Effect of siRNA targeting HER2/neu on the proliferation and viability of prostate cancer PC-3M cells.

The aim of this study was to investigate the effect of a small interfering RNA (siRNA) targeting human epidermal growth factor receptor 2 (HER2/neu) on the proliferation and viability of prostate cancer PC-3M cells. Chemically synthesized siRNA targeting HER2/neu was transfected into PC-3M cells by using liposomes, and cells transfected with empty liposomes, a negative siRNA sequence, or nothing (untransfected) were used as controls. mRNA and protein levels of HER2/neu were detected using reverse transcription-polymerase chain reaction and western blot, respectively. The inhibitory action of HER2/neu siRNA on the in vitro growth of PC-3M cells was assessed by the cholecystokinin 8 assay and apoptosis was detected using flow cytometry. Cells transfected with HER2/neu siRNA showed decreased mRNA and protein levels of HER2/neu compared to control groups (P < 0.05). The survival rate of PC-3M cells decreased significantly after transfection with HER2/neu siRNA compared to that of untransfected cells (55.39 ± 1.60 and 81.42 ± 0.80%, respectively; P < 0.05). The apoptosis rate in cells transfected with HER2/neu siRNA was quite high (45.60 ± 0.70%) compared to that of blank control, empty liposome, and negative siRNA sequence groups (P < 0.05). In conclusion, siRNA targeting HER2/neu inhibits HER2/neu expression in PC-3M cells, resulting in an inhibition in proliferation and an induction of apoptosis.

Interactive toxicity of simple chemical mixtures of cadmium, mercury, methylmercury and trimethyltin: model-dependent responses.

Scientific and societal interest in the analysis of aggregate toxicity derives from the fact that people are seldom exposed to single chemicals, but rather to multiple agents from different sources and even to mixtures of agents from a single source. Many descriptive terms and mathematical, graphical, and statistical models have been used to evaluate the toxicity of simple mixtures. It is not very easy to distinguish clearly the intrinsic differences, distinctions and limitations of these models when applied to characterizing interactive toxicity. A series of experiments were performed to illustrate model-dependent consistencies and differences in interactive toxicity. Cultured murine renal cortical cells, target cells for metal toxicity, were treated with selected concentrations of one metal or binary mixtures of metals to give conditions of dose-additivity, response additivity, or with only one toxic member of the binary mixture. The cytotoxicity was determined at 24h by lactate dehydrogenase release. The data were analyzed graphically and mathematically by (a) Carter's statistical isobologram, (b) Barton's non-linear, and (c) Kodell and Pounds' linear models to characterize the interaction. These models were compared and contrasted for robustness, and consistency using these common data sets. The models gave generally consistent conclusions, but each model has limitations and strengths for assessing particular mixtures scenarios. This comparison illustrates the complexity of extrapolating conclusions between models, and difficulty of public health assessment from exposures to multiple chemicals in the environment.

Inhibitory effects of captopril on c-myc expression during left ventricular hypertrophy.

AIM: To study the molecular mechanism of captopril (Cap) on the inhibition of left ventricular hypertrophy (LVH), disclose the expression and distribution of c-myc in different cell types in left ventricle (LV) in spontaneously hypertensive rats (SHR). METHODS: Cap 100 mg.kg-1.d-1 was given p.o. to SHR. Systolic blood pressure (SBP), left ventricular weight (LVW), and body weight (BW) were measured at 16-wk old. The level of angiotensin II (Ang II), c-myc mRNA, and oncoprotein were determined by immunohistochemical method, Northern blot, and Western blot, respectively. RESULTS: Cap reduced SBP, LVW/BW in SHR, with a decrease of Ang II and c-myc expression in LV. Local cardial Ang II mainly distributed in cardiomyocytes. Cap inhibited cardial Ang II production and c-myc expression (histochemical staining intensity index, 0.49 +/- 0.04 vs 0.83 +/- 0.24, P < 0.01). The c-myc oncoprotein was prevailingly located in cardiac fibroblasts. The c-myc oncoprotein in Cap treated SHR was lower than that of WKY. CONCLUSION: High expression of c-myc in fibroblasts played an important role in the development of LVH in SHR. Inhibitory effects of Cap on LVH was associated with a decreased myocardial Ang II and interstitial fibroblasts c-myc expression. The c-myc oncoprotein post-transcriptional translation was also interrupted by Cap.

Inhibition of left ventricular hypertrophy and expression of proto-oncogenes c-myc other than c-fos in myocardium by early captopril treatment in SHR rats.

AIM: To explore the mechanisms by which angiotensin converting enzyme inhibitor (ACEI) prevents the development of left ventricular hypertrophy (LVH). METHODS: Captopril (Cap 100 mg.kg(-1).d(-1)) was given orally to male spontaneously hypertensive rats from intrauterine period to 16 wk of age. Experiments were performed at 40 wk of age. SBP, left ventricular weight to body weight ratio (LVW/BW) were assessed. The levels of c-myc and c-fos mRNA in the left ventricle were measured by Northern blot. RESULTS: Early-onset Cap therapy significantly decreased SBP. After discontinuance of treatment for 24 wk, SBP of SHRcap was still maintained at a lower level. LVW/BW in SHRcap was markedly reduced. The expression of myocardial c-myc mRNA was decreased by 72% in SHRcap compared with that in the untreated SHR, but the expression of myocardial c-fos mRNA was not different between the untreated SHR, SHRcap, and WKY rats. CONCLUSION: Early Cap treatment may permanently prevent the development of hypertension, inhibit LVH. Furthermore, the prevention of LVH is associated with a decrease in c-myc mRNA levels, and the development and regression of left ventricular hypertrophy may be irrelevant to c-fos expression.

[Mechanism of inhibition in left ventricular hypertrophy by captopril treatment in spontaneously hypertensive rats].

To explore the mechanisms by which angiotensin converting enzyme inhibitor (ACEI) prevents the development of left ventricular hypertrophy (LVH), captopril (Cap 100 mg.kg-1/d was administered orally to male spontaneously hypertensive rats from intrauterine period to 16 weeks of age. Male and age-matched untreated WKY rats and SHR were used as controls. Experiments were performed at 40 weeks of age. SBP, left ventricular weight to body weight ratio (LVW/BW), myocardial hydroxyproline (Hypro) and norepinephrine (NE) were determined. The levels of c-myc and c-fos mRNA in the left ventricle were measured by Northern blot. Early-onset Cap therapy significantly decreased SBP at 16 weeks of age. After discontinuance of treatment for 24 weeks, SBP of SHRcap was still maintained at a level lower than that of untreated SHR. LVW/BW and Hypro in SHR cap were markedly reduced. The expression of myocardial c-myc mRNA (n = 5) was decreased by 72% in SHRcap compared with that in the untreated SHR, but the expression of c-fos mRNA (n = 7) and NE was not different between the untreated SHR, SHRcap and WKY rats. These results indicate that early Cap treatment may permanently prevent the development of hypertension, inhibit myocardial hypertrophy (MH), and interstitial fibrosis. Furthermore, the prevention of MH is associated with a decrease in myocardial c-myc mRNA levels, and the development and regression of MH may be irrelevant to proto-oncogene c-fos expression.

[Effects of clonidine on estrus, estradiol, gonadotropin, graafian follicle, and corpus luteum in rats].

Effects of clonidine (Clo) on female reproductive system were studied in rats. Blood FSH, LH, progesterone, testosterone, and estradiol were measured by radioimmunoassay and the development of secondary follicles and corpus luteum in ovary were investigated by morphometry. After Clo po 0.3 mg.kg-1.d-1 x 14 d, the estrus of rats was prolonged; FSH, LH, and progesterone increased significantly; while estradiol reduced. The development of secondary follicles in ovary was blocked at the stage of prematuration and the numerical density of corpus luteum decreased. After clonidine po 28 d, FSH and LH sustained at high levels, but the estrous cycle, estradiol and progesterone recovered.

A point mutation in the MyoD basic domain imparts c-Myc-like properties.

MyoD and c-Myc, members of the large "basic-helix-loop-helix" family of proteins, regulate diverse aspects of both normal and neoplastic growth and specific gene regulation. These two proteins differ at 9 of the 14 amino acids that comprise the basic domains necessary for DNA binding and transcriptional control. Individual amino acids in the MyoD basic domain were mutated to those found at the analogous positions in c-Myc. Four classes of mutants were obtained: (i) those with no effects on MyoD-site binding or activation of MyoD-responsive genes, (ii) those with no effect on MyoD-site binding but with a loss of activation potential, (iii) those with a loss of both DNA binding and activation potential, and (iv) one mutant (mut 9, Leu122----Arg) that left MyoD-site binding unaffected but imparted a new c-Myc-site binding capability. mut 9 competed with wild-type protein for the activation of MyoD-responsive reporter genes but could, like c-Myc, also suppress the adenovirus major-late promoter, which contains a c-Myc binding site. Our studies thus identify specific amino acid residues in the MyoD basic domain that are important for its activity as a DNA-binding transcriptional activator. Most significantly, our results with mut 9 indicate that Leu122 of MyoD is a critical determinant of specific DNA binding and that mutation at this residue can alter this specificity.

Effects of atrial natriuretic peptide in the canine coronary circulation.

Atriopeptin II has been reported to cause profound coronary vasoconstriction in the isolated perfused guinea pig heart and in the blood perfused canine heart. Consequently, this study was carried out to examine possible mechanisms by which vasomotor effects of human atrial natriuretic peptide (ANP) occur in the canine coronary circulation. Bolus dosages of ANP were administered into the left circumflex coronary artery of in situ dog hearts perfused at constant flow rate. ANP produced dose-related coronary vasodilation with a threshold dosage of 2 ng/kg; a dosage of 2 micrograms/kg caused a 27 +/- 4% decrease in coronary vascular resistance. Coronary vasodilation produced by ANP was not altered by beta-adrenergic blockade with propranolol (1 mg/kg i.v.). In addition, neither adenosine receptor blockade with 8-phenyltheophylline (5 mg/kg i.v.) nor cyclooxygenase inhibition with indomethacin (5 mg/kg i.v.) significantly altered the response to intra-arterial ANP. These data demonstrate that in the in vitro blood perfused canine heart, ANP administered intra-arterially results in coronary vasodilation that does not utilize adenosine-dependent or prostaglandin-dependent mechanisms.