CDDP Injection Research Articles

Enhanced renal accumulation of cisplatin via renal organic cation transporter deteriorates acute kidney injury in hypomagnesemic rats

This study aimed to explore the effects of hypomagnesemia on cisplatin (CDDP)-induced acute kidney injury (AKI) in rats and the relation of hypomagnesemia to the regulation of organic cation transporters and renal accumulation of CDDP. Sprague-Dawley rats were given an Mg-deficient diet starting 7 days before treatment with CDDP. CDDP was administered intravenously to rats in the normal Mg-diet group and Mg-deficient-diet group at 3 mg/kg via the left jugular vein. At the specified periods after injection of CDDP, the amount of platinum in blood and organ samples was determined using inductively coupled plasma-mass spectrometry. Protein expression levels of renal organic cation transporters were determined. Uptake of tetraethylammonium (TEA) bromide in renal slices of rats was measured. Rats fed a Mg-deficient diet showed a significant body weight decrease and a marked decrease in serum Mg levels compared with control rats fed an adequate Mg diet. Serum blood urea nitrogen and creatinine levels were unaltered after CDDP treatment in control rats, whereas these levels were markedly elevated in hypomagnesemic rats. Immunoblotting revealed up-regulation of the organic cation transporter rOCT2 in hypomagnesemic rats before CDDP administration, but not of rOCT1 or rat multidrug and toxin-extrusion 1. TEA uptake by renal slices from hypomagnesemic rats was significantly higher compared with that of control rats. Renal accumulation of CDDP was markedly increased in hypomagnesemic rats. These results suggest that hypomagnesemia could cause dehydration and up-regulation of rOCT2, enhancing renal accumulation of CDDP and the deterioration of AKI.

Protective Effects of the Angiotensin II Receptor Blocker Losartan on Cisplatin-Induced Kidney Injury

Background: In the present study, we investigated the possible modulatory effect of losartan, an angiotensin receptor blocker, on oxidative stress induced by cisplatin (CDDP) as well as on CDDP uptake by the kidney. Methods: Rats were injected with a single dose of CDDP (7 mg/kg) and/or losartan (in either a single dose of 60 mg/kg or divided doses (10 mg/kg daily for 6 days), starting 1 h before CDDP injection. In addition, rat renal cortical slices were incubated with CDDP (2 mM) and/or losartan (2 mM) for 4 h. Nephrotoxicity was evaluated by measuring serum creatinine and blood urea nitrogen (BUN) in vivo and lactate dehydrogenase (LDH) leakage in vitro; histopathological examination of kidney tissue was also done. Oxidative stress markers including reduced glutathione (GSH) and lipid peroxides were also assessed. Furthermore, CDDP uptake by renal cortical slices was determined. Results: Losartan has protective effects against CDDP-induced nephrotoxicity as evidenced by restoration of normal serum levels of creatinine and BUN, and LDH leakage. Histopathological examination of the kidney confirmed these results. Also, losartan significantly counteracted CDDP-induced lipid peroxidation and GSH depletion. However, losartan did not affect CDDP uptake by the kidney. Conclusion: Our results indicate that losartan has proved to be a promising drug for clinical use as a nephroprotectant against CDDP-induced nephrotoxicity.

Endogenous protection against oxidative stress caused by cisplatin: role of superoxide dismutase

Conclusion: The administration of cisplatin induces the activation of superoxide dismutase (SOD) as a response to oxidative stress in the cochleae of Sprague-Dawley rats and this activation is proportional to the activation of the intrinsic pathway of apoptosis. Objectives: To determine the role of the antioxidant endogenous mechanism in the preservation of cochlear integrity and function in an experimental model of cisplatin ototoxicity. Methods: Sixteen Sprague-Dawley rats were studied at 7 days after intraperitoneal injection of CDDP (n = 8) or 10 ml/kg NaCl 0.9% w/v in the control group (n = 8) by means of auditory steady-state responses. These findings were compared with the expression of SOD and caspase-3/7 and caspase-9 activities. Results: Groups receiving cisplatin showed increased auditory thresholds after injection of cisplatin and control groups maintained normal hearing. Measurements of caspase-3/7 and caspase-9 showed a significant increase in cisplatin-treated rats. A significantly increased activity of total SOD in whole cochlear extracts was observed in animals from the CDDP groups vs control animals. Likewise, differences between CDDP groups were also statistically significant.

Interleukin-6 deficiency accelerates cisplatin-induced acute renal failure but not systemic injury

Cisplatin (CDDP), a major chemotherapeutic agent used to treat solid tumors, is known to induce acute renal failure (ARF). The progression of tissue injury involves the coordination of inflammatory and repair responses. Interleukin-6 (IL-6) has been suggested to modulate inflammatory and repair processes in various tissue injuries. In this study, we analyzed IL-6 regulation during CDDP-induced ARF in wild-type (WT) mice and determined the pathological role of IL-6 using IL-6 knockout ( −/−) mice. A correlation between increase in serum IL-6 level and blood urea nitrogen level was found in WT mice. Renal IL-6 expression in most proximal tubular cells and suppressor of cytokine signaling 3 (SOCS3) gene expression significantly increased in WT mice after administration of CDDP, suggesting active IL-6 signaling during CDDP-induced ARF development. Interestingly, renal dysfunction occurred soon after administration of CDDP and became more severe in IL-6 −/− mice than that in WT mice. In contrast, the survival rate of IL-6 −/− mice (50% at 8 days) was better than that of WT mice (10%). Induction levels of proapoptotic Bcl-2 associated X protein (Bax) in renal proximal tubular cells was significantly higher in IL-6 −/− mice than in WT mice at 24 h after CDDP injection. Levels of antiapoptotic proteins, Bcl-2 and Bcl-extra large (Bcl-x L), in IL-6 −/− groups were significantly higher than those in CDDP-treated WT groups throughout the experimental period. Bax might contribute to the development of CDDP-induced ARF at 24 h; however, high expression levels of Bcl-x L and Bcl-2 might overcome the proapoptosis signaling at 72 h in IL-6 −/− mice. These results indicated that local and systemic elevation of IL-6 contributes to the development of CDDP-induced ARF and that IL-6 produced in renal tubular cells prevents progression of ARF at the early stage. IL-6 deficiency accelerates CDDP-induced ARF but not development of systemic injury.

Protective Effect of Royal Jelly and Green Tea Extracts Effect Against Cisplatin-Induced Nephrotoxicity in Mice: A Comparative Study

The aim of the present study was to investigate the protective role of royal jelly (RJ) and green tea (GT) extracts on cisplatin (cDDP)-induced nephrotoxicity in adult albino mice. Albino mice were randomly divided into six groups: Group I (control) received a single intraperitoneal injection of isotonic saline (0.02 mL/g), Group II received a single intraperitoneal injection of cDDP (7 mg/kg of body weight), Group III received RJ (100 mg/kg of body weight), Group IV received GT (100 mg/kg of body weight), Group V received RJ (100 mg/kg of body weight) + cDDP (7 mg/kg of body weight), and Group VI received GT (100 mg/kg of body weight) + cDDP (7 mg/kg of body weight). The concentrations of blood urea nitrogen (BUN) and creatinine were evaluated. In addition, kidney samples were taken for determination of tissue malondialdehyde (MDA) and reduced glutathione (GSH) levels. In addition, histopathological changes in kidneys were investigated. The results indicated that no significant differences in MDA, GSH, BUN, and creatinine levels were observed among the control group and groups treated with RJ alone and GT alone (P > .05). However, there was a significant increase in BUN and creatinine parameters after cDDP application in Groups II, V, and VI. The mice treated with only cDDP exhibited an increase in serum BUN and creatinine levels when compared to Groups V and VI (P < .05). Moreover, cDDP-induced oxidative damage caused a significant decrease in GSH levels and a significant increase in MDA levels in kidneys (P < .05). RJ and GT supplementation attenuated cDDP-induced nephrotoxicity, which was manifested by stopping the elevation in serum creatinine and BUN levels. Moreover, RJ and GT supplementation restored GSH content and MDA production levels in the kidney tissue following cDDP treatment (P < .05). These products were also effective in protecting against cDDP-induced tissue damage in mouse kidneys. In conclusion, 100 mg/kg of body weight doses of RJ and GT provided protection against cDDP-induced nephrotoxicity, and both products can act as protector agents against cDDP-induced kidney damages.

Histone deacetylase blockade improves renal function and attenuates oxidative stress in rats with cisplatin‐induced acute renal failure

Histone deacetylase (HDAC) inhibition has been shown to prevent cytokine induction and to modulate renal disease in rodent models. We tested the hypothesis that HDAC inhibition would improve various renal and oxidative stress parameters in cisplatin (CDDP)‐induced acute renal failure (ARF). 15‐week old male Sprague‐Dawley rats (n=4 per group) were treated with the HDAC inhibitor, valproic acid (VPA; 300 mg/kg, s.c.), or vehicle, for 1 day prior to and 5 days following induction of ARF with a single 6 mg/kg injection of CDDP (or saline vehicle). At the end of 6 days, anesthetized rats underwent acute renal experiments to determine renal blood flow (RBF) and glomerular filtration rate (GFR) using PAH and inulin clearance techniques, respectively. Rats were sacrificed and kidneys removed; renal cortices were isolated for measurement of tissue and mitochondrial free radical levels using electron paramagnetic resonance spectroscopy. Tissue mRNA and protein levels of HDAC and oxidative stress genes were determined using real‐time RT‐PCR and western blot, respectively. No significant changes in mean arterial pressure (MAP) were noted among groups. VPA treatment improved RBF and GFR (p&lt;0.05) and attenuated the higher levels of total tissue and mitochondrial ROS and tissue superoxide in ARF rats. VPA treatment also significantly decreased tissue mRNA and protein expression levels of HDAC and oxidative stress genes, and normalized ATP production and ADP/ATP ratio in rats with ARF. These findings suggest that inhibition of class I HDACs with VPA improves the renal oxidative stress and tissue and mitochondrial injury that accompany CDDP‐induced ARF. Grant support: NIH grant RO1HL080544‐01 for Dr. Joseph Francis.

Renoprotection by α-mangostin is related to the attenuation in renal oxidative/nitrosative stress induced by cisplatin nephrotoxicity

Cisplatin (CDDP) is a chemotherapeutic agent that produces nephrotoxicity associated with oxidative/nitrosative stress. α-Mangostin (α-M) is a xanthone extracted from mangosteen with antioxidant and anti-inflammatory properties. The purpose of this study was to evaluate the renoprotective effect of α-M on the CDDP-induced nephrotoxicity. α-M was administered (12.5 mg/kg/day, i.g.) for 10 days (7 days before and 3 days after CDDP injection). On day 7, rats were treated with a single injection of CDDP (7.5 mg/Kg, i.p.); 3 days after the rats were killed. α-M attenuated renal dysfunction, structural damage, oxidative/nitrosative stress, decrease in catalase expression and increase in mRNA levels of tumour necrosis factor alpha and transforming growth factor beta. In conclusion the renoprotective effect of α-M on CDDP-induced nephrotoxicity was associated with the attenuation in oxidative/nitrosative stress and inflammatory and fibrotic markers and preservation of catalase activity.

The dimethylthiourea-induced attenuation of cisplatin nephrotoxicity is associated with the augmented induction of heat shock proteins

Dimethylthiourea (DMTU), a potent hydroxyl radical scavenger, affords protection against cisplatin (CDDP)-induced acute renal failure (ARF). Since the suppression of oxidative stress and the enhancement of heat shock proteins (HSPs) are both reported to protect against CDDP-induced renal damage, we tested whether increased HSP expression is involved in the underlying mechanisms of the DMTU-induced renal protection. We examined the effect of DMTU treatment on the expression of HSPs in the kidney until day 5 following a single injection of CDDP (5 mg/kg BW). DMTU significantly inhibited the CDDP-induced increments of serum creatinine, the number of 8-hydroxyl-2′-deoxyguanosine (8-OHdG)- and terminal deoxynucleotidyl transferase nick-end labeling (TUNEL)-positive tubular cells, and tubular damage score ( p < 0.05). CDDP significantly increased renal abundances of HO-1, HSP60, HSP72 and HSP90 at days 1, 3, and 5. DMTU significantly augmented only the expression of HSP60 expression mainly in the cytoplasm of the proximal tubular cells at days 1 and 3 in CDDP-induced ARF. DMTU also inhibited the CDDP-induced increment of Bax, a pro-apoptotic protein, in the fraction of organelles/membranes at day 3. The findings suggest that DMTU may afford protection against CDDP-induced ARF, partially through the early induction of cytoplasmic HSP60, thereby preventing the Bax-mediated apoptosis in renal tubular cells.

The anticancer drug cisplatin induces an intrinsic apoptotic pathway inside the inner ear

Ototoxicity is a known adverse effect of cisplatin (CDDP). Since apoptosis is involved in the development of some pathological conditions associated with the administration of anticancer drugs, we examined, using immunohistochemical and electrophysiological techniques, the apoptotic changes in the cochlea of Sprague-Dawley (SD) rats after an injection of CDDP (5 mgkg(-1) body weight). Luciferase assays were used to determine the different caspase activities and ATP levels in protein extracts of whole cochleae. The expression of several apoptotic-related proteins was measured by means of Western blotting. These analyses were performed 2, 7 and 30 days after the CDDP injection. The auditory brain stem response was obtained before and at the different times after the injection of CDDP, before the animals were killed. CDDP significantly increased the levels of caspase-3/7 activity and active caspase-3 protein expression and caspase-3 immunofluorescence staining, caspase-9 activity, and Bax protein expression but decreased Bcl-2 protein expression within the rat cochleae. Threshold shifts were significantly elevated 2 days after CDDP treatment. These findings support the hypothesis that cisplatin-related apoptosis evokes an intrinsic pathway of pro-apoptotic signalling within the rat cochleae. Thus, selective inhibition of the sequence of events involved in the intrinsic apoptotic pathway could provide a strategy to minimize cisplatin-induced ototoxicity.

Chemotherapy Enhances CD8+ T Cell-mediated Antitumor Immunity Induced by Vaccination With Vaccinia Virus

The use of immunotherapy or chemotherapy alone is generally ineffective against well-established tumors. To overcome this intrinsic resistance against therapy for tumors, we have attempted to combine immunotherapy with chemotherapy. In this study, we tried to induce a rapid antitumor effect via chemoimmunotherapy using a vaccinia viral vaccine as an immunotherapeutic agent with anticancer agents including epigallocatechin-3-gallate (EGCG) and conventional anticancer drugs. Although a combination of vaccinia-mediated vaccination and chemotherapy led to a strong inhibition of tumor growth, monotherapy alone failed to completely cure tumors. In contrast, intravenous injection of cisplatin (CDDP) or cyclophosphamide (CTX) after vaccinia virus vaccination led to complete regression of the established tumors. Interestingly, anticancer drugs appear to augment the antitumor effect of the vaccinia virus-mediated immunotherapy. This effect is mainly associated with the enhanced tumor-specific CD8(+) T cell immune response induced by vaccinia virus, which was demonstrated by antibody depletion. However, anticancer drugs alone failed to induce a significant enhancement of the tumor-specific CD8(+) T cell immune response. Taken together, these results suggest that combining vaccinia virus-based immunotherapy with anticancer drugs is particularly effective against established tumors by increasing the tumor antigen-specific CD8(+) T cell immune response, which is primed by vaccinia virus-mediated vaccination.

Cisplatin-induced peripheral neuropathy: Neuroprotection by erythropoietin without affecting tumour growth

This study examined the dose-dependent efficacy of erythropoietin (EPO) for preventing and/or treating cisplatin (CDDP) induced peripheral neurotoxicity (CINP), and its influence on tumour treatment and growth. Rats received eight intraperitoneal (ip) injections of 2 mg/kg CDDP twice weekly. EPO co-administered (50 or 10 μg/kg ip, three times/week) had a dose-dependent effect, partially preventing CINP, but 0.5 μg/kg ip was not effective. The neuroprotective effect lasted at least 5 weeks after the last dose of EPO and CDDP. In addition, EPO (50 μg/kg ip three times/week) after the last injection of CDDP still induced a significant recovery of CINP. In a separate experiment in rats bearing mammary carcinoma EPO treatment (50 μg/kg ip) given concurrently with CDDP (1.0 and 1.5 mg/kg twice a week for four weeks) was neuroprotective without influencing the effectiveness of the treatment or tumour growth. EPO thus appears to be an effective neuroprotectant that does not interfere with tumour treatment.

Silymarin Modulates Cisplatin-Induced Oxidative Stress and Hepatotoxicity in Rats

Cisplatin (CDDP) is a widely used anticancer drug, but at high dose, it can produce undesirable side effects such as hepatotoxicity. Because silymrin has been used to treat liver disorders, the protective effect of silymarin on CDDP-induced hepatotoxicity was evaluated in rats. Hepatotoxicity was determined by changes in serum alanine aminotransferase [ALT] and aspartate aminotransferase [AST], nitric oxide [NO] levels, albumin and calcium levels, and superoxide dismutase [SOD], glutathione peroxidase [GSHPx] activities, glutathione content, malondialdehyde [MDA] and nitric oxide [NO] levels in liver tissue of rats. Male albino rats were divided into four groups, 10 rats in each. In the control group, rats were injected i.p. with 0.2 ml of propylene glycol in saline 75/25 (v/v) for 5 consecutive days [Silymarin was dissolved in 0.2 ml of propylene glycol in saline 75/25 v/v]. The second group were injected with CDDP (7.5 mg /kg, I.P.), whereas animals in the third group were i.p. injected with silymarin at a dose of 100 mg/kg/day for 5 consecutive days. The Fourth group received a daily i.p. injection of silymarin (100 mg/kg/day for 5 days) 1 hr before a single i.p. injection of CDDP (7.5 mg/kg). CDDP hepatotoxicity was manifested biochemically by an increase in serum ALT and AST, elevation of MDA and NO in liver tissues as well as a decrease in GSH and the activities of antioxidant enzymes, including SOD, GSHPx in liver tissues. In addition, marked decrease in serum NO, albumin and calcium levels were observed. Serum ALT, AST, liver NO level, MDA was found to decreased in the combination group in comparison with the CDDP group. The activities of SOD, GSHPx, GSH and serum NO were lower in CDDP group than both the control and CDDP pretreated with silymarin groups. The results obtained suggested that silymarin significantly attenuated the hepatotoxicity as an indirect target of CDDP in an animal model of CDDP-induced nephrotoxicity.

Inhibition of p21 modifies the response of cortical proximal tubules to cisplatin in rats

The purpose of this study was to evaluate whether upregulated p21, a cell cycle-inhibitory protein, contributes to cisplatin (CDDP)-induced acute renal failure (ARF) and to acquired resistance to rechallenge injury with CDDP in rats. ARF was induced in rats by injection of CDDP (5 mg/kg) and rechallenge injury to CDDP by the same dose of CDDP 14 days after the first CDDP injection. Rats were treated with p21 antisense oligodeoxynucleotide (ODN) or its vehicle, p21 sense ODN, every 36 h from days 0 to 5 for single CDDP and from days 13 to 19 for rechallenge injury and killed at day 3, 5, 16, or 19. The uptake of FITC-labeled p21 antisense ODNs by cortical proximal tubule (PT) cells was much greater than by PT cells in the outer stripe of outer medulla (OSOM). Administration of antisense induced partial downregulation of p21 mRNA and protein levels in whole kidneys with single CDDP treatment and its rechallenge injury. Antisense significantly aggravated PT necrosis and decreased the number of p21-positive PT cells in the cortex but not in the OSOM in both CDDP-induced ARF and its rechallenge injury. However, antisense did not alter serum creatinine (Scr) and blood urea nitrogen (BUN) levels. Our findings suggested that p21 plays, at least in part, a cytoprotective role in cortical PTs exposed to CDDP, although this does not contribute to renal dysfunction when judged by Scr and BUN levels. Because antisense may not adequately be taken up and/or function in PTs in the OSOM, the role of p21 in PTs in the OSOM in CDDP-induced ARF remains to be clarified.

Magnetic resonance imaging-based prospective detection of intraperitoneal human ovarian carcinoma xenografts treatment response

The feasibility of applying magnetic resonance imaging (MRI) for conducting prospective studies of intraperitoneal (i.p.) tumor treatment response to chemotherapy and resultant effects on survival in human ovarian carcinoma/nude mouse orthotopic xenograft models was evaluated. Female nude mice were implanted i.p. with either NMP-1 or SKOV-3ip. human ovarian carcinoma cells on day 0. Initial T2-weighted magnetic resonance (MR) images of the abdomens of NMP-1-implanted mice were obtained on day 7 to confirm the presence of nascent tumors; similar confirmations were made on day 14 with mice bearing SKOV-3ip. xenografts. On the initial imaging days, a multiple-dose regimen of cisplatin (CDDP; qd7 x3) was commenced, using 4 or 6 mg/kg treatments with the NMP-1 model and using 6 mg/kg treatments with the SKOV-3ip. model. Mice were reimaged multiple times, 2 days following each CDDP injection and at later times as well, depending on host survival. The images for each mouse from the last imaging day (day 30 for NMP-1, day 44 for SKOV-3ip.) were used in a blinded fashion to attempt to visually distinguish control from treated mice and to determine whether MRI could predict a survival benefit. For SKOV-3ip. mice, ten out of ten mice were correctly segregated into the control or the CDDP treatment group based solely on these blinded, nonquantified MR results. In this model, the 6 mg/kg multiple-dose regimen achieved a modest response, improving life span by approximately 24%. However, for the NMP-1 mice, only six out of nine evaluable mice were correctly segregated into the control or one of the treatment groups by similar MRI criteria, a virtually random distribution; further, neither CDDP treatment regimen achieved a significant improvement in survival in this model. In another study, NMP-1-implanted mice were treated on day 7 after tumor implantation with a single injection of a hyaluronic acid-paclitaxel copolymer. Control and treated mice were MR imaged on day 28, which revealed marked reductions in tumor burden in treated mice, correlating well with a subsequently observed improved survival of approximately 40%. Our results suggest that MRI can be used to serially and noninvasively monitor treatment response and predict ongoing treatment effects on survival.

Magnetic resonance imaging–based prospective detection of intraperitoneal human ovarian carcinoma xenografts treatment response

The feasibility of applying magnetic resonance imaging (MRI) for conducting prospective studies of intraperitoneal (i.p.) tumor treatment response to chemotherapy and resultant effects on survival in human ovarian carcinoma/nude mouse orthotopic xenograft models was evaluated. Female nude mice were implanted i.p. with either NMP-1 or SKOV-3ip. human ovarian carcinoma cells on day 0. Initial T2-weighted magnetic resonance (MR) images of the abdomens of NMP-1–implanted mice were obtained on day 7 to confirm the presence of nascent tumors; similar confirmations were made on day 14 with mice bearing SKOV-3ip. xenografts. On the initial imaging days, a multiple-dose regimen of cisplatin (CDDP; qd7 ×3) was commenced, using 4 or 6 mg/kg treatments with the NMP-1 model and using 6 mg/kg treatments with the SKOV-3ip. model. Mice were reimaged multiple times, 2 days following each CDDP injection and at later times as well, depending on host survival. The images for each mouse from the last imaging day (day 30 for NMP-1, day 44 for SKOV-3ip.) were used in a blinded fashion to attempt to visually distinguish control from treated mice and to determine whether MRI could predict a survival benefit. For SKOV-3ip. mice, ten out of ten mice were correctly segregated into the control or the CDDP treatment group based solely on these blinded, nonquantified MR results. In this model, the 6 mg/kg multiple-dose regimen achieved a modest response, improving life span by ∼24%. However, for the NMP-1 mice, only six out of nine evaluable mice were correctly segregated into the control or one of the treatment groups by similar MRI criteria, a virtually random distribution; further, neither CDDP treatment regimen achieved a significant improvement in survival in this model. In another study, NMP-1–implanted mice were treated on day 7 after tumor implantation with a single injection of a hyaluronic acid–paclitaxel copolymer. Control and treated mice were MR imaged on day 28, which revealed marked reductions in tumor burden in treated mice, correlating well with a subsequently observed improved survival of ∼40%. Our results suggest that MRI can be used to serially and noninvasively monitor treatment response and predict ongoing treatment effects on survival.

452. The Enhancement of Anticancer Effect of Chemotherapy by the Delivery of Rad51 siRNA

Top of pageAbstract Chemotherapy is one of the effective therapies to cancer, however, cancers gradually develop machineries to resist to chemotherapy. To overcome the problem of drug resistance, we tested the combination of short interfering RNA (siRNA) against Rad51, related to the homologous recombination repair, with cis-diamminedichloroplatinum (II) (CDDP; cisplatin). Moreover, to deliver siRNA into cancer cells effectively, we have developed the HVJ (hemagglutinating virus of Japan, Sendai virus) envelope vector which has the high membrane fusion ability. When synthetic Rad51 siRNA was delivered to HeLa cells using HVJ envelope vector, no Rad51 transcripts were detected on day 2, and Rad51 protein completely disappeared for 4 days after siRNA transfer. When HeLa cells were incubated with 0.02 ug/ml CDDP for 3 hours after siRNA transfer, the number of colonies decreased to approximately 10% of that with scrambled siRNA. The sensitivity to CDDP was enhanced in various human cancer cells, but not in normal human fibroblasts. The synthetic Rad51 siRNA was also introduced into subcutaneous tumor mass of HeLa cells in SCID mice with or without intraperitoneal injection of CDDP, and tumor growth was monitored. When Rad51 siRNA was delivered into tumors using HVJ envelope vector, the delivery efficiency was approximately 50%, and the Rad51 transcript level was reduced to approximately 25%. Rad51 siRNA combined with CDDP significantly inhibited the tumor growth when compared to siRNA or CDDP alone. Rad51 siRNA could enhance the sensitivity to CDDP in cancer cells both in vitro and in vivo. Rad51 siRNA also enhanced the sensitivity of cancer cells to bleomycin. Our results suggest that the combination of CDDP and Rad51 siRNA will be an effective anticancer protocol.

Protective effect of dietary azuki bean ( Vigna angularis) seed coats against renal interstitial fibrosis of rats induced by cisplatin

Objective We investigated the effects of azuki bean ( Vigna angularis) seed coats (ABSCs), which mainly contain proanthocyanidins and dietary fibers, on the infiltration of macrophages and the progression of renal interstitial fibrosis induced by cisplatin (CDDP). Methods Male rats were divided into two groups: controls received intraperitoneal injections of saline and the other rats were injected intraperitoneally with 3 mg of CDDP per kilogram once a week for 5 wk. The CDDP-injected animals received one of four diets: 1) control diet (commercial diet only), 2) 0.5% red ABSC (RABSC) diet, 3) 2.0% RABSC diet, and 4) 2.0% white ABSC (WABSC) diet. The saline-injected animals were given the commercial diet. Five weeks after the final CDDP injection, macrophage kinetics and interstitial fibrotic areas were examined. Results The content of polyphenols in the RABSC (76.3 g/kg of plant material) was higher than that in the WABSC (18.1 g/kg). Proanthocyanidins were detected in the RABSC (20.4 g/kg) but not in the WABSC. Histologically, the fibrotic areas consisting of fibroblastic cells and mononuclear cells developed around the dilated or atrophic tubules in the corticomedullary junction in CDDP-treated rat kidney, whereas the extent and magnitude of damage were less in the WABSC- and RABSC-treated rats. In immunohistochemical analysis, ED1-positive macrophages in CDDP-treated rats showed a significant increase in number compared with the control. The number of macrophages in CDDP plus WABSC or RABSC groups was significantly smaller than that in CDDP-treated rats. In addition, the number of macrophages in the RABSC group was significantly smaller than in the WABSC group, indicating that ABSC, especially RABSC, prevented macrophages from infiltrating into areas of interstitial fibrosis. Conclusions These results suggest that ABSC, especially RABSC, suppress the increase of infiltrating macrophages in the damaged kidney and may lead to amelioration of interstitial fibrosis. Based on the composition of ABSC, molecules such as proanthocyanidins and/or dietary fibers may be associated with the amelioration of renal damage.

Rad51 siRNA delivered by HVJ envelope vector enhances the anti‐cancer effect of cisplatin

Every cancer therapy appears to be transiently effective for cancer regression, but cancers gradually transform to be resistant to the therapy. Cancers also develop machineries to resist chemotherapy. Short interfering RNA (siRNA) has been evaluated as an attractive and effective tool for suppressing a target protein by specifically digesting its mRNA. Suppression of the machineries using siRNA may enhance the sensitivity to chemotherapy in cancers when combined with an effective delivery system. To enhance the anti-cancer effect of chemotherapy, we transferred siRNA against Rad51 into various human cancer cells using the HVJ (hemagglutinating virus of Japan, Sendai virus) envelope vector in the presence or absence of cis-diamminedichloroplatinum(II) (CDDP, cisplatin). The inhibition of cell growth was assessed by a modified MTT assay, counting cell number, or fluorescence-activated cell sorting (FACS) analysis after Annexin V labeling. The synthetic Rad51 siRNA was also introduced into subcutaneous tumor masses of HeLa cells in SCID mice with or without intraperitoneal injection of CDDP, and tumor growth was monitored. When synthetic Rad51 siRNA was delivered into HeLa cells using the HVJ envelope vector, no Rad51 transcripts were detected on day 2, and Rad51 protein completely disappeared for 4 days after siRNA transfer. When HeLa cells were incubated with 0.02 microg/ml CDDP for 3 h after siRNA transfer, the number of colonies decreased to approximately 10% of that with scrambled siRNA. The sensitivity to CDDP was enhanced in various human cancer cells, but not in normal human fibroblasts. When Rad51 siRNA was delivered into tumors using the HVJ envelope vector, the Rad51 transcript level was reduced to approximately 25%. Rad51 siRNA combined with CDDP significantly inhibited tumor growth when compared to siRNA or CDDP alone. Rad51 siRNA could enhance the sensitivity to CDDP in cancer cells both in vitro and in vivo. Our results suggest that the combination of CDDP and Rad51 siRNA will be an effective anti-cancer protocol.

Cisplatin-Induced Renal Interstitial Fibrosis in Neonatal Rats, Developing as Solitary Nephron Unit Lesions

Cisplatin (CDDP)-induced renal lesions in rats prove a useful model for analysis of the pathogenesis of post-tubular injury-renal interstitial fibrosis. This study investigated the histopathological changes in 10-day-old neonatal rats induced by a single injection of CDDP (4.5 mg/kg). Compared with age-matched controls, on postinjection (PI) days 1 to 6, the number of apoptotic cells, demonstrable with TUNEL method, was significantly increased in CDDP-treated neonates, and there was no marked epithelial necrosis nor fibrotic lesions. Fibrotic lesions began to be developed solitarily around some nephrons with dilated ducts in the corticomedullary junction on PI day 10 and the lesions became more prominent until PI day 20. The alpha-SMA-positive myofibroblastic cells were seen exclusively in the fibrotic lesions. Additionally, the numbers of macrophages reacting with EDI (specific for exudate macrophages), ED2 (for resident macrophages), and OX6 (recognizing MHC class II antigens expressed in antigen-presenting macrophages/dendritic cells) were significantly increased around the affected renal tubules. A greater immunoreaction for TGF-beta1 was seen mostly in the renal epithelial cells of CDDP-treated neonates. These findings indicated that macrophage populations and myofibrolastic cells as well as TGF-beta1 may be responsible for the production of neonatal renal interstitial fibrosis. Compared with CDDP-injected adult rats that develop extensive interstitial fibrosis (Yamate et al., J Comp Pathol, 1995), the formation of fibrotic lesions was delayed, and the lesions were limited to the area around the affected nephrons; this could be attributable to differences in renal morphology between neonates and mature kidney of adult rats.

Does cisplatin (CDDP) function as a modulator of 5-fluorouracil (5-FU) antitumor action? A study based on a clinical trial

To clarify whether CDDP acts as a modulator of 5-FU antitumor action in gastric cancer, patients were treated preoperatively with 5-FU + CDDP (FP) chemotherapy. From September 2000 to November 2001 at Takarazuka Municipal Hospital, 29 patients preoperatively diagnosed with stages II-IV gastric cancer were enrolled. Written informed consent was obtained from all patients. The patients were randomly assigned to two groups: the FU group, in which patients received a continuous intravenous infusion of 5-FU 320 mg/m2 per day over 24 h a day for 5 days beginning 5 days prior to surgery, and the FP group, in which patients received bolus intravenous injections of CDDP 3.5 mg/m2 per day for 5 days prior to surgery in addition to the same infusion of 5-FU as the FU group. As indicators of the intracellular effect of 5-FU treatment, thymidylate synthase (TS) inhibition rates, TS protein levels, TS and dihydropyrimidine dehydrogenase (DPD) activity, and F-RNA concentrations were measured. Using Scheffe's multiple comparison test, in both treatment groups the tumor regions were found to have significantly higher TS inhibition rates than the nontumor regions (P<0.05). No significant differences in TS protein levels, TS activity, DPD activity or F-RNA concentrations were found between the four regions. Our results show that CDDP clinically may act to enhance the antitumor effects of 5-FU in terms of the inhibition of DNA synthesis and could therefore act as a modulator of 5-FU.