[Palliation by radiation is also our business !] / La radiothérapie palliative, c'est aussi notre rayon !

Radiotherapy (RT), both with a curative and a palliative intent, is one of the cornerstones of oncological treatments. A variety of symptoms linked to cancer can be relieved with RT (such as pain, bleeding, compression exerted by a tumour lesion…). Very often, palliative RT is proposed when other medical treatments (painkillers, morphine…) are no longer efficient, or the patient does not tolerate them anymore. Palliative RT is an integral part of the global supportive oncological care. Indeed, patients' wishes and prognosis are taken into account in each and every step of the treatment pathway. Every treatment deserves an individualized approach and benefits from the best available techniques.

La radiothérapie, à la fois à visée curative et palliative, est l'un des piliers des traitements oncologiques. Une multitude de symptômes liés au cancer (douleurs, saignements, diverses conséquences liées à une compression exercée par une lésion tumorale…) peuvent être soulagés grâce à une radiothérapie palliative (RTP). Bien souvent, la RTP est proposée lorsque les traitements médicamenteux dits «classiques¼ ne font plus suffisamment effet ou si le patient ne les tolère plus (antidouleurs, morphine…). La RTP fait partie intégrante des soins oncologiques de support. En effet, le pronostic du patient, ainsi que ses souhaits, sont pris en compte à chacune des étapes qui constituent le trajet de soins, y compris en RTP. Ainsi, chaque traitement est individualisé et bénéficie des meilleures techniques disponibles.

Assessment of the functional impact of germline BRCA1/2 variants located in non-coding regions in families with breast and/or ovarian cancer predisposition.

PURPOSE: The molecular mechanism of breast and/or ovarian cancer susceptibility remains unclear in the majority of patients. While germline mutations in the regulatory non-coding regions of BRCA1 and BRCA2 genes have been described, screening has generally been limited to coding regions. The aim of this study was to evaluate the contribution of BRCA1/2 non-coding variants. METHODS: Four BRCA1/2 non-coding regions were screened using high-resolution melting analysis/Sanger sequencing or next-generation sequencing on DNA extracted from index cases with breast and ovarian cancer predisposition (3926 for BRCA1 and 3910 for BRCA2). The impact of a set of variants on BRCA1/2 gene regulation was evaluated by site-directed mutagenesis, transfection, followed by Luciferase gene reporter assay. RESULTS: We identified a total of 117 variants and tested twelve BRCA1 and 8 BRCA2 variants mapping to promoter and intronic regions. We highlighted two neighboring BRCA1 promoter variants (c.-130del; c.-125C > T) and one BRCA2 promoter variants (c.-296C > T) inhibiting significantly the promoter activity. In the functional assays, a regulating region within the intron 12 was found with the same enhancing impact as within the intron 2. Furthermore, the variants c.81-3980A > G and c.4186-2022C > T suppress the positive effect of the introns 2 and 12, respectively, on the BRCA1 promoter activity. We also found some variants inducing the promoter activities. CONCLUSION: In this study, we highlighted some variants among many, modulating negatively the promoter activity of BRCA1 or 2 and thus having a potential impact on the risk of developing cancer. This selection makes it possible to conduct future validation studies on a limited number of variants.

Clinical value of R-spondins in triple-negative and metaplastic breast cancers.

BACKGROUND: RSPO ligands, activators of the Wnt/ß-catenin pathway, are overexpressed in different cancers. The objective of this study was to investigate the role of RSPOs in breast cancer (BC). METHODS: Expression of RSPO and markers of various cancer pathways were measured in breast tumours and cell lines by qRT-PCR. The effect of RSPO on the Wnt/ß-catenin pathway activity was determined by luciferase assay, western blotting, and qRT-PCR. The effect of RSPO2 inhibition on proliferation was determined by using RSPO2 siRNAs. The effect of IWR-1, an inhibitor of the Wnt/ß-catenin pathway, was examined on the growth of an RSPO2-positive patient-derived xenograft (PDX) model of metaplastic triple-negative BC. RESULTS: We detected RSPO2 and RSPO4 overexpression levels in BC, particularly in triple-negative BC (TNBC), metaplastic BC, and triple-negative cell lines. Various mechanisms could account for this overexpression: presence of fusion transcripts involving RSPO, and amplification or hypomethylation of RSPO genes. Patients with RSPO2-overexpressing tumours have a poorer metastasis-free survival (P=3.6 × 10-4). RSPO2 and RSPO4 stimulate Wnt/ß-catenin pathway activity. Inhibition of RSPO expression in a TN cell line inhibits cell growth, and IWR-1 significantly inhibits the growth of an RSPO2-overexpressing PDX. CONCLUSIONS: RSPO overexpression could therefore be a new prognostic biomarker and therapeutic target for TNBC.

[Functional imaging and radiotherapy]. / Utilisation de l'imagerie fonctionnelle en radiothérapie.

Medical imaging plays a crucial role in the diagnosis, staging and therapeutic strategy of oncologic patients. The development of medical imaging over the last decade has allowed significant progresses in radiotherapy. Indeed, medical imaging is now considered the corner stone of radiotherapy. The main challenge for the radiation oncologist consists in the tumour identification with a view to irradiate the tumour at a curative dose while avoiding healthy tissues. To achieve these goals, the radiotherapist daily uses anatomical imaging such as computed tomography (CT) or magnetic resonance imaging (MRI). Since several years now, the development of functional imaging such as positron emission tomography (PET) combined with CT or functional MRI has opened new perspectives in the management of oncologic diseases. Indeed, these imaging techniques offer new information on tumour metabolism that may be taken into account to plan the radiotherapy treatment. This article illustrates the different imaging techniques used in radiotherapy and the role of functional imaging for establishing new therapeutic strategies in radiation oncology.

Single and combined impacts of irradiation and surgery on lymphatic vasculature and fibrosis associated to secondary lymphedema.

Lymphedema (LD) refers to a condition of lymphatic dysfunction associated with excessive fluid accumulation, fibroadipose tissue deposition and swelling. In industrialized countries, LD development mainly results from a local disruption of the lymphatic network by an infection or cancer-related surgery (secondary LD). In the absence of efficient therapy, animal models are needed to decipher the cellular and molecular mechanisms underlying LD and test putative drugs. In this study, we optimized and characterized a murine model of LD that combines an irradiation of the mice hind limb and a radical surgery (lymph node resection associated to lymphatic vessel ligation). We investigated the respective roles of irradiation and surgery in LD formation by comparing their impacts, alone or in combination (with different intervention sequences), on eight different features of the pathology: swelling (paw thickness), indocyanine green (ICG) clearance, lymphatic vasculature remodeling, epidermal and dermal thickening, adipocyte accumulation, inflammatory cell infiltration and collagen deposition. This study supports the importance of radiation prior to surgery to experimentally induce a rapid, severe and sustained tissue remodeling harboring the different hallmarks of LD. We provide the first experimental evidence for an excessive deposition of periostin (POSTN) and tenascin-C (TNC) in LD. Through a computerized method of digital image quantification, we established the spatial map of lymphatic expansion, as well as collagen, POSTN and TNC deposition in papillary and reticular dermis of lymphedematous skins. This mouse model is available to study the patho-physiology of LD and test potential therapeutic targets.

[Cyberknife and benign pathologies]. / Cyberknife et pathologies bénignes.

Conventional radiotherapy is known to be an effective treatment approach even for "benign" pathologies. However, this kind of treatment yields a high potential for side effects. The Cyberknife, a robotic stereotactic radiotherapy device, enables to offset a large proportion of the disadvantages encountered with conventional radiotherapy essentially through the high precision of dose administration and sparing of healthy tissues. Therefore, it seems to be a treatment of choice in the approach of some benign intracranial diseases. We review published data on indications and outcome of Cyberknife for intracranial "non-malignant" disease.

Biological conversion of a water-soluble prodrug of cyclosporine A.

UNIL088 is a water-soluble prodrug of cyclosporine A (CsA) designed for topical ocular delivery. The pro-moiety is grafted via an ester function to CsA and the solubilizing group is a phosphate ion. The aim of this study was to elucidate the conversion mechanisms by which UNIL088 generates CsA. UNIL088 was incubated in rabbit tears at physiological temperature to study its enzymatic and chemical conversion, respectively. Metabolites and intermediates were identified using a quadrupole-time of flight (QqTOF) mass spectrometer, which allowed biotransformation pathways to be deduced. Conversion is activated by the chemical or enzymatic hydrolysis of the terminal ester function of the pro-moiety, leading to the phospho-serine-sarcosine-cyclosporine A that spontaneously converts into CsA. In addition to the main biotransformation pathway, a secondary reaction involved hydrolysis of the phosphate ester group of the pro-moiety, probably by phosphatases present in tears.

Evaluation of a topical cyclosporine A prodrug on corneal graft rejection in rats.

PURPOSE: To assess the efficacy of a topical cyclosporine A (CsA), water-soluble prodrug, for promoting the survival of allogenic rat corneal grafts after penetrating keratoplasty (PKP). METHODS: Corneas of Brown-Norway rats (donors) were transplanted to Lewis rats (recipients). Transplanted rats were divided in three treatment groups: group I (PBS) and group II (0.26% Debio088) received drops five times per day. Group III received a daily intramuscular CsA injection (10 mg/kg/day). Blood CsA concentrations were measured on days 2 and 14. On day 4, 10, 13 after PKP, grafts were scored for corneal transparency, edema and extent of neovascularization. An opacity score of greater than or equal to 3 was considered as a nonreversible graft rejection process. On day 14, the experimental eyes were processed for histology. RESULTS: On day 13, 12 of the 18 corneal transplants (67%) in group I showed irreversible graft rejection. Three of 18 transplants (19%) in group II and 5 of 16 transplants (28%) in group III showed irreversible graft rejection (p=0.013/p=0.019, OR=0.14/0.06 versus vehicle). Each mean clinical score for edema, opacity, and neovessels in group II were significantly lower than those of the grafts in group I (respectively p=0.010, p=0.013, p=0.024) and III except for neovessels (respectively p=0.002, p=0.001, p=0.057). Histology confirmed the clinical results. The mean CsA blood levels for groups II and III were, respectively 54+/-141 mug/l and 755+/-319 mug/l on day 2 and 14+/-34 mug/l and 1318+/-463 mug/l on day 14. CONCLUSIONS: Debio088 CsA prodrug drops given five times daily are as effective as intramuscular injection of 10 mg/kg/day for the prevention of acute corneal graft rejection in rats.

Cyclin D1 overexpression enhances radiation-induced apoptosis and radiosensitivity in a breast tumor cell line.

Overexpression of cyclin D1, a G1 cell cycle regulator, is often found in many different tumor types, such as breast carcinoma and squamous cell carcinoma of the head and neck. The overexpression of this protein is, in several cases, associated with a poor prognosis. In this study, the effect of cyclin D1 on radiosensitivity was investigated in a breast tumor cell line, MCF7, containing a cyclin D1 gene construct under the control of a tetracycline-sensitive regulator. MCF7 cells cultured without tetracycline resulted in a 6-fold increase in the cyclin D1 protein. Cyclin D1-overexpressing MCF7 cells were more sensitive to ionizing radiation than the nonoverexpressing counterparts. The cyclin D1-overexpressing cells also exhibited a higher induction of apoptosis. Treatment with a dose of 5 Gy resulted in a rapid increase of p53 and p21 in the cyclin D1-overexpressing cells. Nonoverexpressing cells showed a more transient expression of these proteins after ionizing radiation. A pronounced G2-M block was observed in both cell lines. The cyclin D1-overexpressing cells were, however, released earlier from the block than the control cells. These data suggest that overexpression of cyclin D1 alters sensitivity toward ionizing radiation by modulating gamma-radiation-induced G2-M transition.

Constitutive overexpression of cyclin D1 does not prevent inhibition of hormone-responsive human breast cancer cell growth by antiestrogens.

Cyclin D1 is a target for positive regulation by estrogens in growth-responsive cells, in which it mediates their mitogenic effects. Amplification and overexpression of the cyclin D1 gene (CCND1) might thus represent a genetic lesion inducing hormone-independent growth of transformed cells. Indeed, cyclin D1 overexpression has been found in up to 50% of primary breast cancers, and in about one-third of these cases, this is linked to amplification of the 11q13 chromosomal region, which also includes the CCND1 gene. These tumors are predominantly estrogen receptor-positive, and for this reason, these patients are often selected for adjuvant antiestrogen therapy. No information is available, however, as to whether cyclin D1 overexpression due to gene amplification might interfere with and reduce antiestrogen efficacy. This was investigated here by taking advantage of an experimental model that reproduces cyclin D1 overexpression resulting from increased CCND1 gene dosage in hormone-responsive human breast cancer cells. For this, MCF-7 cells stably transfected with a tet-inducible cyclin D1 expression vector were tested for their in vitro response to steroidal (ICI 182,780) and nonsteroidal (trans-4-hydroxytamoxifen) antiestrogens under condition of low (endogenous only) or high (exogenous) cyclin D1 levels. Results show that although cyclin D1 overexpression seems to interfere with the early cell cycle effects of antiestrogens, it does not prevent their cytostatic actions, so that growth of cyclin-overexpressing MCF-7 cells is still efficiently inhibited in vitro by these drugs.

Smad7 inhibits the survival nuclear factor kappaB and potentiates apoptosis in epithelial cells.

In this study, we examined the effect of the stable expression of Smad7 in two different cell lines on apoptosis induced by various stimuli including TGF-beta, serum withdrawal, loss of cell adhesion (anoikis) and TNF-alpha. Smad7 increased TGF-beta-mediated apoptosis in Mv1Lu cells as well as anoikis and/or serum withdrawal-induced apoptosis in Mv1Lu and MDCK cells. Smad7 markedly decreased the activity of the survival NF-kappaB transcription factor in MDCK cells. Interestingly, the stable expression of oncogenic Ras in MDCK cells which suppressed Smad7 inhibition of NF-kappaB also suppressed Smad7 potentiation of serum withdrawal-induced apoptosis and anoikis. In addition, Smad7 inhibited TNF-alpha stimulation of NF-kappaB and increased TNF-alpha-mediated apoptosis in MDCK cells. Our results provide the first evidence that Smad7 induces sensitization of cells to different forms of cell death. They moreover demonstrate that Smad7 inhibits the survival NF-kappaB factor, providing a potential mechanism whereby Smad7 potentiates cell death.

Improvement of endothelial function by chronic angiotensin-converting enzyme inhibition in heart failure : role of nitric oxide, prostanoids, oxidant stress, and bradykinin.

BACKGROUND-Chronic heart failure (CHF) impairs the endothelium-dependent, flow-mediated dilation (FMD) of small arteries. However, whether chronic angiotensin-converting enzyme (ACE) inhibition affects the impairment of FMD in CHF is unknown. We investigated the effects of long-term ACE inhibition on the FMD of peripheral arteries in rats with CHF and the mechanism(s) involved. METHODS AND RESULTS-FMD was assessed in isolated, perfused gracilis muscle arteries from sham-operated, and untreated or ACE inhibitor-treated (perindopril 2 mg. kg(-1). day(-1) for 10 weeks) rats with CHF (coronary artery ligation). The role of nitric oxide (NO), prostaglandins, and free radicals was assessed by pretreating the vessels with the NO synthase inhibitor N(W)-nitro-L-arginine, the cyclooxygenase inhibitor diclofenac, or the free radical scavenger N-2-mercaptopropionyl-glycine (MPG). Endothelial NO synthase mRNA expression was determined by reverse transcriptase polymerase chain reaction. In animals with hemodynamic and echographic signs of CHF, FMD was converted into vasoconstriction, and this was prevented by ACE inhibition. FMD of arteries from sham-operated or ACE inhibitor-treated CHF rats was abolished by N(W)-nitro-L-arginine. In untreated CHF rats, FMD was increased by diclofenac and MPG. In contrast, in arteries from ACE inhibitor-treated rats, neither diclofenac nor MPG affected FMD. In parallel, ACE inhibition prevented the reduction of endothelial NO synthase mRNA by CHF. CONCLUSIONS-In CHF, ACE inhibition normalized NO-dependent dilatation and suppressed the production of vasoconstrictor prostanoid(s), resulting in improved FMD. The improvement of FMD might contribute to the beneficial effects of ACE inhibition during CHF.

Functional evidence for a role of vascular chymase in the production of angiotensin II in isolated human arteries.

BACKGROUND: In human arteries, angiotensin-converting enzyme (ACE) inhibitors incompletely block the production of angiotensin (Ang) II from Ang I. This ACE-independent production of Ang II appears to be caused by serine proteases, one of which presumably is chymase. However, several serine proteases may produce Ang II, and the exact role of chymase in the vascular production of Ang II has never been directly evaluated using selective chymase inhibitors. METHODS AND RESULTS: Rings of human mammary arteries were subjected to either Ang I or the chymase-selective substrate [pro,(11) D-Ala(12)] Ang I in the absence or the presence of the ACE inhibitor captopril, the serine protease inhibitor chymostatin, or the selective chymase inhibitor C41. Captopril only partially inhibited (by 33%) the response to Ang I. In the absence of captopril, C41 markedly reduced (by 44%) the response to Ang I, and this effect was identical to that of chymostatin. C41 also significantly reduced the response to Ang I in the presence of captopril, although this inhibitory effect was slightly less than that of captopril in combination with chymostatin. [Pro,(11)D-Ala(12)] Ang I induced potent contractions that were not affected by captopril but were abolished by chymostatin and markedly reduced by C41. In addition, we found that prior treatment of the patients with an ACE inhibitor did not affect the in vitro response to Ang I (in the absence or the presence of captopril) or to [Pro,(11)D-Ala(12)] Ang I. CONCLUSIONS: Our results reinforce the hypothesis that chymase is a major serine protease implicated in the ACE-independent production of Ang II in human arteries.

Protective effects of preconditioning in cultured rat endothelial cells: effects on neutrophil adhesion and expression of ICAM-1 after anoxia and reoxygenation.

BACKGROUND: Preconditioning with brief periods of ischemia protects the coronary endothelium against acute and chronic reperfusion injury, but the mechanisms of this endothelial protection remain unknown. We hypothesized that preconditioning protects endothelial cells through a decreased production of endothelial adhesion molecules such as intercellular adhesion molecule-1 (ICAM-1), leading to a lesser adhesion of neutrophils to the endothelium. METHODS AND RESULTS: Cultured rat aortic endothelial cells were subjected to 6-hour anoxia followed by various durations of reoxygenation. Preconditioning was induced by 1-hour anoxia and 1-hour reoxygenation. ICAM-1 gene expression was measured by polymerase chain reaction, and the percentage of cells expressing ICAM-1 was assessed by confocal laser fluorescence microscopy. Anoxia/reoxygenation increased expression of ICAM-1, with a peak occurring after 6 hours of reoxygenation for mRNA and 9 hours for protein. Preconditioning prevented the increase in ICAM-1. Similar reductions were observed with the free radical scavenger N-2 mercaptopropionyl glycine (MPG). The inhibitory effect of preconditioning on ICAM-1 expression was abolished by an inhibitor of protein kinase C, an inhibitor of nitric oxide synthesis, and by MPG but was not affected by an adenosine receptor antagonist. Finally, both preconditioning and MPG partially prevented the increased adhesion of human neutrophils to reoxygenated endothelial cells. CONCLUSIONS: Preconditioning prevented reoxygenation-induced, free radical-mediated expression of ICAM-1 by a mechanism involving activation of protein kinase C and production of nitric oxide and free radicals, and this is associated with a lesser adhesion of neutrophils to endothelial cells. Such prevention of neutrophil adhesion may contribute to the protective effect of preconditioning against reperfusion-induced endothelial injury.

Exercise improves flow-mediated vasodilatation of skeletal muscle arteries in rats with chronic heart failure. Role of nitric oxide, prostanoids, and oxidant stress.

BACKGROUND: Flow-mediated dilatation (FMD) of the peripheral arteries may be impaired in chronic heart failure (CHF), and this could contribute to the increased peripheral resistance and exercise intolerance that occur with this disease. Physical exercise improves the FMD of large conduit arteries in CHF, but whether a similar impairment also occurs in smaller arteries is unknown. The mechanisms of the changes in FMD after CHF or exercise are also unknown. METHODS AND RESULTS: FMD was assessed in isolated, perfused, and preconstricted gracilis muscle arteries from sham-operated rats or CHF rats (coronary artery ligation) who were either sedentary or exercised (30-minute swimming period twice a day for 10 weeks, starting 7 days after ligation). In animals with hemodynamic and echographic signs of CHF, FMD was abolished and converted into vasoconstriction (percent change in diameter after 370 microL/min flow: sham, 42+/-5%; CHF, -4+/-3%; P<0.05). Exercise partially restored FMD (18+/-3%; P<0.05 versus CHF). In sham rats, FMD was abolished by the nitric oxide-synthase inhibitor Nomega-nitro-L-arginine (L-NA) but unaffected by the cyclooxygenase inhibitor diclofenac or the free radical scavenger N-(2-mercaptopropionyl)-glycine (MPG). In arteries from sedentary CHF rats, FMD was not modified by L-NA, but it was partially restored by diclofenac or MPG. In exercised CHF rats, FMD was abolished by L-NA and only moderately improved by diclofenac or MPG. Likewise, endothelial nitric oxide synthase mRNA expression (determined by reverse transcription polymerase chain reaction at the level of the gracilis muscle) was reduced by CHF, and this was prevented by exercise. CONCLUSIONS: CHF abolishes the FMD of small arteries by impairing the nitric oxide pathway, increasing oxidant stress, and releasing a prostanoid-contracting factor. Exercise partially restores FMD by increasing expression of endothelial nitric oxide synthase and preventing the production of vasoconstrictor prostanoids and free radicals. Such restoration of FMD might contribute to the increase in exercise capacity after physical exercise in CHF.

Increased survival after long-term treatment with mibefradil, a selective T-channel calcium antagonist, in heart failure.

OBJECTIVES: We sought to investigate the effects of mibefradil on survival, hemodynamic variables and cardiac remodeling in a rat model of chronic heart failure (HF) and to compare these effects with those of the angiotensin-converting enzyme (ACE) inhibitor cilazapril. BACKGROUND: The use of calcium channel blocking agents in chronic HF has been disappointing. Most studies have shown that these drugs have either no or even detrimental effects due in part to the negative inotropic effects they induce. Mibefradil is a calcium channel blocker that selectively blocks T channels and displays moderately negative inotropic properties only at high doses. Because T channels are upregulated in the hypertrophied heart and could mediate hypertrophic signals and increase arrhythmogenicity, blockade of these channels might be beneficial in chronic HF. METHODS: Rats were subjected to coronary artery ligation and 9 months of treatment with mibefradil (15 mg/kg body weight per day) or cilazapril (10 mg/kg per day) or no treatment. Survival and systolic blood pressure were assessed over the 9-month treatment period, after which cardiac hemodynamic variables and structure were determined. RESULTS: Mibefradil increased survival rate to the same extent as cilazapril (71% for mibefradil vs. 75% for cilazapril and 44% for no treatment). Mibefradil decreased systolic blood pressure, although to a lesser extent than cilazapril. Both treatments decreased left ventricular (LV) end-diastolic and central venous pressures, without any change in the first derivative of LV pressure over time or heart rate. Mibefradil decreased LV weight (although less than cilazapril) without affecting right ventricular weight. Finally, both drugs normalized LV collagen density. CONCLUSIONS: Mibefradil in a rat model improved survival to the same extent as an ACE inhibitor, without impairing LV function, and was associated with a reduction in LV weight and fibrosis. Thus, mibefradil might be beneficial in the treatment of chronic HF.

Conversion of cyclosporine A prodrugs in human tears vs rabbits tears.

The aim of this study was to evaluate the rate and mechanism of conversion of two water-soluble prodrugs of cyclosporine A (CsA) intended for topical delivery to the eye. The new molecules were designed according to the double prodrug concept: a solubilizing moiety was grafted onto CsA via an ester function, which could be hydrolysed via a two-step process (enzymatic and chemical). Prodrug solutions were prepared extemporaneously in an isotonic and neutral aqueous medium compatible with ophthalmic use. The rates of conversion into the parent molecule were determined by incubating the prodrugs in fresh rabbit or human tears or in a phosphate buffer solution (PBS) at pH 7.4. Both prodrugs were converted into CsA within the first minute in the presence of rabbit tears with rate constants of k=5.9x10(-3)min(-1) and k=3.8x10(-3)min(-1), respectively, for UNIL088 and UNIL089, whereas chemical conversion in PBS was negligible (k=0.5x10(-3)min(-1) for both molecules). Incubation of UNIL088 in human tears showed a significantly high conversion rate. It is concluded that the developed double prodrugs underwent a bioconversion in physiological media and thus represent promising candidates for topical delivery of CsA to the eye.

A novel water-soluble cyclosporine A prodrug: ocular tolerance and in vivo kinetics.

The purpose of this study is to demonstrate that a novel water-soluble prodrug of cyclosporine A (CsA) intended for topical ocular administration, does not induce eye irritation in a rabbit model and is able to generate therapeutic concentrations of CsA in the precorneal area immediately after administration. The eye irritancy of the prodrug and CsA control solution was assessed by the Draize test and by confocal laser ophthalmoscopy (CLSO). Residence time and tear concentrations of prodrug and CsA in the rabbit eye were assessed by HPLC. The Draize test showed an excellent tolerance for the prodrug solution while the reference CsA oil solution induced lachrymation and irritation. The CLSO-measured corneal lesions, subsequent to treatment with the prodrug and reference solutions, were 3% and 9%, respectively. The prodrug transformed rapidly, leading to relatively stable CsA concentrations in tears with a maximal concentration of 94 microg ml(-1) over the observation period. This study demonstrated that the prodrug solution was well tolerated and that clinically significant CsA tear concentrations were achieved. UNIL088 is a promising molecule in the treatment of immune-related disorders of the eye.

Effects of high frequency ultrasound irradiation on incorporation of SiO2 particles within polypyrrole films.

This paper deals with the effect of ultrasound on polypyrrole/SiO2 composite film elaboration through various steps (particle dispersion, electrosynthesis). Experiments were carried out on stainless steel in phosphoric acid solution. An efficient method for dispersion of SiO2 particles prior to electropolymerization, based on low frequency irradiation (20kHz), was proposed. It was shown that mechanical effects of high frequency ultrasound (i.e. mass transfer improvement) led to enhancement of electropolymerization kinetics. Scanning electron microscopy imaging and glow discharge optical emission spectroscopy revealed localization of SiO2 particles in the outer region of the films as well as better incorporation of particles under high frequency ultrasound irradiation. Finally, anticorrosion behavior of formed films was investigated in sodium chloride solution by Open Circuit Potential and anodic polarization methods. The results showed that polypyrrole/SiO2 films elaborated under ultrasound irradiation exhibit the best protective performances.

Activation of mitogen-activated protein kinase cascades is involved in regulation of bone morphogenetic protein-2-induced osteoblast differentiation in pluripotent C2C12 cells.

Bone morphogenetic protein (BMP)-2, a member of the transforming growth factor-beta (TGF-beta) superfamily, is able to induce osteoblastic differentiation of C2C12 cells. Both Smad and mitogen-activated protein kinase (MAPK) pathways are essential components of the TGF-beta superfamily signaling machinery. Although Smads have been demonstrated to participate in the BMP-2-induced osteoblastic differentiation of C2C12 cells, the role of MAPK has not been addressed. This report shows that BMP-2 activates ERK and p38, but not JNK, in C2C12 cells. Pretreatment of cells with the p38 inhibitor, SB203580, dramatically reduced BMP-2-induced expression of the osteoblast markers alkaline phosphatase (ALP) and osteocalcin (OC). Nevertheless, overexpression of MKK3, a protein kinase that phosphorylates and activates p38, failed to induce ALP or OC expression in the absence of BMP-2, indicating that p38 activation is necessary but not sufficient for the acquisition of the osteoblast phenotype by these cells. Although ALP induction was increased slightly in the presence of PD-98059, a selective inhibitor of the ERK cascade, this compound significantly inhibited both steady-state and BMP-2-induced OC RNA levels. Our results indicate that p38 and ERK cascades play a crucial role in the osteoblast differentiation of C2C12 cells mediated by BMP-2.