 3 days after irradiation also exhibited a similar decreasein the level of NC expressed(unpublished observations). (3) The Efect of Altering the Number of Cells Transferred to Irradiated Hosts The increase in LNR and concomitant decreasein blood radioactivity seen with irradiation of syngeneic hosts suggestedthe possibility that a simple “traffic diversion” was occurring in these animals, resulting in retention of cells in the lymph nodes. Irradiation produces a significant depression in the number of recirculating lymphocytes ( 17-19) as well as striking changes in the lymph nodes themselves, leaving them almost, but not entirely, devoid of lymphocytes within a few days (19). Table 5 shows that a dose of 800 rad whole-body gamma radiation produces a significant decrease(50%) in the weight of the spleen and lymph nodes. This reduction remains constant for 8 days after irradiation, raising the possibility that, by decreasing the number of cells in the lymph nodes, irradiation produces increased space that may be available for occupation by “Cr-labeled lymphocytes. Table 6 shows that the proportion of cells accumulating in the lymph nodes did not vary greatly over a dosage range between 20 X lo6 and 150 X 106. The increase in LNR in irradiated as compared to nonirradiated syngeneic hosts was 5-7% of the total radioactivity injected. Similarly the increase in LNR for irradiated allogeneic hosts over nonirradiated allogeneic hosts was constant over the range of cell numbers investigated, and amounted to approximately 15% of the total dose injected. These results are consistent with observations of other workers who have compared the effects of altering the cell dosage in nonirradiated hosts (2, 3, 20), and make it unlikely that the increase in LNR seenwith irradiation is due primarily to an increase in the space available in the lymph nodes following irradiation. DISCUSSION Other workers (3, 5, 6, 8, 10, 21) have examined the effect of host irradiation on the distribution of injected lymphoid cells (results summarized in Table 7). The majority found that whole-body irradiation maintained the syngeneic-allogeneic difference in LNR observed with nonirradiated hosts (5, 6, 8, 10). Both Viklicky et al. (5) and Rolstad (6) commented that, while the syngeneic-allogeneic difference was not abolished by host irradiation, the localization of cells to the lymph nodes showed a relatively greater increase in allogeneic hosts. Similarly the data of Pincott and Bainbridge (10) reveal an increase in LNR in irradiated allogeneic hosts. All these authors, however, concluded that the mechanism controlling the decreasein lymph

DAd DA’ DA’

AS2 AS2 -

-I 7

7

7 I I

-7

I

Irradiation injection interval’ (days)

133.48 85.34 129.56

117.42

89.39 90.08 122.94

39.15 120.06

119.40

R

Lymph

3.98 I .49 2.00

1.37

2.57 3.03 4.06

2.45 3.63

2.9s

SEM

NS

NS

NS

NS

P’

75.45 119.67 69.42

91.77

51.13 55.39 84.77

92.28 58.72

66.58

1

<0.02

NS


P

98.29 107.16 93.38

107.69

106.25 loo.73 87.65

128.30 93.56

90.18

I

7 days prior to receiving

3.07 7.60 1.69

2.28

1.93 2.14 4.13

2.78 2.28

1.53

SEM

Spkll

cells and hosts irradiated

nodes

a Recipients were-irradiated n days prior to cell transfer. b Expressed as a percentage of non-irradiated syngeneic controls. rTotal radioactivity recovered from standard set of organs. ‘Hats received 800 rad of whole body irradiation. * Nonirradiated allogeneic controls. ’ P = dil?erence between hosts irradiated I day prior to receiving

69 5

5

AS2d

AS2 -

4

5 5 5

HS’ ASd

AS -

5

No. of rats

AS - HS” AS - HS” AS2 - ASZd

A@

AS -

Donor-host combination

cells.

3.43 I .42 2.03

1.93

3.87 I .90 3.26

4.32 3.41

I .09

SEM

Liver

NS

NS

P

NS

1

73.33 105.16 75.11

83.34

147.27 133.33 79.27

172.73 95.13

I .63 2.15 2.72

3.49

8.89 II.29 I.46

6.03 8.99

9.57

SEM

Kidneys

from

90.20

recovered*


Radioactwity

NS

NS

NS

NS

P

54.90 97.98 49.09

60.00

36.52 41.78 58.35

51.09 43.48

43.56

R

2.41 I .43 1.58

1.82

3.16 3.65 1.34

3.15 2.22

7.38

SEM

Lllood (5CWpl sample)

NS

NS

NS

P

100.85 lcnO2 loo.90

107.21

80.85 84.28 99.48

81.29 93.11

93.91

R

2.16 1.01

0.80

1.56 2.09 2.49

1.32 I.61

I .59

SEM

Total radioactivity recovered’

Localization of 5’Cr-Latxled Lymph Node Cells in Syngeneic and Allogeneic Hosts Irradiated Either I or 7 Days Prior to Cell Transfer

TABLE 4

NS

40.0 I

NS

NS

P

8

F

is

5

ii

P

EFFECT OF RADIATION

ON NATURAL

213

CYTOTOXICITY

TABLE 5 The Effect of Exposure to Whole-Body

Irradiationon Organ Weights Qwn weight(9)

Donor-host combination’

No. of cells injected (X 106)

No. of host rats

Irradiation injection intervalb (days)

Lymph nodes

Spleen

1

SEM

PC

d

SEM

P

AS AS -

AS AS*

150 150

4 4

I

0.389 0.233

0.017 0.004


0.351 0.183

0.01 I 0.007

10.001

AS AS -

AS ASd

34 34

4 4

7

0.500 0.276

0.010 0.019


0.362 0.156

0.009 0.015

co.00 I

AS AS -

HS HSd

150 150

4 5

1

0.614 0.288

0.054 0.029

to.0 I

0.546 0.299

0.033 0.029


AS-HS AS - HSd

34 34

4 5

7

0.613 0.342

0.048 0.016


0.656 0.348

0.022 0.012


a Hosts receiving the same number of donor cells were injected with cells from the same cell suspension. ’ Recipients were irradiated n days prior to cell transfer. ‘P = the difference fmm nonirradiated hosts given cells from the same cell suspension. d Hosts received 800 rad whole-body irradiation 24 hr prior to cell transfer.

node localization of injected cells in allogeneic hosts was relatively radioresistant and did not result from a conventional cellular immune response. The results from the present study indicate that host irradiation: (i) increases LNR in both syngeneic and allogeneic hosts irrespective of the level of NC expressed by the nonirradiated control hosts, this increase is unlikely to be due primarily to an increase in the space available in the irradiated lymph nodes; (ii) abolishes NC in allogeneic hosts normally expressing a low to intermediate level of NC toward the injected cells; (iii) decreasesNC (as judged by lymph node, kidney, urine, and liver radioactivity) in allogeneic hosts normally expressing a high level of NC toward the injected cells, but does not abolish it; (iv) produces a relocalization of cells from the blood to the lymph nodes in all irradiated hosts; (v) produces the above effects when administered either 24 hr or 7 days prior to cell transfer. In general, the above results agree with the findings and conclusions of Zatz and her colleagues (3). Some caution, however, needs to be expressedwhen considering either of the above two interpretations of the effect of host irradiation, as it could be very difficult to distinguish between the effects of an irradiation-induced traffic diversion which allowed an increased accumulation of cells in the lymph nodes, when compared with nonirradiated controls. With reduced NC in the absence of a traffic diversion, it seemslikely that increased LNR would be accompanied by a decrease in liver, kidney, and urine radioactivity. On the other hand, one might expect a traffic diversion in the presence of unaltered levels of NC to be associatedwith a reduction in liver, kidney, and urine radioactivity. In the present series, for allogeneic hosts expressing a high level of NC toward the donor cells (AS - HS), the decrease in kidney, urine, and liver radioactivity was greater than the increase in LNR produced by host irradiation (Tables 1 and 2). Bearing in mind the major parameters of NC (decreasedLNR, increased kidney and urine radioactivity, and to a lesser extent increased liver radioactivity in allogeneic

5

4

4 45

6

4 5 4 5 4 5

AS - AS AS - ASd

AS - BS BSd AS --t AS

AS - AS”

AS - HS AS - HS” AS + AS AS - AS” AS-HS AS - HS”

25.13 25.19

9.69

32.43 7.69 22.48

150 I50 I50

33.03

8.47 27.08 22.42

23.38 28.80

R

15.15 20.98

II.67

0.18 0.50

19.21 9.26 22.6 1

12.35

16.78 17.65

22.73

11.94 13.88

d

0.18

0.35 0.24 0.45 0.33 0.25

0.40

0.48 0.65I 0.4

0.44 0.39

SEM

Spleen

0.52 0.34 0.43 0.34

0.31

0.59 0.27 0.40

0.20 0.36

SEM

Lymph nodes

52 52 150

52

20 52 20

20 20

No. of cells injected (X10”)

20.12 19.12 21.38 19.77 21.66 19.68

17.88

22.12 25.01 19.29

23.81

25.85

R

Liver

0.34 0.50 0.25 0.25 0.26 0.39

0.32

0.28 0.32

0.19

0.22 0.34

SEM

2.18 I .25

0.91

1.12

1.39

0.95

0.18

1.53

0.86 1.01

1.22

1.00

x

0.05 0.05 0.03 0.07 0.05

0.01

0.02

0.04 0.03 0.06

0.02 0.63

SEM

Kidneys

0.18

0.22

0.16

0.09 0.07 0.41

0.09

0.007 0.09 0.21

0.08

0.19

x

0.002

0.01 0.01 0.01 0.01 0.0 I

0.003

0.02 0.002

0.01

0.004 0.003

SEM

59.07

56.01

52.30 57.90 74.53 71.35

66.19

57.07 69.29 68.52

71.66

72.60

d

I.19

1.04 0.57 1.98 0.53 0.69

0.58

0.33 0.56 0.66

0.55

0.86

SEM

Total radioactivity recovered‘

” Both syngeneic and allogeneic hosts injected with the same number of cells received cells from the same cell suspension. BS hosts were used instead of HS hosts in one instance. The BS and HS strains exhibit similar levels of NC toward AS (I 1). h Expressedas a percentage of the total dose injected. ‘ Total radioactivity recovered from the standard set of organs. ‘Hosts receiving 800 rad of whole body irradiation, twenty-four hours prior to cell transfer.

No. of rats

Donor-host combination”

Blood (500-~1 sample)

Lymph Node Cells in Syngeneic and Allogeneic Hosts

Radioactivity recovered” from

The Effect of Altering the Dose of Cells Injected on the Localization of “0-labeled

TABLE 6

3 pi $

az

?2 ~

3 $

5

Mice

Rats

Bone marrow

Thoracic duct lymphocytes Thoracic duct lymphocytes Thoracic duct lymphocytes Spleen

Mice Mice Mice Mice Mice Rats Rats Rats Rats Rats Rats

Mice

Rats

400 700 loo0 1000 1000 200 800 800 800 800

950

6OO/Delay“

loo0

850

950

900 850

900

(rad)

Dose of

Cells in Irradiated

Increased Increased Increased Increased Increased Increased Increased Increased Increased Increased Increased

No effect

Small increase

Increased

Decreased Decreased Decreased Decreased Decreased

DeCreased

Increased Increased Increased No effect No effect

NT

Decreased

Decreased

Decreased

Increased

No effect No effect No effect Increased Decreased Decreased Decreased Decreased NT Decreased

NT

No effect

No effect

No effect

No effect

NT

NT No effect

NT No effect

No effect

Liver

No effect Decreased

No effect

No effect’

No effect

No effect Decreased

No effect

Spleen

Effect of Irradiation on radioactivity recovered from

Summary of the Results in the Literature

Lymph nodes

Hosts-A

7

Decreased Decreased Decreased

Decreased

NT NT NT NT NT Decreased Decreased

NT

Increased

Increased

Increased

NT

NT

NT NT

NT”

Blood

(3) (3) (3) (3) (3) This paper This paper This paper This paper This paper This paper

(8)

05)

(6)

(6)

(5)

(8)

(5)

(21)

(21)

Literature reference

’ NT, not tested. b Taken from Table 6, Viklick? el al. (5). ‘No effect as interpreted by Viklicki d al. (5). However, Table 6 shows a significant reduction in lymph node radioactivity for irradiated allogeneic hosts compared with syngeneic control hosts but an increase (statistical significance not given) when compared with nonirradiated allogeneic hosts. d Delay of 7 days.

Allogeneic (CBA-E) (CBA-E) (CBA-E) (E-EK) @K-E) Syngeneic (AS-AS) (AS-AS) (ASZ-AS2) Allogeneic (AS-HS) (AS2-HS) (AS-(HS X AS)F,)

Allogeneic (BIO.D2BlO.BR)

(DA-HO)

node node node node node node node node node node node

Mice

Spleen

Lymph Lymph Lymph Lymph Lymph Lymph Lymph Lymph Lymph Lymph Lymph

Mice Mice

Thymus Bone marrow

Rats

Mice

Lymph nodes

Syngeneic (CBACBA) (CBA-CBA) Syngeneic (B I OBIO)b Syngeneic (BIO.DZBIO.D2) Allogeneic (B IOBIO.A)b Allogeneic (DA-HO)

(DA-HO)

Animal

Type of cells injected

of Lymphoid

Donor-host combination

Localization

TABLE

N t;

3 ‘3

3

2

z

F

4 B 2 z=!

g

3 3

m

216

MC NEILAGE AND HESLOP

as compared with syngeneic hosts (9, 1l), the effect of irradiation is most plausibly explained as representing a decreasein the level of NC. Since NC is reduced rather than abolished, it seemsthat NC is at most partially radiosensitive. The effect of host irradiation was quite different in the syngeneic donor-host combination. Urine and kidney levels remained constant implying that the number of injected cells dying in the irradiated syngeneic hosts was the same as in the nonirradiated controls. There was considerable movement of label from the blood to the lymph nodes (Table 3), suggesting that host irradiation in the syngeneic situation produces a traffic diversion alone. Experiments examining the effect of host irradiation on the distribution of injected lymphocytes in syngeneic and allogeneic hosts need to be carefully controlled, with results of transfers in irradiated hosts being compared with similar transfers in nonirradiated controls. Decisions concerning the dose of irradiation, the time of irradiation relative to the injection of labeled cells, the source of radiation (e.g., X rays versusgamma rays) may unwittingly influence the results and subsequent conclusions. These factors, together with speciesdifferences, may explain the apparently differing effects of irradiation on phenomena which appear to be analogous to NC (Table 7). It is, however, also likely that some of the conflicting results represent differences in the interpretation of what may well be fairly similar experimental results. ACKNOWLEDGMENTS We thank Karen Skinner and Anne Marie McCarthy for their competent technical assistance.This work was supported by the Medical Research Council of New Zealand.

REFERENCES I. 2. 3. 4. 5. 6. 7. 8. 9. 10. 1I.

Bainbridge, D. R., and Gowland, G., Ann. N.Y. Acad. Sci. 129, 257, 1966. Heslop, B. F., and Hardy, B. E., Transplant&ion 11, 128, 197 I. Zatz, M. M., Gingrich, R., and Lance, E. M., Immunology 23,665, 1972. Frost, H., Cahill, R. N. P., and Trnka, Z., Eur. J. Immunol. 5, 839, 1975. Viklicky, V., Peknicova, J., and PolackovL, M., Folia Biol. (Prague) 22, 159, 1976. Rolstad, B., Cell. Immunol. 45, 389, 1979. Vikhcky, V., and Mickovi, M., J. Immunogenet. 6, 245, 1979. Degos, L., Pla, M., and Colombani, J. M., Eur. J. Immunol. 9, 808, 1979. McNeilage, L. J., and Heslop, B. F., Cell. Immunol. 50, 58, 1980. Pincott, C. E., and Bainbridge, D. R., Eur. J. Immunol. 10, 250, 1980. McNeilage, L. J., Heslop, B. F., Heyworth, M. R., and Gutman, G. A., Cell. Immunol. 72, 340, 1982. 12. Moller, G., (Ed.), Immunol. Rev. 44, 1979. 13. Hochman, P. S., Cudkowicz, G., and Dausset, J., J. Nat. Cancer Inst. 61, 265, 1978. 14. Stark, O., Kren, V., and Gunther, E., Transplant. Proc. 3, 165, 1971. 15. Sprent, J., Cell. Immunol. 7, 10, 1973. 16. Woodruff, J., and Gesner, B. M., Science 161, 176, 1968. 17. Anderson, R. E., Sprent, J., and Miller, J. F. A. P., Eur. J. Immunol. 4, 199, 1974. 18. Sprent, J., and Miller, J. F. A. P., Cell. Immunol. 3, 385, 1972. 19. Anderson, R. E., and Warner, N. L., In “Advances in Immunology” (F. J. Dixon and H. G. Kunkel, Eds.), Vol. 24, p. 215. Academic Press,New York, 1976. 20. Taub, R. N., and Lance.,E. M., Transplantation 11, 536, 1971. 2 I. Lance, E. M., and Taub, R. N., Nature (London) 221, 841, 1969.