Transplanted senescent renal scattered tubular-like cells induce injury in the mouse kidney.

Cellular senescence, a permanent arrest of cell proliferation, is characterized by a senescence-associated secretory phenotype (SASP), which reinforces senescence and exerts noxious effects on adjacent cells. Recent studies have suggested that transplanting small numbers of senescent cells suffices to provoke tissue inflammation. We hypothesized that senescent cells can directly augment renal injury. Primary scattered tubular-like cells (STCs) acquired from pig kidneys were irradiated by 10 Gy of cesium radiation, and 3 wk later cells were characterized for levels of senescence and SASP markers. Control or senescent STCs were then prelabeled and injected (5 × 105 cells) into the aorta of C57BL/6J mice. Four weeks later, renal oxygenation was studied in vivo using 16.4-T magnetic resonance imaging and function by plasma creatinine level. Renal markers of SASP, fibrosis, and microvascular density were evaluated ex vivo. Per flow cytometry, irradiation induced senescence in 80-99% of STCs, which showed increased gene expression of senescence and SASP markers, senescence-associated ß-galactosidase staining, and cytokine levels (especially IL-6) secreted in conditioned medium. Four weeks after injection, cells were detected engrafted in the mouse kidneys with no evidence for rejection. Plasma creatinine and renal tissue hypoxia increased in senescent compared with control cells. Senescent kidneys were more fibrotic, with fewer CD31+ endothelial cells, and showed upregulation of IL-6 gene expression. Therefore, exogenously delivered senescent renal STCs directly injure healthy mouse kidneys. Additional studies are needed to determine the role of endogenous cellular senescence in the pathogenesis of kidney injury and evaluate the utility of senolytic therapy.

Urinary miRNAs as Biomarkers for Noninvasive Evaluation of Radiation-Induced Renal Tubular Injury.

Radiation nephropathy is one of the common late effects in cancer survivors who received radiotherapy as well as in victims of radiation accidents. The clinical manifestations of radiation nephropathy occur months after exposure. To date, there are no known early biomarkers to predict the future development of radiation nephropathy. This study focuses on the development of urinary biomarkers providing readout of acute responses in renal tubular epithelial cells. An amplification-free hybridization-based nCounter assay was used to detect changes in mouse urinary miRNAs after irradiation. After a single LD50 of total-body irradiation (TBI) or clinically relevant fractionated doses (2 Gy twice daily for 3 days), changes in urinary levels of microRNAs followed either an early pattern, peaking at 6-8 h postirradiation and gradually declining, or later pattern, peaking from 24 h to 7 days. Of 600 miRNAs compared, 12 urinary miRNAs showed the acute response and seven showed the late response, common to both irradiation protocols. miR-1224 and miR-21 were of particular interest, since they were the most robust acute and late responders, respectively. The early responding miR-1224 also exhibited good dose response after 2, 4, 6 and 8 Gy TBI, indicating its potential use as a biomarker for radiation exposure. In situ hybridization of irradiated mouse kidney sections and cultured mouse primary renal tubular cells confirmed the tubular origin of miR-1224. A significant upregulation in hsa-miR-1224-3p expression was also observed in human proximal renal tubular cells after irradiation. Consistent with mouse urine data, a similar expression pattern of hsa-miR-1224-3p and hsa-miR-21 were observed in urine samples collected from human leukemia patients preconditioned with TBI. This proof-of-concept study shows the potential translational utility of urinary miRNA biomarkers for radiation damage in renal tubules with possible prediction of late effects.

The impact of 177Lu-octreotide therapy on 99mTc-MAG3 clearance is not predictive for late nephropathy.

Peptide Receptor Radionuclide Therapy (PRRT) for the treatment of neuroendocrine tumors may lead to kidney deterioration. This study aimed to evaluate the suitability of 99mTc-mercaptoacetyltriglycine (99mTc--MAG3) clearance for the early detection of PRRT-induced changes on tubular extraction (TE). TE rate (TER) was measured prior to 128 PRRT cycles (7.6±0.4 GBq 177Lu-octreotate/octreotide each) in 32 patients. TER reduction during PRRT was corrected for age-related decrease and analyzed for the potential to predict loss of glomerular filtration (GF). The GF rate (GFR) as measure for renal function was derived from serum creatinine. The mean TER was 234 ± 53 ml/min/1.73 m² before PRRT (baseline) and 221 ± 45 ml/min/1.73 m² after a median follow-up of 370 days. The age-corrected decrease (mean: -3%, range: -27% to +19%) did not reach significance (p=0.09) but significantly correlated with the baseline TER (Spearman p=-0.62, p<0.001). Patients with low baseline TER showed an improved TER after PRRT, high decreases were only observed in individuals with high baseline TER. Pre-therapeutic TER data were inferior to plasma creatinine-derived GFR estimates in predicting late nephropathy. TER assessed by 99mTc-MAG3-clearance prior to and during PRRT is not suitable as early predictor of renal injury and an increased risk for late nephropathy.

Biochemical and histopathological responses of the Swiss albino mice treated with uranyl nitrate and its recovery.

Uranium is a radioactive heavy metal ubiquitous in the natural environment. In its chemical form, it is known to induce nephrotoxicity both in human and in animals. Its toxicity is dose and time dependent, also varies with form of uranium. In the present study, we assessed the nephrotoxicity induced by a single dose of uranyl nitrate (UN) in mice at different time intervals and recovery from its toxicity. Two doses of 2 and 4 mg/kg body weight of uranyl nitrate was injected intraperitoneally and animals were sacrificed after 1, 3, 5, 14, and 28 d of administration. Histopathological and biochemical alterations of post-UN dosing in comparison to control were evaluated. Tubular damage to about 75% was observed after 3 d (4 mg/kg) and the biochemical parameters such as serum creatinine, urea, and blood urea nitrogen levels were also significantly increased. Progression of tubular damage was not found after 5 d. Dose-dependent recovery of uranyl nitrate-treated animals was observed after 14 and 28 d of dosing. The concentration of uranium retained in kidney correlates with biochemical and histopathological analysis.

Extracellular nucleotides from dying cells act as molecular signals to promote wound repair in renal tubular injury.

Acute kidney injury (AKI) often correlates with poor prognosis and is followed by various severe unfavorable systemic outcomes. It is important to understand the pathophysiology of AKI for the development of novel therapeutic approaches toward promoting renal regeneration after injury. Recent studies have indicated that AKI-induced tubular cell death plays an active role in the onset of tissue regeneration; however, the mechanisms underlying renal tubular repair after injury have yet to be understood. In the present study, we explored molecules that might serve as "danger" signals in mediating tubular regeneration. Kidneys of rats systemically administered the nephrotoxicant cisplatin (to induce AKI) exhibited massive cell proliferation. The proportion of proliferating cells in the total cell distribution was highest in the outer stripe of the outer medulla coincided with where the tubular damage was the most severe in this study. This finding suggests that soluble factors may have been released from damaged cells to stimulate the proliferation of neighboring tubular epithelial cells. In elucidating the mechanism of dying cell-to-surviving cell communication using normal rat kidney NRK-52E epithelial cells, we found a significant increase in ATP levels in supernatants of these cells after the induction of cell death using ultraviolet irradiation. Furthermore, treatment of conditioned supernatants with apyrase or suramin, which inhibits purinergic signaling, resulted in significant decreases in cell proliferation and migration activities. These results demonstrate a novel role for extracellular nucleotides, probably as danger signals in aggravating tubular regeneration after AKI.

Thalidomide ameliorates inflammation and vascular injury but aggravates tubular damage in the irradiated mouse kidney.

PURPOSE: The late side effects of kidney irradiation include vascular damage and fibrosis, which are promoted by an irradiation-induced inflammatory response. We therefore treated kidney-irradiated mice with the anti-inflammatory and angiogenesis-modulating drug thalidomide in an attempt to prevent the development of late normal tissue damage and radiation nephropathy in the mouse kidney. METHODS AND MATERIALS: Kidneys of C57Bl/6 mice were irradiated with a single dose of 14 Gy. Starting from week 16 after irradiation, the mice were fed with thalidomide-containing chow (100 mg/kg body weight/day). Gene expression and kidney histology were analyzed at 40 weeks and blood samples at 10, 20, 30, and 40 weeks after irradiation. RESULTS: Thalidomide improved the vascular structure and vessel perfusion after irradiation, associated with a normalization of pericyte coverage. The drug also reduced infiltration of inflammatory cells but could not suppress the development of fibrosis. Irradiation-induced changes in hematocrit and blood urea nitrogen levels were not rescued by thalidomide. Moreover, thalidomide worsened tubular damage after irradiation and also negatively affected basal tubular function. CONCLUSIONS: Thalidomide improved the inflammatory and vascular side effects of kidney irradiation but could not reverse tubular toxicity, which probably prevented preservation of kidney function.

Gold-embedded photosensitive liposomes for drug delivery: triggering mechanism and intracellular release.

Liposomes embedded with gold nanoparticles show light-triggered contents release. We investigated the mechanism of the light-induced changes and functionality of the light-induced release in the cells. The real time small angle X-ray scattering (SAXS) analysis revealed time-dependent phase transitions in distearoylphosphatidylcholine (DSPC)/dipalmitoylphosphatidylcholine (DPPC) liposomes upon heating. Similar changes were observed when gold nanoparticle-embedded liposomes were exposed to the UV light: gold nanoparticles absorb light energy and transfer it to heat, thereby causing lipid phase transition from gel phase to rippled phase, and further to fluid phase. Without UV light exposure the gold nanoparticles did not affect the liposomal bilayer periodicity. The light-triggered release of hydrophilic fluorescent probe (calcein) from the gold nanoparticle-loaded liposomes was demonstrated with fluorescence-activated cell sorting after liposome internalization into the ARPE-19 cells. The liposome formulations did not decrease the cell viability in vitro. In conclusion, the light-triggered release from the liposomes is functional in the cells, and the release is triggered by thermal phase changes in the lipid bilayers.

Amifostine use in radiation-induced kidney damage. Preclinical evaluation with scintigraphic and histopathologic parameters.

PURPOSE: To assess the degree of protective effects of amifostine on kidney functions via semiquantitative static renal scintigraphy and histopathologic analysis. MATERIAL AND METHODS: 30 female albino rats were divided into three equal groups as control (CL), radiotherapy alone (RT), and radiotherapy + amifostine (RT+AMI). The animals in the CL and RT groups were given phosphate-buffered saline, whereas the animals in the RT+AMI group received amifostine (200 mg/kg) by intraperitoneal injection 30 min before irradiation. RT and RT+AMI groups were irradiated with a single dose of 6 Gy using a (60)Co unit at a source-skin distance of 80 cm to the whole right kidney. They were followed up for 6 months. CL, RT, and RT+AMI groups underwent static kidney scintigraphy at the beginning of the experiment and, again, on the day before sacrificing. Histopathologically, tubular atrophy and fibrosis of the kidney damage were evaluated. RESULTS: After irradiation, the median value of right kidney function was 48% (44-49%) and 50.5% (49%-52%) in RT and RT+AMI groups, respectively (p = 0.0002). Grade 1 kidney fibrosis was observed to be 60% in the RT group, while it was only 30% in the RT+AMI group. Grade 2 kidney fibrosis was 30% and 0% in the RT and RT+AMI group, respectively. Grade 1 tubular atrophy was 70% and 50% in the RT and RT+AMI group, respectively. Grade 2 tubular atrophy effect was the same in both groups (10%). CONCLUSION: Static kidney scintigraphy represents an objective and reproducible method to noninvasively investigate kidney function following irradiation. Amifostine produced a significant reduction in radiation-induced loss of renal function.

Interrelations of mitochondrial fragmentation and cell death under ischemia/reoxygenation and UV-irradiation: protective effects of SkQ1, lithium ions and insulin.

Mitochondria-targeted antioxidant 10-(6-plastoquinonyl)decyltriphenyl-phosphonium (SkQ1) as well as insulin and the inhibitor of glycogen-synthase kinase, Li(+) are shown to (i) protect renal tubular cells from an apoptotic death and (ii) diminish mitochondrial fission (the thread-grain transition) induced by ischemia/reoxygenation. However, SkQ1 and LiCl protected the mitochondrial reticulum of skin fibroblasts from ultraviolet-induced fission but were ineffective in preventing a further cell death. This means that mitochondrial fission is not essential for apoptotic cascade progression.

Ifosfamide-induced nephrotoxicity in 593 sarcoma patients: a report from the Late Effects Surveillance System.

BACKGROUND: Ifosfamide is widely used in paediatric oncology, but its use is limited by nephrotoxic side effects. The aim of this study was to evaluate the incidence and risk factors of tubulopathy, with special emphasis on the influence of age, where different findings have been published so far. PROCEDURE: Five hundred ninety three children and adolescents treated for Ewing, osteo- or soft-tissue sarcoma (median age at diagnosis: 11.7 years) were prospectively investigated for nephrotoxicity in the Late Effects Surveillance System (LESS) study. Tubulopathy was diagnosed in case of continuing hypophosphatemia and proteinuria. RESULTS: After a median follow up of 19 months, 27 patients (4.6%; 95% CI: 3.0-6.6%) had newly developed tubulopathy. This incidence was 0.4% (95% CI: 0-2.4%) in patients treated with a cumulative ifosfamide dose of < or =24 g/m2, 6.5% (95% CI: 3.6-10.7%) after 24-60 g/m2, and 8.0% (95% CI: 4.2-13.6%) after > or = 60 g/m2. In multivariate analysis, children younger than 4 years at time of diagnosis had an 8.7-fold (95% CI: 3.5-21.8) higher risk for tubulopathy than older patients. Neither carboplatin treatment nor abdominal irradiation showed any significant influence. CONCLUSION: Ifosfamide-induced nephrotoxicity was found in 4.6% of patients. Risk factors were the cumulative ifosfamide dose and young age at treatment.

Mitigation of radiation nephropathy after internal alpha-particle irradiation of kidneys.

PURPOSE: Internal irradiation of kidneys as a consequence of radioimmunotherapy, radiation accidents, or nuclear terrorism can result in radiation nephropathy. We attempted to modify pharmacologically, the functional and morphologic changes in mouse kidneys after injection with the actinium ((225)Ac) nanogenerator, an in vivo generator of alpha- and beta-particle emitting elements. METHODS AND MATERIALS: The animals were injected with 0.35 muCi of the (225)Ac nanogenerator, which delivers a dose of 27.6 Gy to the kidneys. Then, they were randomized to receive captopril (angiotensin-converting enzyme inhibitor), L-158,809 (angiotensin II receptor-1 blocker), spironolactone (aldosterone receptor antagonist), or a placebo. RESULTS: Forty weeks after the (225)Ac injection, the placebo-control mice showed a significant increase in blood urea nitrogen (BUN) (87.6 +/- 6.9 mg/dL), dilated Bowman spaces, and tubulolysis with basement membrane thickening. Captopril treatment accentuated the functional (BUN 119.0 +/- 4.0 mg/dL; p <0.01 vs. placebo controls) and histopathologic damage. In contrast, L-158,809 offered moderate protection (BUN 66.6 +/- 3.9 mg/dL; p = 0.02 vs. placebo controls). Spironolactone treatment, however, significantly prevented the development of histopathologic and functional changes (BUN 31.2 +/- 2.5 mg/dL; p <0.001 vs. placebo controls). CONCLUSIONS: Low-dose spironolactone and, to a lesser extent, angiotensin receptor-1 blockade can offer renal protection in a mouse model of internal alpha-particle irradiation.

Shockwave lithotripsy: dose-related effects on renal structure, hemodynamics, and tubular function.

BACKGROUND AND PURPOSE: Shockwave lithotripsy (SWL) predictably damages renal tissue and transiently reduces function in both kidneys. This study characterized the effects on renal function of a supraclinical dose of shockwaves (SWs) (8000) in porcine kidneys and tested the hypothesis that such excessive treatment would intensify and prolong the resulting renal impairment. MATERIALS AND METHODS: Pigs aged 6 to 7 weeks were anesthetized and assigned to one of three groups. Groups 1 (N=8) and 2 (N=6) each received 8000 SWs at 24 kV (Dornier HM3) to the lower-pole calix of one kidney. Group 3 (7 pigs) received sham treatment. Renal function was monitored for the first 4 hours after SW treatment in Group 1 and for 24 hours in Group 2. Plasma renin activity was measured in Groups 2 and 3. RESULTS: The renal lesions produced by 8000 SWs comprised 13.8%+/-1.4% of the renal mass. In the 4-hour protocol, this injury was associated with marked reduction of the glomerular filtration rate (GFR), renal plasma flow (RPF), and urinary sodium excretion in both kidneys, although fractional sodium excretion was reduced only in the shocked kidneys. In the 24-hour protocol, GFR and RPF remained below baseline in shocked kidneys at 24 hours. Evidence of progressive ischemic injury was noted in shocked tissue at 24 hours after SW treatment. CONCLUSIONS: These findings support the hypothesis that the severity of the renal injury caused by SWL is related to the number of SWs administered and demonstrate the connection in this relation between renal structure and function.

Role of heat shock protein 47 on tubulointerstitium in experimental radiation nephropathy.

The molecular mechanisms of fibrosis in radiation nephropathy have received scant attention. Heat shock protein 47 (HSP47), a collagen-binding stress protein, helps in the intracellular processing of procollagen molecules during collagen synthesis. We investigated the role of HSP47 in the progression of radiation nephropathy using experimental radiation nephropathy. Experimental rat groups were as follows: (i) group I, sham operated (n = 12); (ii) group II, single doses of irradiation, either 7, 15 or 25 Gy to left kidney (n = 60); and (iii) group III, a similar irradiation procedure as group II after right nephrectomy (n = 60). The rats were followed up until 9 months after renal exposure to radiation. Renal dysfunction (as determined by serum creatinine and blood urea nitrogen) and hypertension were noted in group III rats, along with inflammatory cell infiltration and interstitial fibrosis (as determined by increased deposition of collagens). Compared to control rat kidneys, an increased expression of HSP47 was noted in kidneys obtained from irradiated rats. By double immunostaining, HSP47-expressing cells were identified as alpha-smooth muscle actin-positive myofibroblasts and vimentin-positive tubular epithelial cells. Increased expression of HSP47 was closely associated with increased deposition of collagens in the widened interstitium of irradiated rats. Overexpression of HSP47 by phenotypically altered tubulointerstitial cells might play a role in excessive assembly/synthesis of collagens and could contribute to tubulointerstitial fibrosis in radiation nephropathy.

Angiotensin II blockade reduces radiation-induced proliferation in experimental radiation nephropathy.

Total-body irradiation or renal irradiation is followed by a well-defined sequence of changes in renal function leading eventually to renal failure. Previous studies in a rat model have shown that inhibition of angiotensin-converting enzyme or blockade of angiotensin II receptors can prevent the structural and functional changes that occur after renal irradiation, and that these interventions are particularly important between 3 and 10 weeks after irradiation. We have now shown that in the same rat model, total-body irradiation induces proliferation of renal tubular cells (i.e., an increase in the number of cells staining positive for proliferating cell nuclear antigen) within 5 weeks after irradiation. Treatment with an angiotensin II receptor blocker delays this radiation-induced tubular proliferation and decreases its magnitude. Renal radiation also induces proliferation of glomerular cells, but the relative increase in glomerular proliferation is not as great as that seen in renal tubular cells, and the increase is not delayed or decreased by treatment with an angiotensin II receptor blocker. We hypothesize that angiotensin II receptor blockers exert their beneficial effect in radiation nephropathy by delaying the proliferation (and hence the eventual mitotic death) of renal tubular cells that have been genetically crippled by radiation.

Radiation-induced kidney injury: a role for chronic oxidative stress?

Kidney irradiation clearly leads to a progressive reduction in function associated with concomitant glomerulosclerosis and/or tubulointerstitial fibrosis. However, the particular cell types, mediators and/or mechanisms involved in the development and progression of radiation nephropathy remain ill defined. Angiotensin II (Ang II) plays a major pathogenic role; administration of Ang II blockers markedly abrogates the severity of radiation nephropathy in experimental models. Both ionizing radiation and Ang II signal via generation of reactive oxygen species (ROS). Thus, we hypothesized that localized kidney irradiation might lead to a chronic oxidative stress. In view of the difficulty in measuring ROS in vivo we adopted an indirect immunohistochemical approach in which we used a monoclonal antibody specific for 8-hydroxy-2'-deoxyguanosine (8-OHdG), one of the most commonly used markers of DNA oxidation. The right kidney of 7-8 week-old male Sprague-Dawley rats was removed. Five to 6 weeks later the remaining hypertrophied kidney was irradiated with single doses of 0-20.0 Gy X-rays. Groups of rats, three per dose, were killed at 4, 8, 16 and 24 weeks post-irradiation, their kidneys fixed, and sections stained with the 8-OHdG-specific antibody N45.1. For quantitation of glomerular DNA oxidation with the N45.1 antibody stained sections, 50 glomeruli/animal were counted. The presence of any intensely stained nuclei within the glomerular tuft was scored as positive. Quantitation of tubular DNA oxidation employed a 10 x 10 point ocular grid. Sections were examined at 400 magnification; 250 tubular profiles were counted. All tubules with any nuclear staining were scored as positive.Sham-irradiated kidneys showed little evidence of DNA oxidation over the experimental period. In contrast, localized kidney irradiation led to a marked, dose-independent increase in glomerular and tubular cell nuclear DNA oxidation. This increase was evident at the first time point studied, i.e. 4 weeks after irradiation, and persisted for up to 24 weeks postirradiation. DNA oxidation in the irradiated kidney was only seen in apparently viable glomerular and tubular cells. Thus, while from 16 to 24 weeks post-irradiation structural alterations had progressed to glomerular sclerosis and tubular atrophy, positive staining for 8-OHdG was not observed in severely atrophic tubules. Similarly, fewer positive staining cells were noted in glomeruli undergoing sclerosis, while none were seen in totally sclerotic glomeruli. These data support the hypothesis that renal irradiation is associated with a chronic and persistent oxidative stress.

In vitro and in vivo expression of endothelial von Willebrand factor and leukocyte accumulation after fractionated irradiation.

Previous investigations have demonstrated an increased release of von Willebrand factor (VWF; also known as vWF) in endothelial cells after high single-dose irradiation in vitro. We have also found increased levels of Vwf protein in mouse glomeruli after a high single dose of renal irradiation in vivo. In addition, increased numbers of leukocytes were observed in the renal cortex after irradiation in vivo. The aim of the present study was to investigate and quantify these biological processes after clinically relevant fractionated irradiation and to relate them to changes in renal function. A significantly greater increase in release of VWF was observed in cultured human umbilical vein endothelial cells (HUVECs) after fractionated irradiation (20 x 1.0 Gy) than after a single dose of 20 Gy (147% compared to 115% of control, respectively, P < 0.0005). In contrast with the in vitro observations, glomerular Vwf staining was lower after fractionated irradiation in vivo (20 x 2.0 Gy or 10 x 1.6 Gy +/- re-irradiation) than after a single dose of 16 Gy. The number of leukocytes accumulating in the renal cortex was also lower after fractionated in vivo irradiation than after a single radiation dose. The onset of these events preceded renal functional and histopathological changes by approximately 10 weeks. These data indicate that radiation-induced changes in endothelial VWF expression after in vivo irradiation may be distinct from the in vitro observations. Increased VWF expression may reflect pivotal processes in the pathogenesis of late radiation nephropathy and provide a clue to appropriate timing of pharmacological intervention.

Long-term effects of total-body irradiation on the kidney of Rhesus monkeys.

PURPOSE: To investigate the long-term effects of total-body irradiation (TBI) on kidneys in non-human primates. METHODS AND MATERIALS: The kidneys of Rhesus monkeys were histologically examined at 6-8 years after TBI with low single doses of 4.5-8.5Gy or two fractions of 5.4Gy. The kidneys of age-matched non-irradiated monkeys served as controls. Irradiation was performed on adult monkeys aged about 3 years; 6-8 years later animals were sacrificed and the kidneys removed and processed for histology. A semi-quantitative scoring system was used to evaluate overall histological damage. Glomerular changes were also morphometrically analysed according to previously published criteria. In selected dose groups (pro)thrombotic and inflammatory changes were investigated by immunostaining cryosections with antibodies against von Willebrand factor (vWF), leukocytes and macrophages. RESULTS: Histological changes were generally mild and only seen in kidneys irradiated with doses higher than 7 Gy. Glomerular changes were characterized by increased mesangial matrix and capillary dilatation. Tubulo-interstitial changes included hypercellularity, fibrosis and mild tubular atrophy. The mean glomerular area expressing vWF protein in the irradiated kidneys was not different from that in the age-matched controls. Numbers of infiltrating leukocytes were not significantly different between irradiated kidneys and controls. However, slightly increased numbers of macrophages were present in the renal cortex after irradiation. CONCLUSIONS: Renal damage after TBI of Rhesus monkeys with single doses of 4.5-8.5 Gy or two fractions of 5.4 Gy was mild, even after follow-up times of 6-8 years.

Irradiation of rat tubule epithelial cells alters the expression of gene products associated with the synthesis and degradation of extracellular matrix.

PURPOSE: To determine the ability of radiation to modulate kidney tubule epithelial cell expression of various molecules involved in regulating extracellular matrix accumulation (collagen types I and III, fibronectin, plasminogen activator-inhibitor 1 (PAI-1), TGF-beta and tissue inhibitor of metalloproteinases-2 (TIMP-2)) and degradation (plasminogen activators u-PA or t-PA, MMP-2 and MMP-9). MATERIALS AND METHODS: NRK52E rat tubule epithelial cells were placed in serum-free medium 24 h prior to irradiation with single doses of 1.0-10.0 Gy 137Cs gamma-rays. After irradiation, cells were maintained in serum-free medium for up to 48 h. RESULTS: Irradiation of NRK52E cells was associated with significant dose-dependent increases in collagen I (p<0.05) and PAI-1 (p< or =0.002) mRNA. Collagen III mRNA levels also exhibited a dose-dependent increase; however, this increase failed to reach levels that were statistically significant. Fibronectin mRNA levels were unaltered following irradiation. Radiation also led to an isoform-specific alteration in TGF-beta expression; TGF-beta1 mRNA increased, TGF-beta3 mRNA decreased. The amount of TGF-beta protein secreted into the medium was unchanged following irradiation; however, there was a fivefold increase in the relative amount of active TGF-beta. Irradiation was also associated with differential changes in MMP expression: active MMP-2 levels increased, while MMP-9 levels were unaltered; PA secretion into the medium was unchanged following irradiation. CONCLUSIONS: Irradiation of rat kidney tubule epithelial cells leads to altered production of various molecules associated with extracellular matrix accumulation and degradation.

Stenotic glomerulotubular necks in radiation nephropathy.

Fibrosis of the renal interstitium is well correlated with kidney function and a greater extent of fibrosis predicts renal failure. Recent work has shown striking fibrotic constrictions at the glomerulotubular neck in porcine radiation nephropathy and in Wegener's granulomatosis in man. The present studies were designed to identify stenotic necks in a third species and to evaluate the effect of captopril treatment. Experimental radiation nephropathy was established with 17 Gy total body irradiation of barrier-maintained rats. Kidneys were obtained at 99 and 203 days for histology, using perfusion fixation. There was renal injury, with a rise in BUN, as expected, which was attenuated by captopril treatment. There was stenotic neck formation at 99 and 203 days. Captopril did not influence the absolute fraction of necks that were stenotic but it did prevent the evolution of glomeruli with necks to atubular glomeruli. It is concluded that stenosis of the glomerulotubular neck is a general phenomenon of any scarring kidney disease; that stenotic necks are probably an intermediate step in the evolution towards atubular glomeruli; and that timely use of captopril may prevent the progression of a stenotic neck towards an atubular glomerulus.

The influence of dose per fraction on the pathogenesis of radiation nephropathy.

Both kidneys of male Wistar rats were irradiated with either a 10-Gy single dose or 26 Gy at a rate of 2 Gy per fraction per day. Serum blood urea nitrogen (BUN), creatinine and blood haematocrit levels were assessed prior to radiotherapy and at intervals of 8 weeks thereafter. A subset of animals from each dose group were killed at 4, 8, 16 and 24 weeks and both kidneys of each animal were examined by electron microscopy. In both dose groups a significant increase in BUN and creatinine levels, together with a decrease in haematocrit level, was observed at 16 weeks, and this was followed by an apparent improvement at 24 weeks. There was no statistical difference in these responses between the two groups. The morphological changes in both dose groups were essentially similar, but differed in severity. At 4 weeks after irradiation glomerular and proximal tubular injury were observed in both groups. A marked increase of glomerular and tubular injury in the 10-Gy dose group, without any apparent progression in the 26-Gy dose group, was detected at 8 weeks. By 16 weeks a noticeable improvement in both tubular and glomerular lesions (especially in the 10-Gy dose group) was observed. No apparent difference from the 16th week of evaluation was found at 24 weeks. These findings indicate that there is some recovery in kidney after irradiation, but the extent of the recovery process is somewhat limited.