Effect of α-difluoromethylornithine on cardiac polyamine content and hypertrophy

Cardiac hypertrophy produced by treatment with thyroxine for 7 days was accompanied by a significant increase in polyamine content. Putrescine was elevated by 4- to 5-fold, spermidine by 1.6-fold and spermine 1.3-fold. Administration of α-difluoromethylornithine, a potent irreversible inhibitor of ornithine decarboxylase, prevented these increases provided that a high concentration of the drug was maintained by intraperitoneal injection of 200 mg/kg body wt every 12 h and supplementation of the drinking water with $\text{3 g of the drug}\text{100 ml}$. Such treatment with α-difluoromethylornithine prevented the rise in spermidine and spermine and reduced putrescine content to undetectable levels but did not prevent the increase in heart weight showing that the enhancement of polyamine concentration is not essential for thyroxine-induced cardiac hypertrophy.     

The effects of hypertrophy,hypobaric conditions and diet on myocardial ornithine decarboxylase activity

Myocardial ornithine decarboxylase activity has been measured in the right and left ventricles of rats subjected to hypobaric conditions. Control normobaric groups included both free-fed animals and animals whose diet was restricted to that chosen by the hypobaric group.In normal animals, left ventricular ornithine decarboxylase activity is higher than right ventricular. A diminution in the intake of food causes a decreased myocardial activity of the enzyme. Hypoxia also appears to cause a transient decrease in activity distinct from the effects of anorexia. Hypertrophy of the right ventricle due to hypoxic pulmonary hypertension is associated with an increased ornithine decarboxylase activity in that ventricle.     

Studies of noradrenergic influence on cardiac ornithine decarboxylase in rats.

Ornithine decarboxylase activity in rat heart was increased significantly by the injection of l-norepinephrine or nisoxetine, an inhibitor of norepinephrine uptake. The enzyme was also increased after 3 h of forced exercise. Propranolol, a β-noradrenergic blocking agent, prevented the enzyme increase after norepinephrine, nisoxetine and exercise. Chemical denervation with 6-hydroxydopamine prevented the effect of nisoxetine but not the effect of l-norepinephrine or exercise. Neither propranolol nor 6-hydroxydopamine significantly affected enzyme activity by themselves, suggesting that tonic noradrenergic input is not necessary for maintenance of basal ornithine decarboxylase activity in heart. These results extend previous literature indicating that cardiac ornithine decarboxylase can be elevated by increased noradrenergic input acting via β-receptors.     

Local inhibition of ornithine decarboxylase reduces vascular stenosis in a murine model of carotid injury

Objectives

Polyamines are organic polycations playing an essential role in cell proliferation and differentiation, as well as in cell contractility, migration and apoptosis. These processes are known to contribute to restenosis, a pathophysiological process often occurring in patients submitted to revascularization procedures. We aimed to test the effect of α-difluoromethylornithine (DFMO), an inhibitor of ornithine decarboxylase, on vascular cell pathophysiology in vitro and in a rat model of carotid arteriotomy-induced (re)stenosis.

Methods

The effect of DFMO on primary rat smooth muscle cells (SMCs) and mouse microvascular bEnd.3 endothelial cells (ECs) was evaluated through the analysis of DNA synthesis, polyamine concentration, cell viability, cell cycle phase distribution and by RT-PCR targeting cyclins and genes belonging to the polyamine pathway. The effect of DFMO was then evaluated in arteriotomy-injured rat carotids through the analysis of cell proliferation and apoptosis, RT-PCR and immunohistochemical analysis of differential gene expression.

Results

DFMO showed a differential effect on SMCs and on ECs, with a marked, sustained anti-proliferative effect of DFMO at 3 and 8 days of treatment on SMCs and a less pronounced, late effect on bEnd.3 ECs at 8 days of DFMO treatment. DFMO applied perivascularly in pluronic gel at arteriotomy site reduced subsequent cell proliferation and preserved smooth muscle differentiation without affecting the endothelial coverage. Lumen area in DFMO-treated carotids was 49% greater than in control arteries 4 weeks after injury.

Conclusions

Our data support the key role of polyamines in restenosis and suggest a novel therapeutic approach for this pathophysiological process.     

鸟氨酸脱羧酶抗酶抑制因子1多态性与乙型肝炎肝硬化的关系

目的 研究一种高加索人群丙型肝炎肝硬化风险评分模型(cirrhosis risk score,CRS)相关基因的多态性与中国人群乙型肝炎肝硬化的关系,评估该模型在乙型肝炎肝硬化中的应用价值.方法 用基质辅助激光解析电离飞行时间质谱法检测相关的11个单核苷酸多态性位点,研究人群包括肝硬化组患者和对照组无肝硬化表现者,分析各位点与肝硬化的关系,比较肝硬化组和对照组CRS值的差异.用x2检验检测哈维平衡验证样本的群体代表性.两组计量资料的比较用t检验或Wilcoxon秩和检验.单个单核昔酸多态性基因型等分类数据与肝硬化发生的关系用Pearson x2检验,差异有统计学意义的单核苷酸多态性位点进一步用Logistic回归分析判断在显性模型、隐性模型和加性模型下基因型与肝硬化发生的关系,计算比值比及95%可信区间,比值比经性别和年龄校正.结果 鸟氨酸脱羧酶抗酶抑制因子1基因的rs2679757位点与肝硬化的发生相关(x2=6.79,P＜0.05),与AA纯合子相比,携带危险等位基因G的GG+AG基因型显著增加肝硬化的发生风险,比值比为1.63,95%可信区间为1.13～2.35.位于Toll样受体4基因的rs4986791位点和鸟氨酸脱羧酶抗酶抑制因子1基因的rs62522600位点在中国人群中无多态性基因型.其他研究位点与肝硬化的发生无相关性(P＞0.05).肝硬化组和对照组的CRS中位数分别为0.57和0.62,两组比较,差异无统计学意义(P＞0.05).结论 鸟氨酸脱羧酶抗酶抑制因子1多态性rs2679757与乙型肝炎肝硬化的发生相关.高加索人群CRS模型相关的两个多态性位点在中国人群中缺乏多态性,该CRS对评估有无乙型肝炎肝硬化的发生无明显价值,其他多态性位点仍需进一步临床验证.

Abstract：

Objective A cirrhosis risk score (CRS) comprised of single nucleotide polymorphisms (SNPs) in seven genes that predicts the risk of cirrhosis in Caucasian hepatitis C has been reported. The present study was to evaluate the association of 11 separate but related SNPs and the CRS with cirrhosis risk in Chinese hepatitis B patients. Methods A total of 563 Chinese subjects with persistent HBV infection (349 with evident liver cirrhosis and 214 without cirrhosis clinically or pathologically) were studied. The candidate SNPs were detected with a matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) method. The allele frequency and genotype distribution of each polymorphism as well as the CRS value within the cirrhosis and non-cirrhosis subjects were compared. Results The rs2679757 polymorphism of the antizyme inhibitor 1 (AZIN 1) gene was associated with the risk of cirrhosis ( x 2 = 6.79,P = 0.03, odds ratio for GG+AG versus AA = 1.63, 95% confidence interval = 1.13-2.35). A gene variant (rs886277) in the transient receptor potential cation channel subfamily M, member 5 gene (TRPM5) was associated with liver cirrhosis, but did not reach statistical significance ( x 2 = 5.77, P = 0.06). Two SNPs (rs4986791, rs62522600) are not polymorphic in Chinese. Genotype frequencies of other SNPs were not different between the cirrhosis and non-cirrhosis groups. The overall CRS values were not different between the cirrhotic and non-cirrhotic groups (median value 0.57 versus 0.62, Z = -1.05, P = 0.29). Conclusions SNP rs2679757 in the AZIN1 gene is associated with the risk of HBV-related liver cirrhosis in Chinese. The CRS for Caucasian population has limited applicability for predicting liver cirrhosis in Chinese hepatitis B patients. SNPs associated with cirrhosis prognosis in hepattis B patients and liver diseases with other etiologies warrant further clinical validation.     

Expression of ornithine decarboxylase in precancerous and cancerous gastric lesions

AIM: To investigate the expression of ornithine decarboxylase (ODC) in precancerous and cancerous gastric lesions. METHODS: We studied the expression of ODC in gastric mucosa from patients with chronic superficial gastritis (CSG,n = 32),chronic atrophic gastritis CAG,n = 43; 15 with and 28 without intestinal metaplasia (IM),gastric dysplasia (DYS,n = 11) and gastric cancer (GC,n = 48) tissues using immunohistochemical staining. All 134 biopsy specimens of gastric mucosa were collected by gastroscopy. METHODS: The positive rate of ODC expression was 34.4%,42.9%,73.3%,81.8% and 91.7% in cases with CSG,CAG without IM,CAG with IM,DYS and GC,respectively (P < 0.01),The positive rate of ODC expression increased in the order of CSG < CAG (without IM) < CAG (with IM) < DYS and finally,GC. In addition,ODC positive immunostaining rate was lower in well-differentiated GC than in poorly-differentiated GC (P < 0.05). CONCLUSION: The expression of ODC is positively correlated with the degree of malignity of gastric mucosa and development of gastric lesions. This finding indicates that ODC may be used as a good biomarker in the screening and diagnosis of precancerous lesions.     

Prostanoids, ornithine decarboxylase, and polyamines in primary chemoprevention of familial adenomatous polyposis

BACKGROUND & AIMS: Familial adenomatous polyposis because of germline mutation of the adenomatous polyposis coli gene is characterized by development of colorectal adenomas and, ultimately, colorectal cancer. The usefulness of colorectal mucosal compounds to predict the effect on adenoma development of primary chemoprevention with the nonsteroidal anti-inflammatory drug sulindac was evaluated. METHODS: A randomized, double-blind, placebo-controlled study of 41 subjects genotypically affected with familial adenomatous polyposis but phenotypically unaffected was conducted. Patients received either sulindac or placebo for 48 months, and development of new adenomas was evaluated. The levels of 5 prostanoids, ornithine decarboxylase, and polyamines were measured serially in normal-appearing rectal mucosa. RESULTS: There were no statistically significant differences between treatment groups in baseline levels of prostanoids, ornithine decarboxylase, or polyamines. At conclusion of the study, 4 of 5 prostaglandin levels were statistically significantly lower in the sulindac group than in the placebo group. Among the subset of patients taking sulindac, 3 of 5 prostaglandin levels were statistically significantly lower in patients who were polyp free than in those who developed polyps. By contrast, there were no statistically significant differences in ornithine decarboxylase or polyamines between treatment groups or in those on sulindac who were polyp free compared with those who developed polyps. CONCLUSIONS: Colorectal mucosal prostaglandin levels, but not ornithine decarboxylase or polyamines, may be valuable biomarkers to assess appropriate drug dosage and medication compliance in patients undergoing primary chemoprevention therapy with sulindac. Reduction of mucosal prostaglandin levels may be necessary to achieve chemopreventive benefit from this agent.     

心肌肥厚的PTEN负性调控与卡托普利干预对其影响的实验研究

目的检测异丙肾上腺素(isoproterenol,ISO)诱导的心肌肥厚大鼠PTEN mRNA、蛋白水平表达及卡托普利(captopril,Cap)对其表达的影响,从而探讨PTEN的负性调控在心肌肥厚中的作用。方法24只大鼠随机分为对照组、ISO组、Cap+ISO组。利用小剂量ISO持续背部皮下注射大鼠,建立心肌肥厚模型。在观察期末,分别测定各组大鼠体重、心脏湿重、左室湿重,计算出心脏重量/体重及左室重量/体重;电镜观察超微结构的变化,并测定左室收缩末压、左室舒张末压、左心室压力上升及下降最大速率等指标。RT-PCR测定心肌组织PTEN mRNA,Western blot测定其蛋白表达。结果(1)与对照组比较,ISO组、Cap+ISO组的左室重量/体重、心脏重量/体重、左室收缩末压、左室舒张末压均升高(P≤0.05),左室压力上升及下降最大速率(±dp/dtmax)均下降(P≤0.05)。(2)与ISO组相比,Cap+ISO组的左室重量/体重、心脏重量/体重、左室收缩末压、左室舒张末压均下降(P≤0.05,P≤0.01),左室压力上升及下降最大速率(±dp/dtmax)均升高(P≤0.05,P≤0.01)。(3)与对照组比较,ISO组、Cap+ISO组的PTEN mRNA、蛋白表达均增加。(4)与ISO组比较,Cap+ISO组的PTEN mRNA、蛋白表达增加。结论ISO诱导心肌肥厚PTENmRNA、蛋白表达升高,心肌肥厚过程中存在负性调控,PTEN是一种内源性抑制心肌肥厚的重要因子。卡托普利不仅能明显抑制心肌肥厚,改善血液动力学参数,而且还能上调心肌PTEN水平,这是其抑制心肌肥厚的又一机制。     

Involvement of polyamines in apoptosis of cardiac myoblasts in a model of simulated ischemia

Apoptotic cell death of cardiomyocytes is involved in several cardiovascular diseases including ischemia, hypertrophy, and heart failure. The polyamines putrescine, spermidine, and spermine are polycations absolutely required for cell growth and division. However, increasing evidence indicates that polyamines, cell growth, and cell death can be tightly connected. In this paper, we have studied the involvement of polyamines in apoptosis of H9c2 cardiomyoblasts in a model of simulated ischemia. H9c2 cells were exposed to a condition of simulated ischemia, consisting of hypoxia plus serum deprivation, that induces apoptosis. The activity of ornithine decarboxylase, the rate limiting enzyme of polyamine biosynthesis that synthesizes putrescine, is rapidly and transiently induced in ischemic cells, reaching a maximum after 3 h, and leading to increased polyamine levels. Pharmacological inhibition of ornithine decarboxylase by alpha-difluoromethylornithine (DFMO) depletes H9c2 cardiomyoblasts of polyamines and protects the cells against ischemia-induced apoptosis. DFMO inhibits several of the molecular events of apoptosis that follow simulated ischemia, such as the release of cytochrome c from mitochondria, caspase activation, downregulation of Bcl-xL, and DNA fragmentation. The protective effect of DFMO is lost when exogenous putrescine is provided to the cells, indicating a specific role of polyamine synthesis in the development of apoptosis in this model of simulated ischemia. In cardiomyocytes obtained from transgenic mice overexpressing ornithine decarboxylase in the heart, caspase activation is dramatically increased following induction of apoptosis, with respect to cardiomyocytes from control mice, confirming a proapoptotic effect of polyamines. It is presented for the first time evidence of the involvement of polyamines in apoptosis of ischemic cardiac cells and the beneficial effect of DFMO treatment. In conclusion, this finding may suggest novel pharmacological approaches for the protection of cardiomyocytes injury caused by ischemia.     

低分子肝素对大鼠心肌肥厚纤维化保护作用研究

目的探讨低分子肝素(LMWH)对大鼠心肌肥厚和纤维化的影响及作用机制。方法异丙肾上腺素(ISO)皮下注射致大鼠心肌肥厚纤维化,并观察经LMWH(200 U·kg-1·d-1)治疗10 d后,对大鼠的心脏质量指数、组织形态、血清中一氧化氮(NO)、心肌组织中胶原水平、钙调神经磷酸酶(CaN)活性的影响。结果与对照组比较,损伤组心脏质量指数增加(P<0．01),出现纤维化病变,心肌中的CaN活性增高(P<0．01),胶原水平增加(P<0．01),血清中NO水平减少(P<0．01);而LMWH保护组与损伤组相比,心肌肥厚和纤维化病变减轻,心肌中的胶原水平减少(P<0．01),CaN活性降低(P<0．01),血清中的NO水平显著增高(P<0．01)。结论 LMWH有良好的减轻心肌肥厚和抗心脏纤维化作用,该作用可能与其抑制CaN活性、减少胶原沉积和增加NO水平有关。     

异丙肾上腺素致大鼠心肌损伤肥大时骨架蛋白的变化及意义

目的复制异丙肾上腺素(isoproteronol,Iso)致大鼠心肌损伤及肥大模型,观察细胞骨架蛋白微管蛋白(tubulin)、结蛋白(desmin)变化特点,探讨心肌支持蛋白在心肌肥厚过程中的关系和变化规律,为抑制心室肥厚的发生和发展提供实验依据。方法①将雄性Wistar大白鼠随机分成对照组和损伤肥大组(Iso组)。Iso组皮下多点注射中等剂量Iso,建立大鼠损伤、代偿性肥厚模型。②光镜观察心肌组织的病理变化。③RT-PCR法检测tubulin、desminmRAN表达。④免疫组化法检测tubulin、desmin蛋白表达。结果光镜下Iso组心肌组织有明显的损伤肥大及纤维化,tubulinmRNA(0.9252±0.3765)和desminmRNA(1.2453±0.5326)表达与对照组(0.6846±0.2372)和(0.7142±0.1416)比较明显升高,差异有统计学意义(t=2.3258,P<0.05;t=4.0972,P<0.01)。Iso组tubulin(0.75±0.32)和desmin(0.85±0.29)蛋白表达与对照组(0.56±0.25)和(0.64±0.31)比较,差异也有统计学意义(t=2.5627,P<0.05;t=2.7544,P<0.01)。结论Iso致大鼠心肌损伤肥大时,tubulin、desmin无论在蛋白水平,还是在基因水平都增加,提示异丙基肾上腺素对大鼠心肌tubulin、desmin蛋白水平和基因水平的影响是平行、一致的。     

Inhibition of ornithine decarboxylase induction by retinobenzoic acids in relation to their binding affinities to cellular retinoid-binding proteins

Summary Retinobenzoic acids induce differentiation of human promyelocytic leukemia cells (HL-60). Like retinoic acid, 14 retinobenzoic acids inhibited the induction of ornithine decarboxylase (ODC) by teleocidin in mouse skin. The mechanism(s) of inhibition of ODC induction by 7 retinobenzoic acids, Am 80, Am 81, Am 580, Am 590, Am 68, Sa 80, and Ch 55 was compared with those by all-trans-retinoic acid and the arotinoid compound 19. Application of 114 nmol of Am 80, Am 81, Am 580, Am 590, Am 68, Sa 80, or Ch 55, 10 min before 11.4 nmol of teleocidin, resulted in 76.7%, 82.0%, 76.2%, 28.3%, 48.4%, 58.6%, and 85.1% inhibition of ODC induction, respectively. Since all-trans-retinoic acid and compound 19 were also inhibitory, we determined whether retinobenzoic acids bind to cellular retinoic acid-binding protein (CRABP) isolated from bovine adrenal glands. Am 80 and Am 580 inhibited the specific binding of ³H-retinoic acid to CRABP, but also showed less affinity than authentic unlabeled retinoic acid and compound 19. Am 81, Am 590, Am 68, Sa 80, and Ch 55 at up to 10 M were not effective competitors of the binding of either ³H-retinoic acid or ³H-retinol. These results suggest that the inhibition of ODC induction can be mediated by pathways that do not involve CRABP or the cellular retinol-binding protein.     

心肌特异性Hsp27对ISO诱导小鼠心肌肥厚的保护作用

目的探讨人热休克蛋白(Hsp27)基因产物对异丙肾上腺素(ISO)诱导的小鼠心肌肥厚的影响。方法以本研究室建立的心肌特异性表达Hsp27转基因(TG)小鼠为模型,腹腔注射ISO[30mg/(kg·d)]共7d。实验在规定时间结束后,测定心脏质量与胫骨长度的比值(HW/TL)、心脏二维超声(echo),Masson三色染色观察心肌纤维化改变,各组均以野生型鼠(WT)为对照组。结果(1)ISO处理使TG鼠和WT鼠的HW/TL与生理盐水处理对照组比较分别增加了7·46%和17·65%,WT组增加比率有统计学差异(P<0·01);(2)echo显示ISO使TG心脏收缩期后壁厚度增加(P<0·05),而WT组心脏前、后壁在收缩、舒张末期均有显著的增加(P<0·05~0·01)。(3)每搏输出量、短轴缩短率、心脏射血分数没有统计学差异。WT鼠心肌纤维化明显多于TG鼠。结论Hsp27可显著抑制ISO诱导的小鼠早期心肌重构。其机制有待进一步探讨。     

食管鳞癌中鸟氨酸脱羧酶mRNA、内皮抑素mRNA表达和血管生成相关性研究

目的 研究鸟氨酸脱羧酶(0DC)mRNA、内皮抑素mRNA和微血管密度(MVD)在食管鳞癌中的表达及相关关系，探讨0Dc在食管鳞癌血管生成中的可能作用，及其与肿瘤浸润和转移的关系。方法 用半定量RT-PcR法测定41例食管鳞癌患者痛组织(T)和相应癌旁组织(N)中的ODC mRNA和内皮抑素mRNA表达，求得各自T／N值；同时用免疫组化方法测定癌组织和癌旁正常组织MVD的T／N值。结果在41例患者中，39例ODC mRNA的T／N值＞1．0，占95．1％：40例MVD的T／N值＞1．0，占97．6％。36例内皮抑素mRNA的T／N值＞1．0，占87．8％。ODC mRNA、内皮抑素mRNA和MVD与食管鳞癌的肿瘤大小、浸润深度和淋巴结转移有关，ODC mRNA、内皮抑素mRNA表达与肿瘤分化程度有关。ODC mRNA的T／N值与MVD的T／N值呈正相关，与内皮抑素mRNA的T／N值呈负相关，内皮抑素mRNA的T／N值与MVD的T／N值呈负相关。结论 DOC与食管鳞癌血管生成和肿瘤浸润转移密切相关。DOC过表达可能是通过抑制内皮抑素而促进血管生成．从而促进食管鳞癌的浸润和转移。     

Effect of remedies for enhancing resistance and relieving blood stasis on metastasis in postoperative gastric cancer and ornithine decarboxylase levels

AIM: To study the action of remedies for enhancing resistance and relieving blood stasis on metastasis in postoperative gastric cancer and its influence on ornithine decarboxylase (ODC).METHODS: Sixty-three postoperative patients with gastric cancer were randomly divided into two groups. Thirty-one patients were treated with western medicine consisting of the FAP (5-fluorouracil, adriamycin, cisplatin) and CODP regimens (cyclophosphamide, vincristine, daunorubicin, prednisone), whereas 32 patients were treated with the FAP regimen and traditional Chinese medicine. Correlations were made between the ODC levels detected before and after treatment and other factors such as tumor diameter, infiltration depth, histological type, and lymph node metastasis.RESULTS: The ODC levels in the gastric cancer tissue and adjacent normal gastric mucosal tissue were significantly higher in the patients than in the controls. There was an obvious correlation between increased ODC and tumor size, infiltration depth, degree of differentiation, and lymph node metastasis. Six months later, there were no significant changes in the ODC levels of the group using only Western medicine, while the ODC levels decreased markedly in the group using combined Western and traditional Chinese medicine (P < 0.01).CONCLUSION: The effects of traditional Chinese medicine remedies on metastases in postoperative gastric cancer are related to the reduction of ODC activity.     

alpha-Difluoromethylornithine, an irreversible inhibitor of ornithine decarboxylase, inhibits tumor promoter-induced polyamine accumulation and carcinogenesis in mouse skin.

The role of ornithine decarboxylase (OrnDCase, EC 4.1.1.17) and of the polyamines [putrescine (Put), spermidine (Spd), and spermine (Spm)] in mouse skin tumor promotion was investigated by the use of alpha-difluoromethylornithine (CHF2-Orn), an enzyme-activated irreversible inhibitor of OrnDCase. 12-O-Tetradecanoylphorbol 13-acetate (TPA), mezerein, and ethyl phenylpropiolate (EPP) were employed as complete, stage II specific, and nonpromoting agents, respectively. TPA and mezerein, but not EPP, provided for a dose-dependent increase in tissue Put accumulation. The Put level in papillomas developed by TPA (2 micrograms) treatment was approximately equal to 15-fold higher than that of the surrounding skin tissue; Spd accumulation was 2- to 3-fold greater in the papillomas. Put administered (intraperitoneally) with TPA greatly enhanced papilloma yield. CHF2-Orn, given orally or intraperitoneally, abolished the TPA-induced OrnDCase activity and Put accumulation in mouse epidermis. The reduction of polyamine accumulation by CHF2-Orn was directly proportional to reduction of tumor size. CHF2-Orn administered in a two-stage (TPA-mezerein) promotion protocol [Slaga, T. J., Fischer, S. M., Nelson, K. G. & Gleason, G. L. (1980) Proc. Natl. Acad. Sci. USA 77, 3659-3663; Slaga, T. J., Klein-Szanto, A. J. P., Fischer, S. M., Weeks, C. E., Nelson, K. & Major, S. (1980) Proc. Natl. Acad. Sci. USA 77, 2251-2254] reduced tumor size, inhibited by 65-70% the number of papillomas per mouse, and decreased by 40% the percentage of mice with tumors when given with the stage II agent mezerein. CHF2-Orn provided considerably less effect on tumorigenesis when administered with the TPA portion of the protocol, and CHF2-Orn did not inhibit the induction of dark basal keratinocytes by TPA. Based on our results with CHF2-Orn, we suggest that regulation of polyamine biosynthesis, particularly Put, is a critical factor in stage II promotion.     

Ornithine decarboxylase activity in the isolated perfused rat heart.

Ornithine decarboxylase activity in rat hearts perfused by the Langendorff technique decreased by 50% after 1 h. Pyridoxal-5-phosphate, amino acids, cycloheximide, isoprenaline and insulin, when added to the perfusion medium, did not significantly affect myocardial ornithine decarboxylase activity. Growth hormone, T₃ and dibutyryl 3′,5′-cAMP were each able to prevent a loss of enzyme activity in hearts perfused for 1 h. When both T₃ and cycloheximide were added to the perfusion medium the ornithine decarboxylase activity was similar to that of hearts perfused with a medium containing T₃ alone. The results suggested that the biosynthesis of ornithine decarboxylase stopped with the onset of perfusion. The elevated enzyme activity after perfusion with growth hormone, T₃ or dibutyryl 3′,5′-cAMP was probably due to a decreased degradation of the enzyme protein.     

Regulatory effects of corticosterone on ornithine decarboxylase activity during liver regeneration in rats

AIMS: The regulation of ornithine decarboxylase (ODC) gene expression and enzyme activity by corticosterone during rat liver regeneration induced by partial hepatectomy (PH) was evaluated. METHODS: Bilateral adrenalectomies were performed on ether-anesthetized rats 3 days before PH. Corticosterone in sesame oil was injected subcutaneously to adrenalectomized rats. ODC mRNA, ODC protein and enzyme activity were detected by in situ hybridization, Western blot and high performance liquid chromatography (HPLC), respectively. RESULTS: The ODC mRNA levels, protein accumulation and enzyme activity were lower in the intact liver compared to the regenerating liver. After PH, mRNA levels were remarkably enhanced in all groups and peaked at 5 h post-PH, and presented a persistent increase only in adrenalectomy rats during the regeneration process. Corticosterone treatment resulted in a dose-dependent decrease in ODC mRNA content after 5 h post-PH. ODC protein accumulation in adrenalectomy rats was higher than that in sham-adrenalectomy rats, but it decreased in corticosterone-treated (10 mg/kg) rats until 24 h post-PH, with a strong decline seen in 40 mg/kg corticosterone-treated rats. ODC activity was rapidly promoted, and the highest levels were observed at 6 h after PH in all groups. After corticosterone treatment, the activities declined significantly at 6 h post-PH, with the lowest value found in the 40 mg/kg group. CONCLUSIONS: Corticosterone treatment results in dose-dependent decreases in ODC mRNA and enzyme protein both in the intact liver and the regenerating liver. The change in ODC activity is partially related to alterations of ODC mRNA and protein accumulation.     

Influence of gastrin,gastrin receptor blockers,epidermal growth factor,and difluoromethylornithine on the growth and the activity of ornithine decarboxylase of colonic carcinoma cells

Summary Polyamines are essential factors of cell growth and differentiation. Modulation of the cellular polyamine content by 2-difluoromethylornithine (DFMO) inhibiting ornithine decarboxylase (ODC), or by hormones inducing ODC, influences cell growth. Gastrin acts trophically on some colonic carcinomas and their growth is inhibited by gastrin receptor blockers. The mechanism of the trophic action of gastrin on colonic carcinomas is not known. In this study the effect of gastrin, gastrin receptor blockers, epidermal growth factor (EGF) and DFMO on growth and ODC activity of four human colon carcinoma cell lines (SW 403, SW 1116, LS 174 T and Lovo) was investigated. Growth and ODC activity of all cell lines were inhibited by DFMO. Growth of the SW 403 cell line was increased by gastrin and inhibited by the gastrin receptor blocker benzotrypte. The other cell lines did not respond to gastrin and the gastrin receptor blocker. In SW 403 cells ODC activity was increased by gastrin, and was also elevated after treatment with the gastrin receptor blocker. These in vitro results were confirmed by studies on tumours that developed from SW 403 cells in nude mice. Combination of benzotrypte and DFMO did not enhance the antiproliferative effect. EGF increased growth of SW 403 cells, but no induction of ODC activity was measured. LS 174 T cells were not stimulated by EGF. Medium replacement was the strongest stimulus of ODC activity in SW 403 cells already inducing ODC after 3 h. During cell culture ODC activity was high after seeding and decreased continuously with increasing cell density. These data suggest that gastrin induces ODC in gastrin-sensitive colonic carcinoma cells. DFMO appears to be a valuable antiproliferative agent in colonic carcinoma cells.Abbreviations ODC Ornithine decarboxylase - DFMO 2-difluoromethylornithine - EGF epidermal growth factor Supported by Deutsche Forschungsgemeinschaft (DFG)     

Effects of tobacco-specific nitrosamines and snuff extract on cell proliferation and activities of ornithine decarboxylase and aryl hydrocarbon hydroxylase in mouse tongue primary epithelial cell cultures

Summary Tobacco and its related compounds, including snuff, have been implicated in oral cancers. Tobacco-specific nitrosamines have been shown to be the causative agents present in tobacco and its related compounds. Both, N-nitrosonornicotine (NNN) and its butanone derivative (NNK) are carcinogenic in animals. In our in vitro studies using embryonic mouse tongue epithelial cells, NNN is linked to an increase in [³H]dT uptake along with a concomitant increase in ornithine decarboxylase and aryl hydrocarbon hydroxylase activities. NNK, the more potent compared to NNN, causes a further increase in [³H]dT uptake, cell count and ornithine decarboxylase activity. However, aryl hydrocarbon hydroxylase behaves differently in cultures treated with NNK compared to those treated with NNN. Snuff extract has an overall inhibitory effect on cell count, [³H]dT uptake, and ornithine decarboxylase and aryl hydrocarbon hydroxylase activities when administered either alone or in combination with NNN and NNK. How the inhibitory effect of snuff in the presence of tobacco-specific nitrosamines is involved in oral carcinogenesis should be further investigated.Abbreviations used NNN N-nitrosonornicotine - NNK 4-(nitrosomethylamino)-1-(3-pyridyl)-1-butanone