Elizabeth Whitehouse

A morphological study of radiation nephropathy in the pig.

Both kidneys of mature pigs received a single dose of 9.8 Gy 60Co gamma rays. Pigs were killed between 2 and 24 weeks after irradiation and the kidneys examined histologically. Glomerular and tubular changes were observed within 2 weeks of irradiation. Neutrophils and other leukocytes were seen within glomerular capillary loops; mesangial matrix and cell number increased. A progressive increase in thickening of the basement membrane and a decrease in capillary lumina were then noted. Basement membrane duplication occurred within 12 weeks. By 24 weeks these lesions had increased in severity, sclerotic endstage glomeruli, predominantly subcapsular or juxtamedullary, being evident. Tubular lesions initially consisted of focal areas of tubular atrophy in the juxtamedullary region. By 6 weeks subcapsular foci of tubular degeneration, regeneration, and necrosis were found; these appeared to resolve 12 weeks after irradiation. At later times the severity of the tubular lesions varied between pigs, with some exhibiting interstitial fibrosis involving a complete band of subcapsular tissue, while others showed relatively mild changes. There was no apparent change in the vasculature. These findings indicate that (a) there is no one target or dose-limiting cell, and (b) the vasculature does not play a primary role in the development of radiation nephropathy.

Boron neutron capture therapy: A guide to the understanding of the pathogenesis of late radiation damage to the rat spinal cord

PURPOSE Before the commencement of new boron neutron capture therapy (BNCT) clinical trials in Europe and North America, detailed information on normal tissue tolerance is required. In this study, the pathologic effects of BNCT on the central nervous system (CNS) have been investigated using a rat spinal cord model. METHODS AND MATERIALS The neutron capture agent used was 10B enriched sodium mercaptoundecahydro-closododecaborate (BSH), at a dosage of 100 mg/kg body weight. Rats were irradiated on the thermal beam at the Brookhaven Medical Research Reactor. The large spine of vertebra T2 was used as the lower marker of the irradiation field. Rats were irradiated with thermal neutrons alone to a maximum physical absorbed dose of 11.4 Gy, or with thermal neutrons in combination with BSH, to maximum absorbed physical doses of 5.7 Gy to the CNS parenchyma and 33.7 Gy to the blood in the vasculature of the spinal cord. An additional group of rats was irradiated with 250 kVp X rays to a single dose of 35 Gy. Spinal cord pathology was examined between 5 and 12 months after irradiation. RESULTS The physical dose of radiation delivered to the CNS parenchyma, using thermal neutron irradiation in the presence of BSH, was a factor of two to three lower than that delivered to the vascular endothelium, and could not account for the level of damage observed in the parenchyma. CONCLUSION The histopathological observations of the present study support the hypothesis that the blood vessels, and the endothelial cells in particular, are the critical target population responsible for the lesions seen in the spinal cord after BNCT type irradiation and by inference, after more conventional irradiation modalities such as photons or fast neutrons.

The radiation response of the cervical spinal cord of the pig: Effects of changing the irradiated volume

PURPOSE An investigation of the field size effect for the cervical spinal cord of the pig after single doses of gamma-rays. In this study, clinically relevant volumes of the spinal cord were irradiated. METHODS AND MATERIALS The effects of the local irradiation of different lengths of the spinal cord (2.5 cm, 5.0 cm, and 10.0 cm) have been evaluated in mature pigs (37-43 weeks). Single doses of 25-31 Gy were given using a 60Co gamma-source, at a dose rate of 0.21-0.30 Gy/min. The incidence of radiation-induced paralysis was used as the endpoint. The data were analyzed using probit analysis and a normal tissue complication probability (NTCP)-model. RESULTS Twenty-five animals out of a total of 53 developed paralysis, with histological evidence of parenchymal and vascular changes in their white matter. The slope of the dose-response curves decreased with the decrease in field size; however, there was no significant difference at the radiation dose associated with a 50% incidence of paralysis (ED50) irrespective of the method of analysis. The ED50 values +/- standard errors (+/- SE) were 27.02 +/- 0.36 Gy, 27.68 +/- 0.57 Gy, and 28.28 +/- 0.78 Gy for field lengths of 10, 5, and 2.5 cm, respectively. Analysis of the data with a normal tissue complication probability (NCTP) model gave similar results. The latent period for paralysis was 7.5-16.5 weeks with no significant differences between dose and field size. CONCLUSION No significant field size-related differences in response were detectable in the cervical spinal cord of mature pigs after single dose irradiations, specifically at a clinically relevant level of effect (< ED10).

Sequential evaluation of radiation-induced glomerular ultrastructural changes in the pig kidney

Both kidneys of 12 mature female pigs received either a single dose of 9.8 Gy 60Co gamma rays or sham irradiation. At intervals of 1-4 weeks serial renal biopsies were obtained, followed by sacrifice at 24 weeks after irradiation. Individual kidney glomerular filtration rate (GFR), effective renal plasma flow (ERPF), and the hematocrit (Hct) were measured routinely. Renal irradiation resulted in a progressive decline in GFR, ERPF, and Hct, with minimal values being observed within 12 weeks of irradiation. No change in any of these parameters was noted in the sham-irradiated pigs. The initial morphological change in irradiated glomeruli was leukocyte attachment to capillary endothelial cells 3-6 weeks after irradiation followed by activation and swelling of the endothelial cells. This was followed by pronounced increases in capillary permeability with fluid and erythrocyte, leukocyte, and platelet exudation into the subendothelial/mesangial space. This resulted in compression of glomerular capillary lumina, which occurred concomitantly with the reduction in GFR. By 12 to 15 weeks after irradiation the changes in endothelial cells were less evident. However, mesangial cells exhibited evidence of activation and proliferation accompanied by progressive mesangial expansion and sclerosis. Thus the glomerular capillary endothelial and mesangial cells appear particularly important in the pathogenesis of radiation nephropathy.

Repair, repopulation and cell cycle redistribution in rat foot skin

The influence of the phenomena of the repair of sublethal damage, repopulation and the role of the reassortment of surviving clonogenic target cells within the cell cycle have been examined in the foot skin of rats using a series of split dose experiments. The dose-related incidence of moist desquamation was used as an end-point. Initially the iso-effect dose for moist desquamation (ED50) increased with an increasing time interval (1-22 h) between two equal fractions. This effect was attributed to the well established phenomenon of the repair of sublethal damage. This appeared to be maximal with a 22 h gap between fractions. A further increase in the time interval, from 2-7 days, between two equal fractions resulted in a decrease in the ED50 value for moist desquamation. The phenomenon is most likely to be explained by a shortening of the cell cycle time in surviving epithelial target cells as repopulation first initiated. With intervals between two fractions of greater than 10 days the ED50 for moist desquamation again increased. This is likely to represent an increase in the number of epidermal target cells (repopulation). Further evidence for the effect of a reassortment of cells in the cell cycle has come from another study in which a half-tolerance priming dose of 16.8 Gy was followed by three daily fractions starting 48 or 125 h after the priming dose. The ED50 for moist desquamation based on the total fractionated dose (three fractions) was significantly lower (P < 0.05) after the longer time interval, i.e. fractions given on days 5, 6 and 7 after the primary dose. These findings were supported by the results of a cell proliferation kinetic study and jointly question the validity of a frequently made assumption of equal biological effect per fraction in a prolonged fractionated irradiation schedule.

A NEW MODEL OF RADIATION-INDUCED MYELOPATHY: A COMPARISON OF THE RESPONSE OF MATURE AND IMMATURE PIGS

PURPOSE The development of an experimental model of radiation-induced myelopathy in the pig which would facilitate the study of the effects of clinically relevant treatment volumes. METHODS AND MATERIALS The effects of local spinal cord irradiation, to a standard 10 x 5 cm field, have been evaluated in mature (37-42.5 weeks) and immature (15.5-23 weeks) pigs. Irradiation was with single doses of 60Co gamma-rays at a dose-rate of 0.21-0.65 Gy/min. The incidence of paralysis was used as an endpoint. RESULTS Irradiation of mature animals resulted in the development of frank paralysis with animals showing combined parenchymal and vascular pathologic changes in their white matter. These lesions, in common with those seen in patients, had a clear evidence of an inflammatory component. The latency for paralysis was short, 7.5-16.5 weeks, but within the wide range reported for patients. However, it was shorter than that reported in other large animal models. The ED50 value (+/- SE) for paralysis was 27.02 +/- 0.36 Gy, similar to that in rats taking into account dose-rate factors. The irradiation of immature pigs only resulted in transient neurological changes after doses comparable to those used in the mature animals, ED50 value (+/- SE) 26.09 +/- 0.37 Gy. The reasons for these transient neurological symptoms are uncertain. CONCLUSION A reliable experimental model of radiation-induced myelopathy has been developed for mature pigs. This model is suitable for the study of clinically relevant volume effects.

Stenosis of the tubular neck : A possible mechanism for progressive renal failure

Fibrosis in chronic renal failure is closely associated with declining function. Its role in affecting function is less well defined. The radiation model of chronic renal failure was used to examine the tissue distribution of fibrosis and scarring and its role in influencing the loss of function in chronic renal disease. A striking and progressive pattern of fibrosis and narrowing of the glomerulotubular neck was found in irradiated pig kidneys. These narrowed necks increased in prevalence with time after irradiation. At 20 weeks after irradiation, the average neck diameter reduction was 60%, as compared with nonirradiated controls, a percentage that is consistent with a reduction in flow and pressure at this critical point of the nephron. Glomerulotubular neck narrowing may thus directly reduce the glomerular filtration rate of an individual nephron. Fibrotic neck stenoses may be a factor in progressive chronic renal failure.

Age-related changes in the cell kinetics of rat foot epidermis.

Abstract. The durations of the cell cycle and its component phases have been determined for the basal layer of the epidermis of the skin from the upper surface of the hind foot of the rat using single pulse [3H]‐thymidine labelling and the percent labelled mitosis (PLM) technique. Rats of three age groups were used, namely 7, 14 and 52 weeks. The duration of DNA synthesis (Ts) and the G2 plus M phase (Tg2± m) were comparable in 7‐week and 52‐week‐old rats (P > 0–1). The major difference between 7‐week and 52‐week‐old rats was in the duration of the G1 phase (Tg1). In 7‐week‐old rats Tg1 was 15.0 ± 0.8 h and in 52‐week‐old rats Tg1 was 31.2 ± 3.5 h. A consequence of this variation was that the overall duration of the cell cycle was longer in 52‐week‐old rats (53.9 ± 5.3 h) than in 7‐week‐old rats (30.1 ± 1.3 h).

Long-term studies of cisplatin-induced reductions in porcine renal functional reserve

SummaryMature Large White female pigs aged approx. 10 months received single intravenous doses of 1.5, 2, or 2.5 mg/kg cisplatin. The glomerular filtration rate (GFR) and the effective renal plasma flow (ERPF) in individual kidneys were measured prior to and at 4-week intervals for up to 24 weeks after cisplatin administration by renography using [99mTc]-diethylenetriamminepentaacetic acid (DTPA) and iodohippurate sodium I 131, respectively. The left kidney of each cisplatin-treated pig and that of three age-matched control pigs was then removed, and GFR and ERPF values were measured in the remaining kidney at 4-week intervals for a further 24 weeks after unilateral nephrectomy (UN). Pigs treated with cisplatin showed no significant reduction in GFR or ERPF for up to 24 weeks after drug infusion. As measured using inductively coupled plasma mass spectrometry, the mean renal platinum concentration in the left kidney removed at UN was 77.5±9.1 ng/g kidney per mg/kg cisplatin. Histological evaluation of these kidneys revealed narrow interconnecting rays of interstitial fibrosis in the deep cortex and medulla; in these areas, glomeruli exhibited thickened Bowmans capsules and occasionally shrunken sclerotic capillaries. In cisplatin-treated pigs, UN was associated with a marked reduction in the ability of the remaining kidney to increase its function in terms of GFR and, to a lesser exten, of ERPF. The increase seen in GFR following UN in the cisplatin-treated pigs was only ca. 50%–70% of that seen in age-matched UN controls. Histologically, these kidneys revealed resolution of the peritubular fibrosis observed at UN; occasional sclerotic glomeruli were also evident. Platinum remained detectable in these kidneys, the mean levels being 18.8±4.9 ng/g kidney per mg/kg cisplatin. These findings confirm previous observations and illustrate the need for caution in considering further treatment of patients who have previously received cisplatin along with a second potentially nephrotoxic agent.

Radiation Response of Mouse Lip Mucosal Epithelium: A Cell Kinetic Study

Changes in the cell proliferation kinetics of the epithelium of mouse lip mucosa have been assessed after local irradiation with a single dose of 18 Gy of 60Co gamma-rays. By the fifth day after irradiation, two distinct sub-populations of epithelial cells could be discerned. The larger of the two populations consisted of cells exhibiting varying degrees of radiation-induced damage, and the smaller population was composed of cells of normal size and appearance, intermingled between the radiation-damaged cells. There was a progressive decline in the epithelial cell density with time after irradiation, and focal denudation was seen after 11 days. Cell colonies were evident in the remnants of the epithelium by day 7. Degenerate cells could be identified in the basal layer of the mucosal epithelium, both before and after irradiation. The proportion of degenerate cells was increased 5 days after irradiation with the maximum number, approximately 3.6% being counted on days 7 and 8. In the first 2 days after irradiation, there was a reduction in the labelling index (LI) of basal cells in the epithelium. This was followed by recovery to control values on day 3. The LIs of both the radiation-damaged cells and those with a normal appearance remained relatively constant between days 5 and 11, at approximately 10 and approximately 60%, respectively. The LI of basal cells in the cell colonies was very high (approximately 70%). The estimated turnover time (TT) for the basal cell population with a normal appearance and for those in cell colonies (groups of normal cells with a cord length > or = 10 cells), was extremely short < 1 day. There was some fluctuation in TT values for basal cells exhibiting radiation-induced damage, with the shortest value (approximately 3 days) at 7 and 8 days after irradiation. It was concluded from the cell kinetic data that repopulation of the lip mucosal epithelium started between 3 and 5 days after irradiation and the overall response of the mucosa to irradiation was consistent with that predicted by a hierarchical model of cell proliferative organization.