By D. Nuss, June Duursma,
D. J. Pudifm, D. Pirie, and R. E. Mickel
l Seven infants and children with renal tumors underwent nephrectomy and the tumors were cultured to investigate the behavior of their cells. In four children over 1 year of age with histologically typical nephroblastoma the tumor cells were bizarre and disorderly. In one patient 3 months of age where the histology suggested malignancy the tumor culture showed an orderly growth pattern of spindle cells akin to normal kidney and the patient has had no recurrence for 18 months. In one infant seven months of age the histology was consistent with malignancy and this was confirmed on
culture showing bizarre cells similar to those of nephroblartoma. The third infant under 1 year of age had a teratoma on histology and an unusual growth pattern on culture characterised by polyhedral cells with long processes and much overlapping. It is suggested that in infants with kidney tumors, tissue culture be used to help in assessing the malignant potential of the tumor and deciding on adjuvant therapy.
INDEX WORDS: Renal tumors; nephroblastoma; teratoma; tumor culture.
CULTURE HAS several possible applications in the assessment of tumors. Firstly, it may be useful in establishing the tissue of origin in very anaplastic tumors, as was shown by Kadin and Bensch’ when they reported that Ewing’s sarcoma was of myelogenous origin with a manner of growth and spread analogous to plasma cell myeloma. The NeuroblastomaEwing’s sarcoma controversy was thus put to rest. Secondly, it may be used to indicate whether certain cytotoxic drugs inhibit growth in vitro. Thirdly, it may expose the malignant propensity of a tumor by revealing its manner of growth during culture. Cells from a benign tumor show an orderly growth pattern closely resembling the parent tissue, while in a malignant tumor the growth pattern may be bizarre and uncontrolled. It was with this last concept in mind that we decided to study mesoblastic nephromas and compare their cultural characteristics with those of normal kidney and nephroblastoma tissue. MATERIALS Seven kidney tumors
Three of these were derived from infants under over I yr of age, (18 mo, 24 mo, 3 and 4 yr). See Table I. Normal kidney tissue was obtained from parts of the kidney not involved with tumor. Kidneys from cadaver renal transplant donors without renal disease were also cultured for comparison. The culture procedure adopted was as follows: Small explants of tissue were placed in a 30 ml
I yr of age (3 mo, 7 mo and 7 mo) and four were from children
Falcon Tissue Culture flask, dried for I hr at 37°C. and covered with T.C. 199 supplemented with 20”); fetal calf serum. The air in the flasks was replaced with 5% carbon dioxide in air. Cell monolayers were observed from l-3 wks after the setting up of the culture. The pathology report was not conveyed to the department that set up the tissue culture. cultures were photographed.
From rhe Departments Institure of Immunology,
Surgery and Pathology,
and the Natal
Durban, So. Africa.
Address Jbr reprint requests: D. Nuss, FRCS/CI. Norfolk,
Ave., Suite 200,
@ 1977 bv Grune & Stratton,
Journal of Pediatric Surgery, Vol. 12, No. 4 (August), 1977
Table 1. Summary
of a spindle numerous
cells of em-
but a benign
in the undif-
cells of em-
size and shape assumed
bers of mitoses. 7mo
and much overlapping. 4 Y’
0s well (IS
vc~scular and copsulor
sion. 3 Y’
Nephroblostoma and capsular
As in (4) above.
(4) above. 6
As in (4) above
OS in (4)
As in (4) above.
Normal kidney tissue in culture produces a monolayer in which whorls of ovoid cells are seen (Fig. 1). The tumor from patient No. 1 (3 mo old infant) showed a similar picture to that seen with normal kidney cultures. There was an orderly growth pattern of elongated cells similar to normal kidney (Fig. 2). The histology on the other hand was highly suggestive of a malignant tumor. It showed a spindle cell stroma with numerous clusters of undifferentiated cells of embryonic character with tubules of various sizes. The spindle cell stroma con-
Normal kidney culture.
Tumor culture from patient No. 1 age 3 mo, showing orderly growth pattern.
Fig. 3. Low power microscopicslide from tumor of patient No. 1 aged 3 mo, showing clusters of undifferentiated cells in a spindle cell stroma.
Fig. 4. growth.
Tumor culture from patient No. 2 ogod 7 mo, showing bizarre cells with disorderly
Fig. 5. Low power microscopicslide from tumor of patient No. 2 aged 7 mo, showing tubules, mitoses, und necrosis.
Fig. 6. Tumor culture from pcltient No. 3 aged 7 mo, showing polykdml processes.
tolls with long toll
Fig. 7. choroid
No. 3 aged
No. 5 aged
3 yr, showing
Fig. 9. Low power microscopic slide from patient and necrosis.
No. 5 aged 3 yr, showing
tained areas of myxomatous degeneration but had a benign appearance. In the undifferentiated cell clusters there was individual cell necrosis and moderate numbers of mitoses. There was no vascular or capsular invasion (Fig. 3). The culture from patient No. 2 (7 mo old infant) was derived from a tumor which was situated in one pole of a horseshoe kidney. The cells seen here were bizarre in size and shape and assumed no orderly pattern (Fig. 4). The histological examination of this tumor revealed the appearance of a typical nephroblastoma. Undifferentiated cells predominated and were separated by a spindle cell stroma. Tubules were present and there were moderate numbers of mitoses noted both in the tubules and in the undifferentiated cell formations. Areas of necrosis were quite frequent (Fig. 5). The culture from patient No. 3 (7 mo old) was characterized by a disorderly growth of polyhedral cells with long cell processes and much overlapping (Fig. 6). The histopathologist diagnosed a benign teratoma containing recognisable brain and choroid plexus tissue (Fig. 7). The cultures from the four children over I year were all similar in that there appeared to be two types of cell. Small clumps of pleomorphic cells were scattered amongst a heavy growth of fibroblasts (Fig. 8). The histological features in all four were characteristic of malignant nephroblastoma with numerous mitoses, predominantly undifferentiated cells, large areas of necrosis and vascular invasion (Fig. 9). DISCUSSION
The success achieved in the management of childhood malignancies in the last two decades has added great impetus to cancer research. Survival figures of
807; and more have now been recorded with both nephroblastoma and rhabdomyosarcoma,‘-4 whereas before 1950 survival rates of less than 20”” were not uncommon.4-b The improvement in survival has clearly been shown to be related to the use of multiple cytotoxic drugs in combination with surgery and radiotherapy. 2-6 Although these excellent survival figures allow one to view this highly destructive disease with more optimism than previously, one must still regard the future with caution since we may well be sitting on a biological timebomb. The delayed and long term effects of these antinuclear agents on the patients as well as on their offspring is as yet unknown. However, their immediate harmful effects are well documented and may lead to the death of the patient far sooner than would be expected from his disease.7,x Bolande’s9 concept of a benign variety of embryonic kidney tumor (the congenital mesoblastic nephroma) is thus of great importance. The infants fortunate enough to have these tumors can be treated simply by surgical excision without the attendant risks of chemotherapy and radiotherapy. The difficulty arises, however, in making the diagnosis. Beckwith has summarized the features of the mesoblastic nephroma in his editorials in this journal’O~” and has drawn attention to the importance of what he calls the “field effect” or diffuse type of growth pattern, which shows itself by interdigitating bands of spindle cells with normal kidney tissue at the periphery of the tumor. However, there may be considerable “variability in the degree of cellularity and pleomorphism, and regions of rather dense cellularity and high mitotic rate are said not to worsen the outlook for cure by nephrectomy alone.” Most important is his observation that there is a spectrum of tumors varying from typically benign to outspokenly malignant spindle cell tumors with a large grey zone in the middle. It is obvious therefore that the pathologist will often be of little help in advising whether chemotherapy and/or radiotherapy should be used. Electron-microscopy has been used by several investigators’2,‘3 to try to define these tumors more clearly. In the typically benign tumor, the features are those of mature fibroblasts and some features are suggestive of muscle fibers. One reportI does mention a similarity to fibromatosis (low grade fibrosarcoma), and this is also the only documented case of a mesoblastic nephroma recurring. It is possible therefore that electron-microscopy may be of help in advising further management. The use of tissue culture techniques to determine the malignant characteristics of these tumors during growth is straightforward. The results of our study appear to justify the use of tissue culture in the management of these tumors. The first patient (3 mo old) had a histological picture which lead the pathologist (D.P.) to consider the tumor to be malignant (Fig. 3). On tissue cdture (D.J.P. and J.D.) there was no doubt that this tumor illustrated a benign behavior pattern (Fig. 2). Clinically the cultural diagnosis has been supported insofar as a nephrectomy was done when the infant was 3 mo of age and to date 18 mo later there has been no recurrence. Patient No. 2 was 7 mo of age at the time of surgery and the tumor was situated in the right half of a horseshoe kidney. In this instance, despite the patient’s age, the pathologist had no hesitation in labelling the tumor malignant (Fig. 4). The culture in this patient confirmed the malignant potential
as the cells showed a bizarre shape and size and assumed no orderly pattern of growth. Unfortunately the patient had a cardiac arrest after he had been extubated, so we have no follow-up. Under our present protocol this patient would not have received chemo- and/or radiotherapy as he was under 1 yr of age and had no obvious spread (Stage 1). From the above reports a recurrence could have been expected. In Patient No. 3 the culture showed a disorderly growth of polyhedral cells with long cell processes and much overlapping (Fig. 6). The pathologist reported this to be a benign teratoma consisting predominantly of brain and choroid plexus tissue (Fig. 7). Patients 4-7 were all typical nephroblastomas on histology with numerous mitoses, much necrosis and vascular and capsular invasion (Fig. 9). The tissue culture in this group of tumors showed a heavy fibroblastic growth with bizarre cells scattered in clumps among the fibroblasts (Fig. 8). At present, the very important decision of whether the infant should be bombarded with multiple cytotoxic drugs as well as radiotherapy is based either on the stage of the tumor or on the patient’s age. That the age delimitation is fallible is well known and that staging depends on the expertise of the individual surgeon is also well established. Tissue culture thus offers an additional item of information and helps to complete the picture. Its routine use should be encouraged. REFERENCES I. Kadin ME. Bensch KG: On the origin of Ewing’s tumor. Cancer 27:257, 1971 2. Fernbach DJ. Martin DT: Role of dactinomycin in the improved survival of children with Wilms’tumor. JAMA 195:1005, 1966 3. Koop CE, Hope JW, Abir E: Management of nephroblastoma (Wilms’ tumor) and abdominal neuroblastoma. Cancer 14:178, 1964 4. Kilman JW: Reasonable surgery for rhabdomyosarcoma. Ann Surg 178:346, 1973 5. Louw JH: The management of Wilms’ tumour. S. Afr Med J 45:106551067, 1971 6. Sutow WW. Gehan EA. Heyn RM, et al: Comparison of survival curves, 1956 versus 1962, in children with Wilms’ Tumor and neuroblastoma. Pediatrics 45:800-81 I. 1970 7. Waisman J, Cooper PH: Renal neoplasms of the newborn. J Ped Surg 54:407, 1970 8. Reuter MD, Cerilli J. Kontras SB: In-
cidence of infection in neuroblastoma patients receiving chemotherapy. J Ped Surg 4:389, 1972 9. Bolande RP, Brough AJ. Izant RJ: Congenital mesoblastic nephroma of infancy-A report of 8 cases and the relationship to Wilms’ tumor. Pediatrics 40:272, 1967 IO. Beckwith JB: Editorial: Mesenchymal renal neoplasm of infancy. J Ped Surg 5:405, 1970 1 I. Beckwith JB: Editorial: Mesenchymal renal neoplasms of infancy revisited. J Ped Surg 9:803. 1974 12. Fu YS, Kay S: Congenital mesoblastic nephroma and its recurrence. Arch Pathol 96: 66, 1973 13. Tannenbaum M: Ultrastructural pathology of human renal cell tumors, in Sommers SC (ed): Pathology Annual, New York, Appleton-Century-Crofts, 1971, pp 249-277