SOME OBSERVATIONS ON FREE FAT GRAFTS: WITH REFERENCE TO THEIR USE IN MAMMAPLASTY

By JOHN ~rATSON, M.A., M.B., F.R.C.S.E.

Queen Victoria Hospital, East Grinstead

ALTHOUGH it is now seventy-five years since the first human free fat transplant was repor tedby Neuber .in 1893, before being brought into extensive use by Lexer (I925) , the procedure has never been particularly favoured by plastic surgeons on account of the unpredictable degree of shrinkage which is liable to occur in the months following grafting. Peer addressed a questionnaire to an international sample of plastic surgeons in 1956 and found that of 187 surgeons, seventy-three no longer made use of these grafts, and that all who did preferred to use fat with a portion of dermis attached.

In 1938 Gurney produced evidence of fat cell survival in experimental grafts in rats, contrary to the widely held belief that the original fat cells of the graft were absorbed, the scaffolding of the graft being repopulated by fat-carrying cells of histiocyte type (Neuhof, 1923 ; Davis and Traut, 1925). Further evidence that a proportion of fat cells survive was given by Peer (195o), reporting a series of experimental fat grafts in humans. His findings indicated that autogenous fat grafts survive transplantation as fatty tissue, but with an ultimate weight and volume loss amounting to some 45 per cent. in the case of large grafts (up to 26o g.), and 79 per cent. if implanted in the form of small multiple segments. Histological evidence suggested that this loss of weight was due to death of cells which failed to achieve an early and adequate circulation, and Peer believed that this circulation was established within four days by direct anastomosis of the ingrowing vessels of the host tissue with the original vascular system of the graft rather than by an invasion of the graft substance by host capillaries. By the twelfth day the grafts had become invested in a loose connective tissue capsule rich in blood-vessels. Grafts removed three months after transplantation showed cystic spaces representing cellular loss and containing scavenger histiocytes.

The use of free fat grafts containing a portion of attached dermis, denuded of superficial epidermis, appears to have originated as an extension of the buried curls graft procedure ; the cutis graft was defined by Loewe (1913) as " an excised area of skin, usually with an underlying layer of fat, from which the epidermal layer has been removed." Although there is a general feeling that the inclusion of dermis leads to improved results in free fat grafting, there is little information available in the literature to show whether this is in fact so, or of the reasons therefor. Peer (1956), writing of experiences with both fat and dermo-fat grafts, favoured the inclusion of dermis ; he found that large transplants to the cheek of lean patients lost so little bulk that secondary operations were necessary to remove excess tissue, but did not specify whether these particular grafts contained dermis. At first sight it might appear that the inclusion of skin with a higher metabolic activity than subcutaneous tissue might jeopardise the survival of the fat by its own oxygen requirements. Some have felt that the function of the dermis is merely to provide support and some degree of solidity in an otherwise rather sloppy

263

264 B R I T I S H JOURNAL OF P L A S T I C SURGERY

and unwieldy graft, and old appendicectomy scars have been transferred with the underlying scarred fat with this in view.

In our hands the inclusion of dermis has unquestionably improved the results of fat-grafting to an extent which renders transplantation a reliable procedure. The use of large dermo-fat grafts in the correction of hypomastia has the experimental advantage that being inserted in pairs, any inconsistencies between the two sides resulting from some failure of operative technique became evident. With a technique modified by the writer from that of Bames (I953) and described later, no difficulty has been experienced in transplanting gluteal dermo-fat grafts of considerable volume to the retromammary region in repeated instances without post-operative complications. The grafts consist of elliptical portions of dermis, 8 to 9 in. in length, 3 in. in maximum width, and containing the full thickness of gluteal fat, together with the underlying deep fascia. The weight of each graft is of the order of 2oo to 25o g. The complications of tissue necrosis, fat liquefaction, and sepsis have not been encountered with the technique used, and the clinical fate of these grafts has been followed up for a sufficient length of time to show the undoubted survival of the greater part of this mass of tissue ; if loss of bulk should occur in ensuing years it will be due to causes other than tissue necrosis from lack of an adequate initial blood supply. The purpose of this paper, however, is to discuss the means by which these large masses of tissue survive transplantation rather than to advocate, at this stage, this method because, as is well known, reduction in the bulk of these grafts may occur over a period of several years after transplantation (Conway and Smkh, 1958). The reason for this remains obscure, but may be associated with the little understood factors which regulate the distribution of fat over the body surface. Nevertheless, it would seem that if a sufficiently large bulk of tissue is transplanted, by a careful technique that allows initial survival of the main mass of cells, the ultimate resuks will be acceptable in a high proportion of cases. This would be a physiological operation free from the drawbacks and potential hazards associated with procedures involving the introduction o f " inert" materials.

I f these massive pieces of tissue were transferred on a vascular pedicle, and if this provision for their circulation proved inadequate, the inevitable result would be an extensive tissue slough, with fat necrosis, gangrene, abscess formation, and ultimately discharge of the necrotic material through the wound. It is a curious paradox that a dermo-fat graft of large size will survive without any direct provision being made for its initial circulation.

There are three ways in which a large tissue mass might survive free transplantation: the capacity of the cells to live in a state of anaerobiosis, the diffusion of Oz and CO2 through the tissue from the surrounding bed, and the rapid acquisition of a circulation of some kind. So far as the first is concerned, the capacity of mammalian cells to live anaerobically on their reserves at body temperature is usually reckoned as a matter of hours rather than days. Oxygenation by gas diffusion from the surrounding bed would depend on the rate of yield from the surrounding capillaries, and the gas diffusion rate through the tissue. In this instance an adequate 02 tension would have to be maintained in the centre of a mass whose minimum dimension is about 7 cm., a distance bearing no relation to the normal capillary density. Braithwaite et al. (1954) calculated that the endothelial surface available for diffusion in the fully dilated capillary bed of the dermis was about 184 sq. cm. per ml. of tissue. I f a capillary

SOME OBSERVATIONS ON FREE FAT GRAFTS 265

bed of such a density is necessary for normal oxygen requirements, it seems inconceivable that an adequate O~ tension could be maintained in the centre of these large masses by diffusion, even at the lowest estimate of oxygen. Oxygen diffuses with some difficulty even through the pulmonary epithelium. On a basis of gas diffusion alone, only an external layer of tissue about a millimetre thick would be expected to survive.

There remains for consideration the possibility that the grafts pick up a circulation from surrounding tissues at a very early phase. The thesis presented in this paper is that the dermis plays an active and essential part in the rapid vascularisation of these grafts.

The capacity of the denuded dermis, applied in the " reverse" position, t o survive and pick up a blood supply from the host has been amply shown by Hynes (I954 a) in his " skin dermis " graft procedure; he has stated that the grafted dermis " takes " more readily in the reversed position than the standard Wolfe graft, and that it is so vascular when it has " t a k e n " on granulation tissue that it will then support a split-skin graft on its exterior.

The problem of the establishment of an early circulation in skin grafts has received considerable attention from investigators using various types of grafts in animals and man, with a good deal of discussion as to whether revascularisafion is achieved by direct anastomosis of the host-graft vessels with persistence of the original graft vascular tree, by invasion of new capillaries from the sprouting blood-vessels of the host, or by both these methods. Bert (I865) , denuding a rat's tail of skin and introducing it subcutaneously as a graft, first noted connections between the host and graft vessels at the fourth or fifth day. The same was observed by Thiersch (I874), who used the term "inosculation " for this anastomosis of which he claimed proof as early as eighteen hours after grafting, from injection studies on an amputated limb, upon the granulations of which grafts had been applied at varying periods before severance. Garrd (I888), on the other hand, believed that revascularisation was carried out by host capillary buds, after most of the graft vessels had become obliterated, but carried out no injection studies. Davis and Traut (I925) were able by injection to demonstrate the presence of anastomotic channels as early as twenty-two hours after transplanting canine Wolfe grafts, although they stressed that the definitive revascularisation of the graft was by subsequent capillary invasion. Medawar (I944), in his classical paper on the homograft problem, found that full-thickness grafts in rabbits had sustained a through-and-through invasion by blood-vessels which was completed by the fourth day, and commented on the differentiation of the vessels in the lower reaches of the dermis into thin-walled arterioles and venules up to 0.03 ram. in lumen diameter at this period. Conway et al. (I95I), using a transparent chamber techniquein mice, found that invasion by new capillary buds did not take effect for four to five days, with budding from the host vessels which exhibited a strong directional tendency with parallelism towards the graft. The same workers (I957), using an improved technique, have now observed the persistence of the graft circulatory tree with direct anastomosis to the host vessels ; no particular activity was seen on the first day, but on the second day the graft vessels became distended and a slight back and forth movement of blood cells was visible; by the third day unidirectional flow was seen. On the whole the numerous observations support the belief in the persistence of the graft circulatory tree, albeit temporarily, as suggested by Peer (i956), and by Converse and

266 B R I T I S H JOURNAL OF P L A S T I C SURGERY

Rapaport (I956), who found that an early connection between the graft and the host vessels preceded invasion of the graft by host vessels.

This would certainly seem the likely manner in which large masses of dermo-lipomatous material can survive as a free graft, since it is hardly conceivable that new proliferating capillaries from the host could invade several inches of tissue in time to ensure survival. The crux of the problem is whether even by direct vascular anastomosis a circulation can be established with sufficient speed to prevent irreversible anoxic changes. Hynes (I954 b) noted the dilatation of the dermal vessels twenty hours after the application of a human split-skin graft to human granulation tissue, with the appearance of red blood cells in the vessels within twenty-four hours of transplantation, but was unable at this phase to demonstrate actual capillary continuity between the graft and its bed. He postulated the existence of a primitive circulation of fibrinogen-free fluid between the surface of the granulations and the vessels of the graft. Certainly our own histological observations on human dermal and skin grafts, which show consider- able oedema and widely dilated dermal vessels during the first few days, suggest the fluid is abstracted from the surrounding tissues into the graft. Nevertheless it seems doubtful that a circulation so primitive in character could support any sizeable mass of tissue. Unfortunately at the early phase, well described by Medawar (I944) as that of "primary union and vascularisation, characterised by migratory and amoeboid movements of cells of all types," and when the graft is still but loosely adherent to its bed, the demonstration of an early circulation path by injection studies presents obvious technical difficulties in man. In an attempt to throw further light upon this matter a split-skin graft was applied to the surface and a dermal graft buried in the subcutaneous tissues of a finger twenty-three hours before its amputation for Dupuytren's contracture. The digital arteries of the amputated finger were then perfused (IO per cent. colloidal iodine) at a controlled pressure of I3O mm. Hg. Thick sections, however, failed to disclose any early anastomotic channels, and neither the vessels of the surface Thiersch graft nor those of the buried dermal graft showed the presence of any injected material. At this time phase the result of the experiment was thus negative, but it would obviously be worth repeating at increasing time periods after insertion of the graft when opportunity presents.

An attempt was made on quite different lines to obtain indirect evidence of vascularisation by injecting phenolsulphonphthalein (6 mg. in I ml.) into dermo-fat grafts just before and at varying time periods after insertion. When injected into normal subcutaneous tissue, as a control, it was found that 85 per cent. or more of the dye could be recovered in the urine within three hours. When injected into the graft just before insertion, diffusion was so slow that in repeated instances the dye failed to appear in the urine in detectable amounts over a period of two days after injection. Any hope, therefore, that its appearance post- operatively in the urine might provide a clue to the time of vascularisation of the graft had to be abandoned. Similarly, injection of the graft twenty-four hours after insertion failed to produce excretion in five instances with the exception of one patient who showed a minute trace in the urine (this may have been due to extravasation of a small percentage of the dose). At forty-eight hours,

' however, all cases showed a normal excretion. It is not, of course, known whether this is a true test of revascularisation, but the results are suggestive.

It is evident that the dermis is highly active and adaptable tissue, capable of

S O M E O B S E R V A T I O N S O N F R E E F A T G R A F T S 267

rapid modification of its morphology to meet new situations, particularly in respect of its vascular structure. These properties have been exploited in various procedures in plastic surgery. Braithwake (1951) has shown that a modification in the dermal circulation is a major factor in improving the blood flow ill skin flaps after delaying operations. Clau6's dtoile method of attaching a tube pedicle with apposition o f dermis to dermis is a highly effective way of achieving rapid vascularity across the junction. It would seem certain that chondro-cutaneous grafts survive by the rapid establishment of a circulation across the dermal junction (McLaughlin, 1955). There remains for consideration whether the vaso-organisation powers of the dermis are attributable merely to the natural density of its contained capillary bed or whether other more obscure factors are involved. It is evident that a dense capillary bed in transplanted tissue should favour ready anastomosis, available regenerating vessels ill the recipient bed only having to deviate through a few microns ill order to obtain vascular contact. Nevertheless the fact that muscle tissue, in spite of ks high vascularky, responds to transplantation by atrophy and fibrosis, suggests that mere capillary densky may not be the only factor concerned - - the remarkably rapid morphological changes of which the dermis is capable during wound healing, and its effect on tissues with which it is in contact, suggest that it may play the part of an active vaso-inductive agent rather than a merely passive role.

With this in mind it was thought that the superiority of dermo-lipomatous grafts over simple fat grafts might lie in the possibility that the dermal element, wheI1 placed in intimate contact with a competent tissue, might exert a stronger vaso-inductive effect than fat alone. Small dermo-lipomatous grafts were buried beneath skin flaps during delaying operations, with the dermis in contact with the deep fascia, and removed at varying intervals of four to ten days. Histological examination after removal up to ten days showed very little reaction between the host fat and the fat of the implant ; at five days (Fig. I) there still existed a plane of cleavage, occupied by a thin layer of fibrin and a few scattered red blood cells, without identifiable vascular response. The dermis, on the contrary, showed intense activity, with firm adherence to the base and dilatation of blood-vessels which contained injection material in such specimens in which successful perfusioI1 could be carried out through a host vessel. At this phase it is often necessary to place ties round the specimen before histological section, to prevent the host and donor fat from actually falling apart. Whilst the graft is ultimately nourished by vessels traversing the capsule, in the initial phases the histological picture suggests that the dermis plays the major if not the sole part.

The successful induction of all adequate blood supply by the graft dermis implies its application to a competent bed, i.e., one containing an adequate density of capillary endothelial material. The subcutaneous fat of the trunk and limbs has a low potential in this respect, and we have preferred to place the dermis ill contact with the most vascular tissue available, be this muscle, skin dermis, or mammary gland, rather than leave it lying free in the fat. The immediate reaction of the host bed after the dermis is placed in contact is hypera:mia extending over quite a wide area, and persisting for several days, due to the opening up of every available capillary in the vicinity, as a response to the presence of anoxic tissue and presumably mediated by the release of H substance. Armistead (1956) noted such an " inflammatory" reaction in the peritoneum around experimental skin grafts applied to mammalian intestine and attributed it to operative trauma.

268 BRITISH JOURNAL OF PLASTIC SURGERY

Converse and Rapaport (I956) refer to the zone of erythema and oedema surrounding human full-thickness grafts. The coloured illustrations of McLaughlin's paper (1955) show well the hypermmic zone surrounding a chondrocutaneous graft applied to the nose.

I f the theoretical conclusions reached above are correct, the larger the surface area of dermis in relation to the mass of fatty tissue the greater the chances of survival. Our mammary grafts presented a surface area of dermis of approximately

FIG. I Experimental dermo-fat graft five days after insertion in calf.

a, Normal host skin surface. b, Plane of cleavage still apparent between graft fat and host subcutaneous fat. c, Dermis of graft firmly adherent to and integrated with deep fascia covering

gastrocnemius. Vessels in the graft contain injection material.

I25 sq. cm. for a total surface area of 375 sq. cm., i . e . , a " vascular aspect ratio " of about I in 3. The dermal contact area for a given volume of graft can be doubled by the expedient of folding the graft upon itself, dermal surface outwards, and we have exploited this principle in other applications of fat-grafting.

From the technical point of view, close apposition between dermis and bed, with precautions against subsequent shearing stresses and sliding, and absolute hmmostasis are essential ; the same care has to be taken in introducing a fat graft as in the application of a skin graft to a raw surface. Drainage tubes should not be used and should not be necessary.

The introduction of skin beneath the body surface carries the danger of the formation of cysts. Three types of cysts are recognised, and arise from (I) epithelial recurvature, (a) hair follicles or glandular remnants, and (3) central necrosis with

SOME OBSERVATIONS ON FREE FAT GRAFTS 269

encystment of liquefied fat. The natural tendency of a free sheet of epithelium to curl, so that its margins unite to form spherical or tubular structures, has long been recognised (Zimches, I93I ; Glucksmann, I95I ; Reid, I952), and forms the basis of urethral reconstruction on the Duplay principle. This type of cyst does not seem to occur, provided that the major part of the epithelium has been removed from the graft, and the graft so placed on its bed and sufficiently large to preclude reunion of its margins. Cysts arising from follicular and glandular remnants are an unavoidable risk. Such cysts do not appear to attain any size or to give rise to troublesome complications : there is a general tendency towards disappearance of all hair follicles and glands as the dermis becomes modified in its new situation. No such cysts have been evident clinically in the mammary grafts in this series. Cysts containing liquefied fat are an index of cellular necrosis from inadequate vascularisation of the tissue ; the fat may be discharged after several weeks or months, or alternatively undergo absorption with consequent shrinkage and central fibrosis of the graft.

OPERATIVE TECHNIQUES IN MAMMAPLASTY

With the patient prone and the hips slightly flexed and abducted the grafts are first obtained, using the full thickness of available fat and taking the deep fascia also by severing the septa which pass between the fasciculi of the gluteus maximus muscle (Fig. 2, A)--if the grafts are taken in the line of the gluteal crease, the scar is likely to show beneath a bathing costume, and it is likely to produce tightness on full hip flexion; if taken in a direction at right angles to that recommended, the sacral area will be involved, where the fat is likely to be insufficiently thick. Closure of the defects is assisted by buttoned mattress sutures to support the weight of the tissues. Undermining is not required and drainage tubes have not been used.

A thin graft is then taken from the surface of each mass with a Humby-type knife (Fig. 2, c). It is possible to remove the surface epithelium when the grafts are still in situ before excision, but it has been found easier and satisfactory to do this after removal, and may indeed be advantageous by expelling any stagnant blood retained in the vessels of the fat.

The incisions are then dressed with gauze and strapping, and the patient rotated into a semi-sitting position--meantime the grafts are dusted with penicillin and flavazol powder.

A retromammary approach is made through a 3 in. incision in the lateral part of the submammary groove, or where this should be. Dissection is continued upwards in a plane immediately superficial to the pectoral fascia (Fig. 3). Hmmostasis must be absolute, and a moderate degree of induced hypotension shortens this procedure. The dissection must be continued upwards and inwards until a cavity is achieved which will exactly accommodate the graft. The graft is placed with the dermis superficial. It will be found that as its two extremities are approximated (Fig. 2, D), the graft assumes a conical form, at the same time presenting the maximum area of dermis for contact whilst folding the fat layer against itself. The closer the extremities are approximated, the greater the forward position. The desired position is maintained by passing two silk sutures through each extremity of the graft, passing them on straight needles through the skin of the chest wall above the reconstructed breast, and securing them by tying

3F

270 BRITISH JOURNAL OF PLASTIC SURGERY

FIG. 2 Operative technique.

A, Position of the gluteal donor sites, and B, their closure. C, Removing the superficial epithelium. D, Showing how when the graft is folding upon itself, dermis outwards, it tends to assume conical shape, and also presents the maximum amount of dermis and the minimum of fat to the surrounding bed. E, Pre-operative condition with medial extremities of incision lines shown. F, One graft is in position and the manner of fixation against shear by suspension from buttons is shown. The form of the new breast depends on the height of these sling sutures and their distance apart. G, Method of fixation with adhesive strapping. This is not disturbed for twelve days. H, Post-operative condition

at five months.

SOME OBSERVATIONS ON FREE FAT GRAFTS 27I

over buttons (Fig. 2, H). In this way not only is accurate positioning obtained, but all shearing movement is avoided whilst revascularisation is taking place, mid the grafts are held up well clear of the incision lines. Closure is in two layers and it is the writer's practice to use fine catgut for the fat and subcuticular wire for the skin.

Subsequent fixation is best obtained by strapping, with a minimum of gauze and no wool (Fig. x, G). These dressings are not disturbed for twelve days.

FIG. 3 Diagram showing position of graft and manner of supension. The graft lies superfici/d to the pectoral fascia, and beneath the underdeveloped breast tissue. It is folded with the dermis outwards, and the fat is largely included

in this dermal envelopment.

Twelve bilateral cases have been treated by this method over a period o f two years and the only post-operative complication experienced has been one gluteal hmmatoma-- the procedure has, in fact, been most trouble-free. So far as the chest is concerned, hmmatoma, sepsis, or discharge of liquid fat as has been described by some writers has not been seen. In the immediate post-operative phase the grafts swell and become firm with oedema, and this to some extent persists for about three months, the grafts during this period being a little larger than when inserted. Subsequently some degree of softening takes place, although the grafts remain a little firmer than normal breast tissue. Of the twelve patients treated, one was treated two years ago, four over one year, one over nine months,

3 F~

272 BRITISH JOURNAL OF PLASTIC SURGERY

and the remainder more recently. No significant shrinkage has yet been found in any patient, although it is possible that a shrinkage of up to some 15 per cent.

A B Fro. 4

Pre-operative condition (A) and post-operative result (B) at six months. The grafts have remained unchanged at ten months but the patient is

now pregnant.

A B Fro. 5

Pre-operative condition (A) and post-operative result (B) at fourteen months. No change since insertion.

of mass might occur without detection. Slight post-operative sagging from weight has occasionally occurred. The patients have expressed themselves as highly satisfied with the results obtained (Figs. 4 to 7).

SOME OBSERVATIONS ON FREE FAT GRAFTS 273

A B FIG. 6

Pre-operative condition (A) and post-operative result (B) one :year eight months after treatment. Some sagging from weight which was cor~cted at the time of the operation has recurred, but the size of the grafts has not

detectably changed.

A B FIG. 7

Pre-operative condition (A) and post-operative result (B) thirteen months after treatment.

274 BRITISH JOURNAL OF PLASTIC SURGERY

SUMMARY AND CONCLUSIONS

I. Problems relating to the survival of massive free dermo-lipomatous transplants are discussed.

2. Reasons are given for the supposition that the dermis plays an essential part in the rapid revascularisation of these grafts.

3. A technique for the treatment of amastia and hypomastia by dermo- lipomatous transplants is described, together with the results obtained.

I am indebted to Dr A. Sachs for numerous histological and biochemical investigations, and to Mr Gordon Clemetson for the photographs.

REFERENCES

ARMISTEAD, W. W. (1956). Plast. reconstr. Surg., 18, 9. BAMES, H. O. (1953). Plast. reconstr. Surg., IX, 404. BERT, P. (1865). C . R . Acad. Sci., 6I, 587 . BRAITHWAITE, F. (1951). Brit. J. plast. Surg., 4, 28. BRAITHWAITE, F., FARMER, F. T., BELL, R. C., and EDWARDS, J. R. G. (1954). Brit. ft.

plast. Surg., 7, I3. CONVERSE, J. M., and RAPAPORT, F. T. (x956). Ann. Surg., 143 , 306. CONWAY, H., GRIFFITH, B. H., SHANNON, J. E., and FINDLAY, A. (1957). Plast. reconstr.

Surg., 2o, 10 3. CONWAY~ H., and SMITH~ J. (1958). Plast. reconstr. Surg., 21, 8. CONWAY, H., STARK, R. B., and JOSLIN, D. (I95I). Plast. reconstr. Surg., 8, 312. DAVIS, J. S., and TRAUT, H. S. (1925). Ann. Surg., x, 35. GARRY, C. (1888-89). Beitr. klin. ChiT., 17, 318. GLUCKSMANN, A. (I95I). Brit. ft. plast. Surg., 4, 88. GURNEY, C. E. (I938). Surgery, 3, 68o. I-IYNES, W. (I954 a). Brit. ft. plast. Surg., 6, 257 - - (1954 b). Brit. ft. plast. Surg., 7, 97. LEXER, E. (1925). Arch. klin. ChiT., 138, 294. LOEWE, O. (I913). Mi~nch. med. Wschr., 6o, I32o. McLAUGHLIN, C. R. (I955). Brit. J. plast. Surg., 7, 274. MEDAWAR, P. B. (I944). ft. Anat., 78, I76. NEUBER (I893). ChiT. Kongr. Verhandl. dtsch. Ges. ChiT., 22, 66. (Quoted by L. Peer (195o).) NEUHOF, H. (I923). " T h e Transplantation of Tissues," p. 70. New York : Appleton & Co. PEER, L. (195o). Plast. reconstr. Surg., 5,217.

(I956). Plast. reconstr. Surg., r8, 244. REID, D. A. C. (I952). Brit. J. plast. Surg., 4, 235. THmRSCH, K. (1874). Arch. klin. ChiT., I7, 318. ZIMCHES (193I). Frankfurt. Z. Path., 42, 2o3.