Effect of several PEO-PPO amphiphiles on bax, bcl-2, and hTERT mRNAs: An insight into apoptosis and cell immortalization induced in hepatoma cells by these polymeric excipients.

UNLABELLED: Recent data have shown that synthetic polymers and nanomaterials display phenotypic effects in cells and signal transduction mechanisms involved in inflammation, differentiation, proliferation, and apoptosis. AIM: This article aims to investigate the effect of poly(ethylene oxide)-poly(propylene oxide) (PEO-PPO) block copolymers with a wide range of biomedical and pharmaceutical applications on apoptosis and/or cell immortalization, by flow cytometry and multiplex RT-PCR for bax, bcl-2, and human telomerase reverse transcriptase (hTERT). RESULTS: PEO-PPO amphiphiles upregulated bax and hTERT and induced apoptosis of two human hepatoma cell lines. CONCLUSIONS: PEO-PPO block copolymers-considered safe for human use-can drastically alter gene expression profiles of genes related to apoptosis/cell proliferation.

[Isolation of fetal liver oval cells in physiologic conditions form their natural niche]. / Aislamiento de células ovales de hígado fetal en condiciones fisiológicas a partir de su nicho natural.

The liver is characterized by a remarkable ability to proliferate and self-renew. In the situation of mild or moderate liver damage, hepatocytes carry out regeneration. Nevertheless, when liver damage is far too much extensive and the number of residual mature hepatocytes is not enough to accomplish regeneration, or likewise when mature hepatocyte proliferation is inhibited, hepatic regeneration depends on the activation of liver stem cells that give rise to oval cells. The population of liver stem cells is scant in normal liver. It is considered that in fetal liver this population is just over 1% of the cells. For this reason, it is necessary to isolate and enrich them for their study. With this goal several models of hepatic damage that permit the isolation of oval cells af ter the induction of massive hepatic injure have been developed. Here we present a simple methodology that allows the isolation of oval cells from rat fetal liver without prior induction of liver damage. The use of oval cell 2 (OC2) and oval cell 3 (OC3) antigens as molecular markers allowed the highly precise characterization of this cell population. Furthermore, the in vitro culture in presence of HGF yielded a substantial enrichment of the oval cell population.

Expression of caveolin-1 in penile cavernosal tissue in a denervated animal model after treatment with sildenafil citrate.

INTRODUCTION: Radical pelvic surgery is a major cause of erectile dysfunction due to iatrogenic cavernous nerve damage. Endothelial nitric oxide synthase, which generates nitric oxide (NO) in the cavernosal tissues, localizes to specialized plasma membrane invaginations known as caveolae. Growing evidence suggests that caveolae are major components of signal trafficking and that stimuli that affect the concentration of the main structural protein of caveolae, caveolin-1 influence NO signaling. AIM: To evaluate caveolin-1 expression as a marker of cavernous tissue damage and determine the impact of early sildenafil administration on caveolin-1 expression in animal models of partial and total surgical penile denervation. METHODS: Thirty-six rats were divided into six groups (N = 6 per group) that received bilateral or unilateral penile denervation or sham surgery, with and without sildenafil 10 mg daily for 7 weeks. MAIN OUTCOME MEASURES: Sections were taken from the proximal middle portion of the penis of all animals. Cavernous tissue was delineated by the tunica albuginea, then the extent of immunostaining for the following parameters was quantitated to determine (i) cavernous smooth muscle layer in the cavernous space expressed as the percentage of alpha-smooth muscle actin (alpha-SMA) positive immunostaining per area and (ii) caveolin-1 expressed as a percentage of area. RESULTS: A marked decrease in both caveolin-1 and alpha-SMA expression in cavernous smooth muscle tissue and in the endothelium of rats was noted after a bilateral and unilateral neurotomy. Specimens from animals receiving sildenafil exhibited higher mean immunostaining values for both proteins in cavernous tissue. The differences were statistically significant compared with groups receiving the same surgical treatment without sildenafil. CONCLUSION: Caveolin-1 and alpha-SMA expression in cavernous tissue is significantly reduced by pelvic nerve injury, and the loss is related to the extent of the neural damage. Early administration of sildenafil elicits caveolin-1 expression, which appears to preserve cavernous tissue.

F127 poloxamer effect on cytotoxicity induction of tumour cell cultures treated with doxorubicin.

INTRODUCTION: Hepatocellular carcinoma is the most common liver malignancy and the third leading cause of cancer death worldwide. One crucial limitation in the pharmacotherapy for this tumour is its chemotherapy-resistant nature produced by the overexpression of several members of the ATP-binding cassette protein family that efflux drugs out of cells, as observed with the breast cancer resistant protein (BCRP). OBJECTIVES: This study aimed to assess the ability of Pluronic® F127 to reverse the multidrug resistance phenotype in two human hepatocellular cell lines. METHODS: PLC/PRF/5 and SKHep1 cells were exposed to Pluronic® F127 at several concentrations. The effect of F127 on BCRP expression (mRNA and protein), mitochondrial transmembrane potential and cell hypodiploidy was assessed. Finally, the effect of this copolymer on cytotoxicity of doxorubicin in both hepatoma cell lines was investigated, as expressed by its reverse resistance index. KEY FINDINGS: It was demonstrated that F127 in both cell lines contributes to chemosensitization, as shown by BCRP down-regulation, an altered mitochondrial transmembrane potential and hypodiploidy and reverse resistance index values. A remarkable dependence of these effects significantly correlated with the copolymer concentration. CONCLUSIONS: These findings further uncover the potential usefulness of this copolymer as multidrug resistance reversal agent, increasing the efficacy of cancer therapies.

Homeostatic response under carcinogen withdrawal, heme oxygenase 1 expression and cell cycle association.

BACKGROUND: Chronic injury deregulates cellular homeostasis and induces a number of alterations leading to disruption of cellular processes such as cell cycle checkpoints and apoptosis, driving to carcinogenesis. The stress protein heme oxygenase-1 (HO-1) catalyzes heme degradation producing biliverdin, iron and CO. Induction of HO-1 has been suggested to be essential for a controlled cell growth. The aim of this work was to analyze the in vivo homeostatic response (HR) triggered by the withdrawal of a potent carcinogen, p-dimethylaminoazobenzene (DAB), after preneoplastic lesions were observed. We analyzed HO-1 cellular localization and the expression of HO-1, Bcl-2 and cell cycle related proteins under these conditions comparing them to hepatocellular carcinoma (HC). METHODS: The intoxication protocol was designed based on previous studies demonstrating that preneoplastic lesions were evident after 89 days of chemical carcinogen administration. Male CF1 mice (n = 18) were used. HR group received DAB (0.5 % w/w) in the diet for 78 days followed by 11 days of carcinogen deprivation. The HC group received the carcinogen and control animals the standard diet during 89 days. The expression of cell cycle related proteins, of Bcl-2 and of HO-1 were analyzed by western blot. The cellular localization and expression of HO-1 were detected by immnunohistochemistry. RESULTS: Increased expression of cyclin E/CDK2 was observed in HR, thus implicating cyclin E/CDK2 in the liver regenerative process. p21cip1/waf1 and Bcl-2 induction in HC was restituted to basal levels in HR. A similar response profile was found for HO-1 expression levels, showing a lower oxidative status in the carcinogen-deprived liver. The immunohistochemical studies revealed the presence of macrophages surrounding foci of necrosis and nodular lesions in HR indicative of an inflammatory response. Furthermore, regenerative cells displayed changes in type, size and intensity of HO-1 immunostaining. CONCLUSION: These results demonstrate that the regenerative capacity of the liver is still observed in the pre-neoplastic tissue after carcinogen withdrawal suggesting that reversible mechanism/s to compensate necrosis and to restitute homeostasis are involved.