Immunology, I960, 3, 244.

The Effect of Site of Implantation on Host Reaction toOvarian Homografts

DELPHINE M. V. PARROTT*

Division of Experimental Biology, National Institute for Medical Research,Mill Hill, London, N. W. 7

Summary. A method of orthotopic ovarian grafting in a stock of hooded rats hasbeen described. Intra-strain grafts were established in ten out of eighteen animalsand six of these were fertile. Four fertile rats were killed during their first pregnancybut two others kept for I 20 and 154 days had several litters of normal size.When ovarian tissue from donors from a stock of albino rats was implanted

orthotopically into hooded recipients all the grafts were overwhelmed by homo-graft reaction within 24 days. However, both inter- and intra-strain grafts per-sisted in the subcutaneous site.

Factors affecting the establishment and fertility of the intra-strain orthotopicgrafts have been discussed and an explanation for the difference in host response toincompatible grafts according to the site of implantation has been suggested.

INTRODUCTION

Harris and Eakin (I 949) described an extensive series of experiments with ovarianhomografts in two strains of rats. They recorded the number of 'takes' and the endocrinefunction over a period of 3 months of subcutaneous grafts with various combinations ofdonors and recipients. The percentage of 'takes' was almost the same in sibling and intra-strain grafts, about 8o per cent, but was reduced in the inter-strain grafts to 35 per cent.They noted, however, that some inter-strain grafts survived for as long as 3 months in goodcondition. Parkes (I 956) carried out a similar series of experiments of longer duration,using albino and hooded strains of rats. He showed that the proportion of 'takes' in thesubcutaneous site was maximum in intra-strain grafts and only slightly less (i6/20) ininter-strain grafts. Moreover, 7/i8 inter-strain grafts were still functioning endocrinologic-ally I2-I3 months after the grafting operation. By contrast, Krohn (1955) reported thatin his experience inter-strain grafts do not survive more than a few weeks.

However, all these experiments were confined to the assessment of the survival andendocrine function of ectopic grafts; no record of orthotopic grafting in this species hasbeen found in the literature.

In I956 Parrott and Parkes reported a simple method of orthotopic grafting aftersterilization of recipients by X-irradiation. In mice, transfer of ovarian tissue was con-fined to members of the same strain (Parrott and Parkes, i960), but in hamsters ovariesfrom an albino strain were successfully implanted into golden hamster recipients andoffspring were obtained which were genetically identified as deriving from a graft(Parrott, 1959). Inter-strain ovarian grafts also survived in the hamster and functionedin two ectopic sites, the subcutaneous and within the cheek pouch. It seemed possible,therefore, that this technique of orthotopic grafting could also be applied to rats since the

* Present address: Imperial Cancer Research Fund, Burtonhole Lane, Mill Hill, London, N.W.7.

244

Reaction to Ovarian Homografts

same two strains hooded and albino, in which subcutaneous grafts had proved successful(Parkes, I 956) were available and would also provide genetic identification of the progeny.A few of the results presented in this paper have been recorded briefly elsewhere

(Parrott, I 958; Parrott and Parkes, I 960).

MATERIALS AND METHODS

ANIMALS

The albino rats were from a colony that originated in six females and three malesobtained from the Pharmaceutical Society in I946. The hooded rats were from a colonyestablished in I939 with animals obtained from Imperial Chemical Industries. Bothcolonies have been closed since their establishment at this Institute and both have beenmaintained by line-breeding. No form of inbreeding has been practised.

IRRADIATION (i)

In the first experiments (Group ROG) the ovaries of hooded rats were exteriorizedunder anaesthesia and exposed to large doses of irradiation (2000 r. or 4000 r.) whilst therest of the animal was shielded with lead. This method of irradiating the ovaries alonewas described by Mandl and Zuckerman (I956) and was used in some experiments withhamsters (Parrott, I959). These dosages were subsequently found to be excessive for ourrequirements and sometimes caused damage to the ovarian tube. The irradiation pro-cedure was therefore modified and in most of the experiments the recipient rats (GroupSIR) were sterilized by two small doses of whole body irradiation given during infancy.The first dose (200 r.) was administered when they were approximately I week old andthe second (I50 r.) a week later. On each occasion the mother was removed from heroffspring, all of which were irradiated. Each litter was enclosed in a shallow box duringtreatment to prevent the pups shielding one another and all were returned to the nestbefore the mother was replaced in the cage. If two doses of 200 r. were given about 40 percent of the pups died within 4 days of the second dose, but when this was reduced toI5o r. there were no deaths. A single dose of 450 r. or two doses of 300 r. resulted in Iooper cent mortality.

DETAILS OF IRRADIATION DOSAGES

A Newton Victor G.X.io Superficial Therapy Unit was used throughout theseexperiments.

(i) Dose 2000 r. or 4000 r. The dose rate was 200 r. per minute and this was obtainedwith a potential of 8o.5 kV and a current of 6.4 mA. There were no filters.

(ii) 200 r. whole-body irradiation. The dose rate was ioo r. per 2.5 minutes using apotential of 8o kV, a current of 6 mA and a filter of I mm. of aluminium.

OPERATIVE TECHNIQUE

In the intra-strain graft experiments both donor and recipient were from the hoodedstock. In all the inter-strain experiments hooded rats served as recipients and albino ratsas donors, because Parkes (I 956) had shown that this combination was the more successful.Donors of either strain were from 7 to I4 days old and the recipients were between 2 -5months at the time of grafting. The preparation of the graft and the operative technique

245

have already been described in detail in reports on the mouse (Parrott and Parkes, I956),and the hamster (Parrott, I959). The grafted rats were paired with albino males 2-3weeks after the operation. Vaginal smears were taken from all animals every day exceptSunday.

RESULTS

In these experiments X-irradiation has been administered in different ways and indifferent dosages but with the sole purpose of completely sterilizing recipient animals.It is unlikely that the immune response of the rats was significantly affected by theX-irradiation for when large doses were given only the ovaries were exposed, and whenwhole body irradiation was used the doses were small and the animals had at least 2months for recovery. All irradiated rats were tested with males before the grafting opera-tion. Those that were irradiated during infancy became sterile within 3 months oftreatment.

Groups of control irradiated rats that had not received a graft were paired concurrentlywith all grafted animals. None of them became pregnant. In the inter-strain experimentsthe hooded recipients with grafts from albino donors were paired with albino males sothat there would be genetic identification of any offspring. The hooded eye and coatcolour is dominant to albinism and only graft offspring would be albino; any offspringderived from the recipients own ovaries would be hybrids and therefore hooded in colour.

FERTILITY OF INTRA-STRAIN ORTHOTOPIC GRAFTS

Ovarian tissue was implanted into both ovarian capsules of fourteen hooded rats (ROGI I, I2, 22, 23, SIR 2 I-30). A further four animals (SIR 7-IO) were grafted on one sideonly and the sterile ovary on the opposite side was removed. The results obtained aresummarized in Table I. Seven out of fourteen of the rats with bilateral grafts matedwhen paired, and six of these were fertile. None of the four animals with unilateralgrafts mated. Sixteen out of the total of eighteen grafted animals were killed between36-54 days after the grafting operation. Autopsy revealed that eight out of sixteen ratshad grafts that had grown and were established. Four rats were pregnant when theywere killed, each having a small number of foetuses in utero, some of them dead. Thetwo most successful grafted rats were kept for much longer periods (SIR 24, I55 days;SIR 25, I23 days). SIR 24 had three normal litters (9, 8, 6). One graft was found atautopsy. SIR 25 had two live litters each of seven pups and she was pregnant whenkilled; there were large grafts in both ovarian capsules.

Vaginal smears were taken daily from all animals with orthotopic grafts so as to detectmating, by the presence of spermatozoa, and pregnancy by implantation signs. It was notpossible to detect the successful functioning of an established graft by observing thecyclical changes in the cells of the vaginal epithelium because the continuous low level ofsecretion of oestrogen by the sterile host ovary often resulted in persistent cornification ofthe vagina.

HISTOLOGICAL EXAMINATION OF THE INTRA-STRAIN ORTHOTOPIC GRAFTS

Both ovarian sites from all grafted animals were examined histologically. In generalappearance sections of established grafts were very similar to the mouse and hamsterorthotopic grafts (Parrott, I958; Parrott, I959). The small sterile irradiated ovary

246 Delphine M. V. Parrott

Reaction to Ovarian Homografts

contrasted strongly with the large and vigorous graft (Plate I, Figs. I, 2, 3). However,there was evidence of reaction in all established grafts including those from pregnantanimals. In some instances the reaction was limited to a slight plasma cell infiltration andtrivial connective tissue proliferation; in others many follicles had disappeared and hadbeen replaced by mononuclear cells and fibroblasts amongst which plasma cells werepresent in considerable numbers. Although the corpora lutea appeared to be relativelyresistant to host reaction (Plate i, Fig. 4), the mononuclear cells which accumulated in thestroma often extended along the lines ofthe capillary blood vessels into the luteal substance.

TABLE I

THE FERTILITY OF INTRA-STRAIN ORTHOTOPIC GRAFTS IN HOODED RATS

Duration Estimated numberSerial of Established Fertile Total of oocvtes presentNo. experiment graft graft oocyte in normal animal

(days) count comparable in ageto graft tissue.

ROG I 36 4- Did not mate _ (Calculatedfrom12 36 + Did not mate - the figures22 36 + 2nd mating fertile: 3 foetuses published by

found at autopsy - Mandl and23 36 + ist mating fertile: I foetus found Zuckerman, 1951)

at autopsy

SIR 21 43 - -22 54 + + 2nd mating fertile: 2 foetuses

found at autopsy 303 5-6ooo23 43 -24 '55 + + 3 live litters (9, 8, 6) 258 3-400025 I23 + + 2 live litters (7, 7) and 3 foetuses

found at autopsy 2280 3-400026 43 + One infertile mating 214 5-600027 4328 43 -+H 2nd mating fertile: 3 foetuses

found at autopsy 579 5-600029 43 _30 43

Unilateral graftsSIR 7 46

8 469 46 + - Did not mate 381 -6oooI0 46

Counts were made of the numbers of oocytes and large follicles present in some of theestablished grafts using the method described by Green, Smith and Zuckerman (I956).There was a considerable variation in the number of surviving oocytes in the grafts, butall estimates were lower than the calculated number of oocytes present in the ovaries of anormal rat comparable in age to the grafted tissue (Mandl and Zuckerman, 1951).

INTER-STRAIN ORTHOTOPIC GRAFTS

Ovarian tissue from albino donors was implanted in both capsules of ten hooded rats(ROG I, 3, 5, 6, 8, 9, I8, 20, 24, 25), and in one capsule of six hooded rats (SIR I-6).Only four of the rats mated, all within 20 days of grafting; none of the matings was fertile.The animals were killed between 3' to 43 days after the grafting operation but only twoof them had recognizable ovarian elements within graft tissue. In the graft from one rat

247

Delphine M. V. ParrottPLATE I. Intra-strain orthotopic grafts. (SO sterile ovary. OG orthotopic graft.)

FIG. I. Graft and irradiated ovary from pregnantrat killed 54 days after grafting. Note the corporalutea ofpregnancy. (SIR 22: X 20)

FIG. 3. Another section of the same ovarian siteshowing the graft alone. (x I9.4)

FIG. 2. Graft and irradiated ovary from pregnantrat killed 123 days after grafting. There were two

previous litters (SIR 25 X 23-5)

FIG. 4. Corpora lutea of pregnancy in a graftsurrounded but only slightly infiltrated withlymphocytes. (SIR 28: X2I5)

248

Iwil

Reaction to Ovarian Homografts

killed after 34 days there were some corpora lutea and very few oocytes; in another ratkilled at 43 days after grafting there were vestigial remains ofcorpora lutea but no oocytes.In the other fourteen rats there were no remains of any grafted tissue.To determine the time of destruction of inter-strain grafts, ovarian tissue from infantile

albino donors was implanted in both capsules of three groups of five hooded rats. Onegroup was killed at each of the allotted time intervals (7, i6 and 24 days) after grafting,and the results obtained are set out in Table 2.

TABLE 2THE SURVIVAL TIME OF INTER-STRAIN ORTHOTOPIC GRAFTS IN HOODED RATS

Days after grafting 7 i6 24 34No. of animals with some

oocytes in graft tissue 5/5 5/5 1/5 I/Io

All five animals killed 7 days after implantation had surviving grafts but there wasconsiderable variation in histological structure. The graft tissue from one animal wasalmost completely disorganized, follicular structure being replaced by proliferatingconnective-tissue cells extensively infiltrated by lymphocytes. At the other extreme, agraft from another animal showed peripheral accumulations of eosinophils and scantyinfiltration of the stroma with lymphocytes. An intermediate degree of reaction wascharacterized by partial disappearance of follicular basement membranes, and fibro-blastic proliferation with lymphocyte infiltration; eosinophils were not conspicuous.A similar degree of variation was seen in the grafts from five animals killed i6 days

after operation. Some degree of necrosis was observed in the tissue from two animalswhilst in the remainder (Plate II, Fig. I) a proportion of the follicles had been replaced byproliferating connective-tissue cells infiltrated with lymphocytes.

Grafts remained in only one rat killed 24 days after operation. A few anovular folliclesembedded in connective tissue infiltrated with lymphocytes remained in one graft (Plate II,Fig. 2); on the other side, small accumulations of luteal cells could still be identified andlymphocytes were not very numerous (Plate II, Fig. 2).

INTER- AND INTRA-STRAIN SUBCUTANEOUS OVARIAN HOMOGRAFTS

The results obtained when grafting ovarian tissue orthotopically into X-irradiatedrecipients differ considerably from those obtained by Parkes (I 956) who implanted ovariantissue subcutaneously in spayed recipients. It was considered possible that the presence ofthe host ovary, although sterile, might have influenced the fate of an inter-strain graft bycompeting for available gonadotrophin. This was investigated by implanting ovariantissue into the flank of three groups of ten hooded rats, sterilized by X-irradiation or byspaying (Table 3). Vaginal smears were taken daily from spayed recipients to assess theendocrine function of the subcutaneous grafts. No smears were taken from the X-irradi-ated hosts as previous experience with orthotopic grafts had shown that in the presence ofa sterile ovary the endocrine activity of a graft could not be detected.The results obtained (see Table 3) confirm in general the findings of Parkes (I956). A

high proportion of intra- and inter-strain homografts became established and wereendocrinologically functional in the subcutaneous position and the persistence of a sterilehost ovary in no way affected the results. The successful subcutaneous inter-strain graftsin the irradiated rats (SIR 2I-30) were subsequently transferred to the orthotopic site inthe same hosts (see below).

249

Delphine M. V. ParrottPLATE II. Inter-strain orthotopic grafts.

FIG. I. Section of graft tissue from rat killed 16days after operation. Note connective tissue over-growth and lympho cytic infiltration.(SIR 48: x 174)

FIG. 3. Inter-strain graft. Some normal folliclespersist; the stroma contains scattered mono-

nuclear and plasma cells. (SIR i8: X i60)

FIG. 2. Section of graft tissue from rat killed 24days after operation. A few anovular folliclesremain embedded in proliferating connectivetissue infiltrated with mononuclear cells.(ROG I3: X I88) Subcutaneous grafts.

FIG. 4. Intra-strain graft. The section shows acluster ofoocytes and luteal cells. (SIR 33: X I 60)

250

Reaction to Ovarian Homografts

HISTOLOGICAL EXAMINATION OF SUBCUTANEOUS GRAFTS

At autopsy, the interstrain grafts were seen to be embedded in a thick layer of putty-coloured fibrous tissue in which blood vessels could not readily be distinguished. In contrast,the intra-strain grafts were only lightly encapsulated and appeared hyperaemic. All theestablished grafts were examined microscopically. There were normal ovarian elementsin all the grafts but all showed some evidence of homograft reaction similar to that seen inthe intra-strain orthotopic grafts; however, there was a quantitative difference in theextent of disorganization within the inter- and intra-strain subcutaneous grafts. Thegeneral appearance of the inter-strain grafts was of a few normal elements, includingoocytes surviving in a stroma of fibrous tissue infiltrated with mononuclear and plasmacells (Plate II, Fig. 3). The intra-strain grafts were more normal and a slight plasma celland lymphocyte infiltration was often the only sign ofhomograft reaction (Plate II, Fig. 4) .

TABLE 3

INTER- AND INTRA-STRAIN SUBCUTANEOUS GRAFTS

Duration of No. withSerial experiment No. with endocrine activity some oocytes inNo. (days) graft tissue

Inter-strain Recipients ovariectomizedROG 47-56 30 7/IO 7/10

Inter-strain Recipients X-irradiatedSIR I1-20 30 7/1O

Intra-strain Recipients X-irradiatedSIR 31-40 30 8/sO

The established inter-strain subcutaneous ovarian grafts from seven irradiated rats(SIR 2I-30) were removed whilst the animals were anaesthetized. Half of the graft waskept for histological examination. The thick fibrous layer was removed from the rest ofthe graft, which was transferred to the ovarian capsules of the same hosts. None of theseanimals mated when paired. All were killed 32 days after orthotopic grafting and only twoout of the seven rats had any graft tissue left and this consisted solely of luteal tissue.Transfer of an established subcutaneous graft to the orthotopic site therefore resulted inrapid destruction of all graft tissue.

DISCUSSION

A successful method of grafting ovarian tissue to the orthotopic site in the rat has beendescribed. Two rats kept for I 20 and I 55 days had several litters ofnormal size. Four otherfertile grafted rats were killed during their first pregnancy and the numbers of foetusesfound in utero were small and some of these were dead. Foetal losses during pregnancy andsmall litters have been reported in mice with orthotopic ovarian grafts (Parrott andParkes, i960), but were less frequent in the experiments with hamsters (Parrott, I959).The number ofoocytes in most of the grafts was below iooo; Green, Smith and Zuckerman(1 956) observed a marked reduction in the number of oocytes even in autotransplants. Inthe present experiments host reaction has also contributed to the destruction of oocytes.

25I

It is probable that the fertility of the recipient animals was related to the total numberof oocytes and the number of mature follicles within the orthotopic grafts, although noconclusions can be drawn from the small number of counts available in these experiments.Ingram (I 958), who reduced the fertility of rats by graded doses of irradiation, found thata reduction in litter size was most marked when the total oocyte count fell below i000.The pre- and post-implantation loss of foetuses was probably also a consequence of thesmall number of mature ova shed by the grafts, for Frazer (I955) has shown that foetalloss in rats both before and after the ninth day was much more severe when the numberof implanted ova was between i and 6 than when the normal number of 6 to i o ovawas implanted.The hooded strain of rats used in these experiments exhibited a wide range of immune

response from maximum to very weak. Approximately half of the intra-strain grafts weredestroyed rapidly and even in the established grafts there was some sign of host reaction;but it was impossible to decide how long any individual graft would have survived theimmunological onslaught if its host had not been killed.

Certainly, grafts have survived in two rats kept for I 20 and I 55 days after grafting and,in spite of considerable homograft reaction, many grafts continued to function bothendocrinologically and gametophorically without the development of cystic follicles suchas Parkes (I 956) has observed in long-term subcutaneous grafts. Host reaction to the graftswas often concentrated round the oocytes, and although there were lymphocytes close tothe corpora lutea these appear to be the most resistant elements in an ovarian graft. Itseems unlikely, therefore, that any foetal death (see above) was due to destruction of thecorpora lutea of pregnancy by immune reaction.The degree of incompatibility between the two strains of rats, hooded and albino,

appears to be considerable, almost all the inter-strain orthotopic grafts being overwhelmedwithin 24 days of grafting. Host reaction was rapid and resulted in a replacement of thegraft by fibrous tissue.

It might be considered that the proportion of established intra-strain orthotopic grafts(io/i8) was high, for the stock of rats used was not inbred. Similar results with ovariangrafts, however, have been obtained with two outbred stocks and a hybrid stock of mice(Parrott and Parkes, i960). Billingham and Parkes (I955) compared the transplantabilityof subcutaneous ovarian grafts and skin homografts in the albino stock of rats. They notedthat in some animals the skin grafts survived longer than expected, although not as longas the subcutaneous ovarian grafts, and suggested that some degree of inbreeding of thestock might have occurred by chance. More recently, McQuarrie, Kim and Varco(I959) have demonstrated a difference in the rate and severity of the rejection process ofinter- and intra-strain skin grafts in two outbred stocks of rats. Their findings were similarto those obtained in the present experiments with orthotopic ovarian grafts. They observedthat all the inter-strain skin grafts became hard and were sloughed off within I5 days. Afew of the intra-strain grafts were rejected in the same way as the inter-strain grafts, andalthough the majority of the grafts did not regrow hair, complete rejection was generallydelayed for some time; however, in a small proportion of the animals grafts appeared tosurvive indefinitely with continued healthy hair growth. McQuarrie et al. (I959) alsosuggested that sufficient fortuitous inbreeding can occur in a closed colony for graft survivalto be favoured in some animals. Another explanation of the long-term survival of ovarianas compared with skin grafts is that ovarian tissue has fewer genetically defined 'histo-compatibility' antigens than skin (Billingham and Parkes, 1955; Krohn, I959).

Delphine M. V. Parrott252

Reaction to Ovarian Homografts 253

The immune response to grafts implanted orthotopically differed considerably fromthat evoked by those placed in the subcutaneous site. The appearances in the subcutaneousgrafts were similar to those described by Harris and Eakin (I949) and Deanesly (1956),but, as already stated, there was a quantitative difference in the extent of disorganizationbetween the intra- and inter-strain series. Although the animals with subcutaneous graftswere only kept for 30 days in these experiments, in view of earlier experience (Parkes,I956) it is probable that the grafts could have survived in situ for a much longer period.In contrast the inter-strain orthotopic grafts were destroyed within 24 days and establishedsubcutaneous grafts were also rapidly destroyed after transfer to the orthotopic site. It isdifficult to understand why alteration in the site of implantation should increase sodramatically the rate of destruction. However, it was noted that in the subcutaneoustissue the inter-strain grafts were surrounded by a relatively avascular connective-tissulecapsule whereas all the orthotopic grafts had an abundant blood supply and the ovarianbursa did not become fibrotic.

Several workers have concluded that a reduction or delay in vascularity will prolongthe survival time of a graft (Medawar, I948; Billingham and Boswell, I953; Merwin andHill, I954). The marked difference in the host reaction to subcutaneous and to orthotopicovarian grafts may well have been due to difference in vascularity in the two positions. Tothis extent grafts implanted subcutaneously in these stocks of rats were in an immuno-logically privileged site.

ACKNOWLEDGMENTSI should like to thank Dr. A. S. Parkes for the interest he has taken in this work, Dr. Janet S. F.

Niven and Dr. J. H. Humphrey for help in the preparation of the manuscript. Dr. Niven alsoexamined the histological material.

Miss A. Wakeford and Mr. F. Crisp gave invaluable technical assistance.

REFERENCES

BILLINGHAM, R. E. and BOSWELL, T. (I 953). 'Studieson the problem of corneal homografts.' Proc. Roy. Soc.B., 141, 392-406.

BILLINGHAM, R. E. and PARKES, A. S. (I955). 'Studieson the survival of homografts of skin and ovariantissue in rats.' Proc. Roy. Soc. B., 143, 55o-6o.

DEANESLY, R. (I956). 'Cyclic function in ovariangrafts.' J. Endocrin. (1956), I3, 2 I1-20.

FRAZER, S. F. D. (I955). 'Foetal death in the rat.' 7.Embryol. exp. Morph., 3, 13-19.

GREEN, S. H., SMITH, A. U. and ZUCKERMAN, S. (I 956).'The numbers of oocytes in ovarian autografts afterfreezing and thawing.' J. Endocrin., 13, 330-4.

HARRIS, M. and EAKIN, R. M. (I 949). 'Survival oftransplanted ovaries in rats.' J. exp. Zool., I I2,I31-63.

INGRAM, D. L. (I 958). 'Fertility and oocyte numbersafter X-irradiation of the ovary.' Y. Endocrin., 17,81 -90.

KROHN, P. L. (1955). 'Ovarian homotransplantation.'Ann. N.r. Acad. Sci., 59, 443-7.

KROHN, P. L. (I959). 'Ovarian transplantation inmice.' In Biological Problems of Grafting. Ed. F. ALBERTand P. B. MEDAWAR: Blackwell Scientific Publica-tions, Oxford.

MANDL, A. M. and ZUCKERMAN, S. (195I). 'The rela-tion of age to numbers of oocytes.' J. Endocrin., 7,I 90-3.

MANDL, A. M. and ZUCKERMAN, S. (I 956). 'Thereactivity of the X-irradiated ovary of the rat.' J.Endocrin., 13, 243-6 I.

MCQUARRIE, D. G., KIM, J. H. and VARCO, R. L. (I 959).'Long-term survival of intra-strain homografts incommercially bred rats.' Trans. Bull., 6, 97-100.

MEDAWAR, P. B. (I948). 'Immunity to homologousgrafted skin. III. The fate of skin homografts trans-planted to the brain, to subcutaneous tissue, and tothe anterior chamber of the eye.' Brit. J. exp. Path.,Path., 29, 58-69.

MERVIN, R. M. and HILL, E. L. (I 954) . 'Fate ofvascularized and non-vascularized subcutaneoushomografts in mice.' J. nat. Cancer Inst., 14, 8I9-39.

PARKES, A. S. (1956). 'Survival time of ovarian homo-grafts in two strains of rats.' J. Endocrin., 13, 20I-10.

PARROrr, D. M. V. (I958). 'Orthotopic ovarian graftsin mice, rats and hamsters.' J. Endocrin., A6, xi-xii.

PARROTrr, D. M. V. (959). 'Orthotopic ovarian graftsin the Golden Hamster.'J. Endocrin., I9, 126-38.

PARROTT, D. M. V. and PARKES (I956). 'Orthotopicovarian grafting after sterilization by X-rays.' Brit.vet. J., 112, 550-4.

PARROTrT, D. M. V. and PARKES (I960). 'Dynamics ofthe orthotopic ovarian graft in "Sex Differentiationand Development".' Ed. C. R. AUSTIN. Mem. Soc.Endocrin., No. 7, 7'. Cambridge University Press.