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In this study, we aimed to evaluate the toxic effects, changes in life span, and expression of various metabolism-related genes in Caenorhabditis elegans, using RNA interference (RNAi) and mutant strains, after 3-bromopyruvate (3-BrPA) treatment. C. elegans was treated with various concentrations of 3-BrPA on nematode growth medium (NGM) plates, and their survival was monitored every 24 h. The expression of genes related to metabolism was measured by the real-time fluorescent quantitative polymerase chain reaction (qPCR). Nematode survival in the presence of 3-BrPA was also studied after silencing three hexokinase (HK) genes. The average life span of C. elegans cultured on NGM with 3-BrPA was shortened to 5.7 d compared with 7.7 d in the control group. hxk-1, hxk-2, and hxk-3 were overexpressed after the treatment with 3-BrPA. After successfully interfering hxk-1, hxk-2, and hxk-3, the 50% lethal concentration (LC50) of all mutant nematodes decreased with 3-BrPA treatment for 24 h compared with that of the control. All the cyp35 genes tested were overexpressed, except cyp-35B3. The induction of cyp-35A1 expression was most obvious. The LC50 values of the mutant strains cyp-35A1, cyp-35A2, cyp-35A4, cyp-35B3, and cyp-35C1 were lower than that of the control. Thus, the toxicity of 3-BrPA is closely related to its effect on hexokinase metabolism in nematodes, and the cyp-35 family plays a key role in the metabolism of 3-BrPA.
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OBJECTIVE@#To investigate the effect of down-regulation of miR-205-5p on 3-bromopyruvate-induced apoptosis in human nasopharyngeal carcinoma CNE2Z cells.@*METHODS@#Nasopharyngeal carcinoma CNE2Z cells were transfected with miR- 205-5p-mimic or miR-205-5p-inhibitor, treated with 80 μmol/L 3-bromopyruvate alone, or exposed to both of the treatments. The proliferation of the treated cells was examined with MTT assay, and early apoptosis of the cells was detected using a mitochondrial membrane potential detection kit (JC-1). DAPI fluorescence staining was used to detect morphological changes of the cell nuclei and late cell apoptosis; Annexin V-FITC/PI double staining was employed to detect the cell apoptosis rate. Western blotting was used to detect the expressions of Bcl-2, Bax, Mcl-1 and Bak proteins.@*RESULTS@#Exposure to 3-bromopyruvate significantly inhibited the proliferation of CNE2Z cells, and increasing the drug concentration and extending the treatment time produced stronger inhibitory effects. Treatment with 80 μmol/L 3-bromopyruvate for 24, 48 and 72 h resulted in inhibition rates of (45.7±1.21)%, (64.4±2.02)% and (78.3±1.55)% in non-transfected CNE2Z cells, respectively; the inhibition rates were (27.7±1.04)%, (34.8±2.10)% and (44.3±1.57)% in the cells transfected with miR-205-5p-mimic, and were (80.5 ± 0.94)%, (87.9 ± 0.50)% and (93.8 ± 1.16)% in cells transfected with miR-205-5p-inhibitor, respectively. The results of mitochondrial membrane potential detection showed that the relative proportion of red and green fluorescence decreased significantly in miR-205-5p-inhibitor-transfected cells with 3-bromopyruvate treatment. Combined treatment of the cells with 3-bromopyruvate and miR-205-5p-inhibitor transfection obviously increased nuclear fragmentation and nuclear pyknosis and significantly increased cell apoptotic rate as compared with the two treatments alone ( < 0.01), causing also decreased expressions of Bcl-2 and Mcl-1 proteins and increased expressions of Bax and Bak proteins.@*CONCLUSIONS@#Inhibition of miR-205-5p enhances the proapototic effect of 3-bromopyruvate in CNE2Z cells possibly in relation to the down-regulation of Mcl-1 and Bcl-2 and the up-regulation of Bak and Bax proteins.
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ABSTRACT PURPOSE: To investigate the hepatotoxicity and nephrotoxicity of 3-Bromopyruvate (3BP) in mice. METHODS: Fifteen nude mice were grafted subcutaneously in the left flank with MDA-MB-231 cells, then all mice were divided into control group (PBS), 3BP group (8 mg/kg), positive group (DNR: 0.8 mg/kg) when tumor volume reached approximately 100 mm3. 28 days later, tumors, livers and kidneys were stored in 4 % formalin solution and stained with hematoxylin and eosin staining. The Kunming mice experiment included control group (PBS), 3BP group (4mg/kg; 8mg/kg; 16mg/kg), positive group (DNR: 0.8 mg/kg). 24 hours later, the blood were used for the determination of hepatic damage serum biomarkers. Livers were stored in 4 % formalin solution for the later detection. RESULTS: 3BP at the dose of 8mg/kg had a good effect on inhibiting tumor growth in nude mice and did not damage liver and kidney tissues. Kunming mice experiment showed 3BP at the dose of 16mg/kg did damage to liver tissues. CONCLUSION: 3-Bromopyruvate at the dose of suppressing tumor growth did not exhibit hepatotoxicity and nephrotoxicity in nude mice, and the effect on liver was confirmed in Kunming mice.
Subject(s)
Animals , Female , Mice , Pyruvates/toxicity , Enzyme Inhibitors/toxicity , Chemical and Drug Induced Liver Injury/pathology , Acute Kidney Injury/pathology , Kidney/drug effects , Liver/drug effects , Disease Models, Animal , Dose-Response Relationship, Drug , Acute Kidney Injury/chemically induced , Liver Neoplasms, Experimental/drug therapy , Mice, Inbred BALB C , Mice, NudeABSTRACT
Objective To investigate the effect of 3-bromopyruvate (3-BrPA) on transplanted rectal tumors in experimental rabbit models. Methods A total of 60 New Zealand white rabbits with transplanted rectal tumor were randomly and equally divided into low-dose (0.5 mmol/L), medium-dose (1.0 mmol/L), high-dose (2.0 mmol/L) treatment groups and saline control group with 15 rabbits in each group. Arterial perfusion of 10 ml 3-BrPA with concentration of 0.5 mmol/L, 1.0 mmol/L and 2.0 mmol/L via caudal mesenteric artery was respectively employed for the rabbits of the corresponding treatment group; the control group was perfused with equal amounts of saline. Four days later, rectal tumors were removed by vivisection. The necrosis degree of tumor cells was determined by microscopic examination, and the necrosis rate was calculated. The effect of different 3-BrPA concentrations on the rectal tumor was evaluated. Results The rectal tumor transplantation and transcatheter 3-BrPA or saline perfusion was successfully completed in all 60 experimental rabbits. Microscopically, tumor cells showed different degrees of damage in experimental rabbits. In low-dose (0.5 mmol/L) treatment group, gradeⅠnecrosis was observed in 3 rabbits, gradeⅡin 11 rabbits, and gradeⅢin one rabbit;the effective rate was 6.7%. In medium-dose (1.0 mmol/L) treatment group, gradeⅡnecrosis was seen in 2 rabbits, grade Ⅲ in 10 rabbits, and grade Ⅳ in 3 rabbits; the effective rate was 86.6%. In high-dose (2.0 mmol/L) treatment group, gradeⅢnecrosis was detected in 2 rabbits and gradeⅣin 13 rabbits;the effective rate was 100.0%. In the saline control group, grade I necrosis was observed in 15 rabbits. Statistically significant differences in tumor necrosis rate and effective rate existed between medium-dose (1.0 mmol/L) treatment group and high-dose (2.0 mmol/L) treatment group (P<0.05). Statistically significant differences in tumor necrosis rate also existed between each other among the four groups with necrosis of gradeⅠto gradeⅣ(P<0.05). 3-BrPA had obvious therapeutic effect, while it showed no damage to the normal intestinal tissue. Conclusion For the treatment of transplanted rectal tumor in rabbit models, arterial infusion of 3-BrPA has certain therapeutic effect. In the high-dose group, the necrosis rate and effective rate are the highest, and the therapeutic results are the most significant.
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OBJECTIVE: The purpose of this study was to compare the antitumor effect and hepatotoxicity of an intraarterial delivery of low-dose and high-dose 3-bromopyruvate (3-BrPA) and those of a conventional Lipiodol-doxorubicin emulsion in a rabbit VX2 hepatoma model. MATERIALS AND METHODS: This experiment was approved by the animal care committee at our institution. VX2 carcinoma was implanted in the livers of 36 rabbits. Transcatheter intraarterial administration was performed using low dose 3-BrPA (25 mL in a 1 mM concentration, n = 10), high dose 3-BrPA (25 mL in a 5 mM concentration, n = 10) and Lipiodol-doxorubicin emulsion (1.6 mg doxorubicin/ 0.4 mL Lipiodol, n = 10), and six rabbits were treated with normal saline alone as a control group. One week later, the proportion of tumor necrosis was calculated based on histopathologic examination. The hepatotoxicity was evaluated by biochemical analysis. The differences between these groups were statistically assessed with using Mann-Whitney U tests and Kruskal-Wallis tests. RESULTS: The tumor necrosis rate was significantly higher in the high dose group (93% +/- 7.6 [mean +/- SD]) than that in the control group (48% +/- 21.7) (p = 0.0002), but the tumor necrosis rate was not significantly higher in the low dose group (62% +/- 20.0) (p = 0.2780). However, the tumor necrosis rate of the high dose group was significantly lower than that of the Lipiodol-doxorubicin treatment group (99% +/- 2.7) (p = 0.0015). The hepatotoxicity observed in the 3-BrPA groups was comparable to that of the Lipiodol-doxorubicin group. CONCLUSION: Even though intraarterial delivery of 3-BrPA shows a dose-related antitumor effect, single session treatment seems to have limited efficacy when compared with the conventional method.