Regulation of nitric oxide donor JS-K on tumor energy metabolism in H22 tumor-bearing mice / 中国药理学与毒理学杂志

OBJECTIVE To investigate the regulation of{O2 (2,4-dinitrophenyl)1-[(4-ethoxycarbonyl) piperazin-1-yl] diazen-1-ium-1, 2-diolate} (JS-K), a nitric oxide donor, on tumor energy metabolism in H22 tumor-bearing mice. METHODS The hepatoma animal model in BALB/c mice was established with H22 cell line. The inoculated mice were randomly divided into four groups. The JS-K group and model group received JS-K (0.75 and 1.50 mg?kg-1) and saline via tail the vein intravenously once every 3 d for 14 d, and 5 injections, respectively. The fluorouracil (5-FU) group received 5-FU 20 mg·kg-1 by intra-peritoneal injection once a day for 14 d. On the 15th day after the first administration, mice were sacri-ficed and the tumor, thymus and spleen were isolated and weighed immediately. The tumor growth inhibitory rate and organ index were calculated. The activities of hexokinase (HK), phosphofructokinase (PFK), pyruvate kinase (PK), succinate dehydrogenase (SDH), adenosinetriphosphatase (ATPase), and the levels of lactic acid (LD) and adenosine triphosphate (ATP) in tumor tissues were determined by colorimetric method. The expression of hypoxia-inducible factor 1 alpha (HIF-1α) and hexokinaseⅡ(HKⅡ) in the tumor tissue was analyzed by Western blotting. RESULTS Compared with model group, the tumor mass of JS-K 0.75 and 1.50 mg · kg-1 groups was significantly reduced (P<0.01), and the tumor growth inhibitory rate was 23.9%and 50.3%, respectively. There was no diffrence in thymic and splenic indexes between JS-K group and model group. The activity of HK, PFK, SDH, PK and ATPase of tumor tissue in model group was 22.6±3.7, 14.4±2.6, (10.5±2.6) U·g-1protein, (12.9±3.2) kU·g-1 protein and (0.70 ± 0.10) mmolPi · g-1protein · h-1, respectively, which dropped by 42.0%, 26.6%, 22.7%, 23.3%and 21.7%respectively (P<0.01, P<0.05) in JS-K 1.50 mg?kg-1 group. Compared with the model group, the level of ATP of tumor tissue in JS-K 1.50 mg?kg-1 groups dropped by 16.6%(P<0.01) and the level of LD in JS-K 0.75 and 1.50 mg?kg-1 groups dropped by 38.7%and 59.4%(P<0.01), respectively. In addi-tion, the expression of HIF-1αof tumor tissue in JS-K 1.50 mg?kg-1 group was decreased (P<0.01), and the expression of HKⅡ of tumor tissue in JS-K 0.75 and 1.50 mg?kg-1 groups was decreased signifi-cantly (P<0.05, P<0.01). CONCLUSION JS-K can inhibit the growth of tumor in H22 tumor-bearing mice and its mechanism may be related to regulating the tumor energy metabolism by inhibiting glycolysis and aerobic oxidation.

Nitric oxide donor PABA/NO induced mitochondrial-mediated apoptosis in HepG2 cells / 中国药理学与毒理学杂志

OBJECTIVE To investigate the effects of nitric oxide (NO) donor, O2-{2,4-dinitro-5-[4-(N-methylamino) benzoyloxy]phenyl}1-(N,N-dimethylamino)diazen-1-ium-1,2-diolate (PABA/NO) on apop-tosis in human hepatocarcinoma cells. METHODS Proliferation of HepG2 cells treated with PABA/NO 7.5, 15.0 and 30.0μmol · L-1 was measured with Cell Counting Kit-8 (CCK-8) assay, the morphological features were observed under fluorescence microscopy, the level of NO was measured by DAF-FM DA staining, the apoptosis rate was determined by Annexin Ⅴ-FITC staining, mitochondrial membrane potential was determined by Rhodamine 123 staining, and protein expressions of Bcl-2, Bax, cleaved caspase 3, cytochrome c (Cyt c) and apoptosis inducing factor (AIF) were measured by Western blotting analysis. RESULTS Compared with cell control group, PABA/NO could obviously inhibit the proliferation of HepG2 cells (P<0.01). IC50 value of HepG2 cells treated with PABA/NO for 24 h was (10.8±0.6)μmol·L-1. The cells became round, deformed and appeared shrunken.The level of NO was increased and the fluores-cence intensity was 121 ± 9 (P<0.05), 174 ± 31 (P<0.05) and 230 ± 43 (P<0.01). The apoptosis rate was increased from (2.9 ± 0.5)％ to (17.0 ± 4.5)％ (P<0.01), (39.8 ± 5.4)％ (P<0.01) and (74.3 ± 45.2)％ (P<0.01). The fluorescence intensity of Rh123 was reduced from 668±69 to 605±73, 420±65 (P<0.05) and 242±47 (P<0.01). Compared with cell control group, PABA/NO down-regulated Bcl-2, up-regulated Bax and activated cleaved caspase 3. Meanwhile, the expression of Cyt c in the cytoplasm was increased from 0.15±0.04 to 0.27±0.06 (P<0.05), 0.38±0.07 (P<0.01) and 0.82±0.16 (P<0.01). The expression of AIF in the nucleus was increased from 0.183±0.032 to 0.231±0.011, 0.682±0.020 (P<0.01) and 0.966± 0.090 (P<0.01). Addition of carboxy-PTIO (NO scavenger) 50 μmol · L- 1 blocked PABA/NO-induced down-regulation of Bcl-2, up-regulation of Bax and cleaved caspase 3 activation. Additionally, up-regu-lation of Cyt c in the cytoplasm and up-regulation of AIF in the nucleus were also blocked by carboxy-PTIO in PABA/NO-treated HepG2 cells (P<0.01). CONCLUSION PABA/NO may induce HepG2 cell apoptosis through a mitochondrial pathway.

Regulation of nitric oxide donor JS-K on tumor energy metabolism in H22 tumor-bearing mice / 中国药理学与毒理学杂志

OBJECTIVE To investigate the regulation of {O2 (2,4-dinitrophenyl)1-〔(4-ethoxycarbonyl) piperazin-1-yl〕diazen-1-ium-1,2-diolate}(JS-K), anitric oxide donor, on tumor energy metabolism in H22 tumor- bearing mice. METHODS The hepatoma animal model in BALB/c mice was established with H22 cell line. The JS-K group and model group were received JS-K (0.75 and 1.5 mg?kg-1) and saline via tail intravenous once every 3 d for 14 d, received 5 injections, respectively. The positive group was received 5-FU 20 mg·kg- 1 by intraperitoneal injection once a day for 14 d. On the 15th day mice were sacrificed. The tumor growth inhibition rate were calculated. The activities of hexokinase (HK), phospho?fructo kinase (PFK), pyruvate kinase (PK), succinate dehydrogenase (SDH), adenosine triphosphatase (ATPase), and the levels of lactic acid (LD) and adenosine triphosphate (ATP) in tumor tissues were de?termined by colorimetric method. RESULTS Compared with model group, the tumor mass of JS- K 0.75 and 1.5 mg·kg- 1group was significantly reduced (P<0.01),and the tumor growth inhibition rate was 23.9% and 50.3%, respectively. The activity of HK, PFK, PK, SDH and ATPase of tumor tissue in model group was (22.6±3.7, 14.4±2.6, 12.9±3.2 and 10.5±2.6)U·g-1 protein and (0.70±0.10)μmolPi·mg-1 protein per hour, respectively; which in JS-K 1.5 mg?kg-1 group was dropped by 42.0%, 26.6%, 22.7%, 23.3% and 21.7% (P<0.01, P<0.05). Compared with the model group, the level of ATP and LD in JS-K group was dropped (P<0.01). CONCLUSION JS-K can inhibit the growth of tumor in H22 tumor-bearing mice and its mechanism may be related to regulating the tumor energy metabolism with inhibition of glycolysis and aerobic oxidation.

Synthesis and anti-hepatocellular carcinoma activity of novel O-vinyl diazeniumdiolate-based nitric oxide-releasing derivatives of oleanolic acid / 中国天然药物

Considering that high levels of nitric oxide (NO) exert anti-cancer effect and the derivatives of oleanolic acid (OA) have shown potent anti-cancer activity, new O-vinyl diazeniumdiolate-based NO releasing derivatives (5a-l, 11a-l) of OA were designed, synthesized, and biologically evaluated in the present study. These derivatives could release different amounts of NO in liver cells. Among them, 5d, 5i, 5j, 11g, 11h, and 11j released more NO in SMMC-7721 cells and displayed stronger proliferative inhibition against SMMC-7721 and HepG2 cells than OA and other tested compounds. The most active compound 5j showed almost 20-fold better solubility than OA in aqueous solution, released larger amounts of NO in liver cancer cells than that in normal ones, and exhibited potent anti-hepatocellular carcinoma activity but little effect on the normal liver cells. The inhibitory activity against the cancer cells was significantly diminished upon addition of an NO scavenger, suggesting that NO may contribute, at least in part, to the activity of 5j.

Nitric oxide releasing-dendrimers: an overview

Platforms able to storage, release or scavenge NO in a controlled and specific manner is interesting for biological applications. Among the possible matrices for these purposes, dendrimers are excellent candidates for that. These molecules have been used as drug delivery systems and exhibit interesting properties, like the possibility to perform chemical modifications on dendrimers surface, the capacity of storage high concentrations of compounds of interest in the same molecule and the ability to improve the solubility and the biocompatibility of the compounds bonded to it. This review emphasizes the recent progress in the development and in the biological applications of different NO-releasing dendrimers and the nitric oxide release pathways in these compounds.

Plataformas capazes de armazenar, liberar ou capturar NO de forma controlada e específica são de grande interesse tendo-se em vista aplicações biológicas. Dentre as possíveis matrizes que podem ser utilizadas para esse fim, os dendrímeros são excelentes candidatos. Essas moléculas têm sido empregadas em sistemas para o transporte de fármacos e apresentam propriedades interessantes tais como a possibilidade de modificações químicas em sua superfície, a capacidade de estocar altas concentrações de compostos de interesse em uma só molécula e a possibilidade de aumentar a solubilidade e a biocompatibilidade dos compostos a eles ligados. Esta revisão enfatiza os recentes avanços no desenvolvimento e nas aplicações biológicas de diferentes dendrímeros liberadores de NO e a forma em que o óxido nítrico é liberado nesses compostos.