Current gene panels account for nearly all homologous recombination repair-associated multiple-case breast cancer families.

It was hypothesized that variants in underexplored homologous recombination repair (HR) genes could explain unsolved multiple-case breast cancer (BC) families. We investigated HR deficiency (HRD)-associated mutational signatures and second hits in tumor DNA from familial BC cases. No candidates genes were associated with HRD in 38 probands previously tested negative with gene panels. We conclude it is unlikely that unknown HRD-associated genes explain a large fraction of unsolved familial BC.

A combination of pyridine-2, 4-dicarboxylic acid diethyl ester and resveratrol stabilizes hypoxia-inducible factor 1-alpha and improves hair density in female volunteers.

OBJECTIVE: The aim of this study was first to demonstrate that a combination of pyridine-2, 4-dicarboxylic acid diethyl ester and resveratrol could synergize in vitro on biological pathways associated with hair growth and then to demonstrate the benefit on hair density in a clinical study. METHODS: The effects of pyridine-2, 4-dicarboxylic acid diethyl ester and resveratrol directly on the hypoxic inducible factor-1α protein (HIF-1α) and related genes expression were demonstrated on keratinocytes in culture in vitro using western-blot analysis and real time quantitative polymerase chain reaction analysis. The effect of resveratrol against oxidative stress induced by hydrogen peroxide treatment was studied in hair follicle and hair matrix cells in vitro using the sensitive probe Dichloro-dihydro-fluorescein diacetate (DCFH-DA). Finally, a randomized clinical study on hair density was conducted on 79 Caucasian female subjects to assess the effect of this combination of actives. RESULTS: Pyridine-2, 4-dicarboxylic acid diethyl ester and resveratrol stabilized HIF-1a protein and increased the expression of HIF-1α target genes. Resveratrol significantly reduced the oxygen peroxide-induced oxidative stress generated in hair follicle and hair matrix cells. The clinical study showed that a topical treatment with the combination significantly increased the hair density on women from 1.5 months. CONCLUSION: In addition to the antioxidant properties of resveratrol, the association of pyridine-2, 4-dicarboxylic acid diethyl ester and resveratrol revealed a synergistic effect on the HIF-1α pathway. The results of the clinical study confirmed the importance of such a combination to increase the hair density.

L'alopécie peut affecter 50% des femmes au cours de leur vie ce qui induit une perte de leur estime de soi et une diminution de leur qualité de vie. Au-delà des solutions chirurgicales et des traitements pouvant induire des effets secondaires potentiellement dangereux, il y a un besoin d'améliorer l'efficacité des produits cosmétiques qui permettent de prévenir la chute des cheveux tout en préservant la sécurité des patients. Ainsi, nous avons sélectionné une combinaison de pyridine-2, 4-dicarboxylic acide diethyle ester et de resvératrol pour activer des voies biologiques associées à la croissance du cheveu. Nous avons d'abord montré, in vitro, que la combinaison de pyridine-2, 4-dicarboxylic acide diethyle ester et de resvératrol permet de stabiliser la protéine HIF-1α conduisant ainsi à un effet synergique sur l'expression de gènes clés de la voie HIF-1α. Nous avons aussi démontré, in vitro, que le resvératrol permet de protéger significativement les follicules pileux et les cellules de la matrice du stress oxydatif induit par traitement au peroxide d'hydrogène. En final, une étude clinique randomisée mesurant la densité capillaire a été réalisée sur 79 femmes caucasiennes. Cette étude montre qu'une application topique d'une solution contenant de 5% pyridine-2, 4-dicarboxylic acide diethyle ester et 0.25% de resvératrol augmentent significativement la densité capillaire chez les femmes après 1.5 mois. En conclusion, ces résultats démontrent l'intérêt de stimuler la voie HIF-1α tout en protégeant les cheveux et le scalp du stress oxydatif afin d'améliorer la croissance des cheveux chez les femmes.

Tetracycline use in the community may promote decreased susceptibility to quinolones in Escherichia coli isolates.

We previously found that the hospital use of tetracyclines is associated with quinolone resistance in hospital isolates of Enterobacteriaceae. Tetracyclines are heavily used in the community. Our aim was to assess whether their use in the community favors quinolone resistance in community isolates of Escherichia coli. Monthly data of community antibiotics use and E. coli quinolone resistance in a 1.3 million inhabitant French area were obtained from 2009 to 2014, and were analyzed with autoregressive integrated moving average (ARIMA) models. Quinolone use decreased from 10.1% of the total antibiotic use in 2009 to 9.3% in 2014 (trend, - 0.016; p-value < 0.0001), while tetracycline use increased from 16.5% in 2009 to 17.1% in 2014 (trend, 0.016; p < 0.0001). The mean (95% confidence interval) monthly proportions of isolates that were non-susceptible to nalidixic acid and ciprofloxacin were 14.8% (14.2%-15.5%) and 9.5% (8.8%-10.1%), respectively, with no significant temporal trend. After adjusting on quinolone use, tetracycline use in the preceding month was significantly associated with nalidixic acid non-susceptibility (estimate [SD], 0.01 [0.007]; p-value, 0.04), but not with ciprofloxacin non-susceptibility (estimate [SD], 0.01 [0.009]; p-value, 0.23). Tetracycline use in the community may promote quinolone non-susceptibility in E. coli. Decreasing both tetracycline and quinolone use may be necessary to fight against the worldwide growth of quinolone resistance.

Three years of antibiotic consumption evaluation in French nursing homes.

OBJECTIVES: We had for aim to assess antibiotic consumption and to better understand their use in nursing homes so as to target messages on relevant practice procedures sent to prescribers. DESIGN: The MedQual network asked nursing homes with in-house pharmacies to participate in a retrospective collection of yearly antibiotic consumption data with an Excel(®) spread sheet according to the Health Ministry recommendations. RESULTS: Fifty-two nursing homes participated in 2011, 2012, and 74 in 2013, accounting for 10% of the Pays de la Loire region's nursing homes and 15% of beds. The medians of total antibiotic consumption in daily-defined dose for 1000 patient-days were respectively 39 (32.4-49.0), 39.3 (34.4-52.9), and 44.8 (33.6-55.4). There was no significant difference between 2011 and 2013. Penicillins (J01C) were the most commonly used class with a median of 25.7 [IQ 18.8; 33.8] in 2011 and 30.4 [IQ 23.6; 41.3] in 2013. Quinolones (J01M) were the second most commonly used class with a median of 4.6 [IQ 2.9; 5.9] in 2011 and 3.8 [IQ 2.3; 6.5] in 2013, followed by the other beta-lactams (J01D) with a median of 2.5 [IQ 1.7; 4.5] in 2011 and 2,8 [IQ 1.7; 3.8] in 2013. CONCLUSION: The monitoring of antibiotic consumption in nursing homes in the Pays de la Loire Region since 2011 has allowed identifying inappropriate use and helped improve practices. No increase of overall consumption was observed in nursing homes but the distribution according to antibiotic class changed. The current objective is to extend this monitoring and to send personalized messages to prescribers.

Epidemiology of third-generation cephalosporin-resistant community-acquired Enterobacteria isolated from elderly patients.

AIMS: This survey was made to study the epidemiology of multiresistant bacteria (MRB) in the French community, among elderly patients 65 years of age or more, carrying third-generation cephalosporin-resistant (3GC-resistant) Enterobacteriaceae, and the co-resistance of prescribed antibiotics. METHODS: The data was collected in 2009 in the West of France by MedQual, a network of 174 private laboratories. RESULTS: Two thousand one hundred and sixty strains of the 88,255 identified Enterobacteria strains were 3GC-resistant (2.4%) and 945 of these strains (41.8%) were isolated from elderly patients 65 years of age or more. Escherichia coli was the predominant 3GC-resistant strain (72.7%). 51.4% of the 945 patients in whom a 3GC-resistant strain was isolated produced an extended-spectrum ß-lactamase (ESBL). The main risk factors for infection with the 3GC-resistant strain were hospitalization and antibiotic treatment in the previous year (58.2 and 86.9%, respectively). Hospitalization during the previous year was more frequent among elderly patients who lived at home compared with those who lived in nursing homes (P<0.05). The production of ESBL, among the 945 patients who carried the 3GC-resistant strains, was similar among patients who lived at home compared with those who lived in nursing homes (51.4% versus 49.7%). CONCLUSION: Microbiologists should warn family physicians about MRB isolates with a specific antimicrobial resistance pattern (3GC-resistant, fluoroquinolone-resistant, etc.) to prescribe more effective medications.

A national network of infectious diseases experts.

OBJECTIVES: The aim of this study was to consider the implementation of a network of infectious diseases (ID) experts to optimize the antibiotic prescriptions of community and hospital practitioners. METHODS: An observational prospective study was conducted among ID experts in the Pays-de-la-Loire Region to evaluate the number of calls and to determine the practitioner's reasons for soliciting ID expertise. For each phone consultation, four criteria were recorded during 5 days: origin of the call (internal/external), kind of question (diagnostic/therapeutic) time spent for the advice provided, type of advice. RESULTS: A total of 386 phone consultations for 20 infectious disease specialists were recorded during the study period (5 days); 81% were internal to the hospital, 7.7% from another hospital, and 11.3% from private practice, 56.3% of the questions concerned a therapeutic strategy, 21% a diagnostic advice, and 22.6% concerned both diagnosis and therapy. Two third of the questions were answered within 10minutes. In 68.7% of cases, the ID specialist answered immediately, 19.8% of calls required following-up the patient, 6% led to refer the patient to an ID consultation, and 5.5% to hospitalization. CONCLUSION: The survey results stress the important need for such ID expertise, both in hospitals and in ambulatory medicine. Collaboration of ID specialists in a regional network would allow an easy and permanent access to antibiotic therapy advice for prescribers. This network would improve the quality and safety of care.

Chronological ageing of human hair keratin fibres.

Examination of very long hair (length > 2.4 m) using a large range of evaluation methods including physical, chemical, biochemical and microscopic techniques has enabled to attain a detailed understanding of natural ageing of human hair keratin fibres. Scrutinizing hair that has undergone little or no oxidative aggression--because of the absence of action of chemical agents such as bleaching or dyeing--from the root to the tip shows the deterioration process, which gradually takes place from the outside to the inside of the hair shaft: first, a progressive abrasion of the cuticle, whilst the cortex structure remains unaltered, is evidenced along a length of roughly 1 m onwards together with constant shine, hydrophobicity and friction characteristics. Further along the fibre, a significant damage to cuticle scales occurs, which correlates well with ceramides and 18-Methyl Eicosanoic Acid (18-MEA) decline, and progressive decrease in keratin-associated protein content. Most physical descriptors of mechanical and optical properties decay significantly. This detailed description of natural ageing of human hair fibres by a fine analysis of hair components and physical parameters in relationship with cosmetic characteristics provides a time-dependent 'damage scale' of human hair, which may help in designing new targeted hair care formulations.

Susceptibility to the main antibiotics of Escherichia coli and Staphylococcus aureus strains identified in community acquired infections in France (MedQual, 2004-2007).

BACKGROUND: The aim of this study was to determine the susceptibility of bacterial strains identified in community-acquired infections. Surveillance was made by a network of 32 medical analysis laboratories in the five departments of the French Region "Pays de la Loire". METHODS: All Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus) strains isolated in these laboratories over a 4-year period, from January 2004 to December 2007, were included in the investigation. RESULTS: Eighty-four thousand and twenty-nine strains were collected: 90.6% of E. coli and 9.4% of S. aureus. E. coli isolates were mainly isolated from urine (97.2%). S. aureus isolates were more frequently isolated from pus (42.30%), from urinary samples (19.53%), or genital tract samples (14.36%). This study confirms the worrying E. coli evolution of resistance to quinolones. Indeed, during the study period, ofloxacin or norfloxacin susceptibility decreased gradually and the susceptibility rate to ciprofloxacin decreased slightly during and after 2006 (94.01% in 2005; 92.81% in 2006, and 91.62% in 2007). One thousand four hundred and thirty-five methicillin-resistant S. aureus (MRSA) strains were isolated. We observed a decrease of resistance to oxacillin: 20.73% in 2004 and 16.65% in 2006 (p<0.01). In 2007, this resistance to oxacillin seemed to increase (18.26%). CONCLUSIONS: Our data confirms the serious need to monitor transmission of these strains between community and hospitals. A better knowledge of the epidemiological behavior of these BMR will contribute to better-adapted antibiotics strategies.

Who are the carriers of MRSA in the community? A prospective study in the Pays de la Loire region of France.

The aim of this study was to determine the demographic characteristics of methicillin-resistant Staphylococcus aureus (MRSA) carriers in the community, to assess their risk factors and possible past hospitalization history and to describe the different resistance phenotypes of community isolates of S. aureus. Data were collected over the course of 16 months (from June 2005 to September 2006) in the Pays de la Loire region of France by MedQual, a network of private biological analysis laboratories. This work was based solely on the analysis of strains isolated in the community as opposed to isolates from private facilities such as nursing homes or hospitals. The antimicrobial susceptibility results for a total of 313 MRSA isolates were included in this study. The isolates were most frequently recovered from skin and soft tissue infections (41.2%), urine (38.3%) and genital samples (8.3%). We distinguished 36 patients without classical risk factors (WRF), such as demographic individual medical, healthcare exposure, carried MRSA, from the other 277 patients with at least one risk factor (RF). WRF MRSA patients were younger than RF patients and an infection was more often found among WRF patients. MRSA strains isolated from RF patients were resistant to ofloxacin in 81.1% of cases, whereas only 50% of the MRSA strains isolated from WRF patients were resistant (p <0.001). Nine resistance phenotypes were observed among the 313 MRSA strains. MRSA resistance profiles in the community have evolved in recent years. Therefore, it is necessary to study the resistance phenotypes of the circulating strains in order to adapt therapeutic care in the community.

Human eyelash characterization.

BACKGROUND: Few biological data on human eyelash follicles have been reported in the literature. OBJECTIVES: To characterize eyelash follicle growth, cycle and morphology, and further investigate the biological mechanisms that determine eyelash length, curl and pigmentation, compared with scalp hair follicle. METHODS: Twenty-nine caucasian female volunteers aged between 26 and 60 years were enrolled in the study to provide eyelashes. Four of these volunteers were followed weekly for 9 months to characterize their eyelash cycle. Eyelash length and time of renewal were measured using a high-resolution camera and image analysis. Immunohistological study of the bulbs were performed on eyelid biopsies from 17 patients requiring block excision for ectropion repair. RESULTS: The calculated durations of anagen phase and complete cycle of the eyelashes were 34 + or - 9 and 90 + or - 5 days, respectively. Eyelash follicle growth rate was quite variable, with an average rate of 0.12 + or - 0.05 mm daily. Eyelash follicle morphology was very close to that of the scalp hair follicle, but some remarkable differences were noticed. For example, the K19-positive epithelial stem cell population was spread all along the follicle and not split into two reservoirs as seen in scalp hair follicles. Some asymmetry was detected in HSPG and CSPG, as well as K38 (formerly Ha8) and K82 (formerly Hb2) distribution, similar to that observed in curly hair. Finally, dopachrome tautomerase was found expressed in eyelash follicle melanocytes, while it was strikingly absent in scalp hair follicle melanocytes. CONCLUSIONS: The eyelash is structurally very close to curly hair but some biological processes related to follicle cycle and pigmentation differ markedly.

Proteoglycan expression patterns in human hair follicle.

BACKGROUND: Proteoglycans (PGs) are known to play key roles in many cellular signalling pathways involved in hair follicle biology. Although some PG core proteins have previously been described in adult human hair follicles, their glycosaminoglycan (GAG) moieties have been less studied. OBJECTIVES: To add knowledge about PG core protein and GAG distributions in human anagen hair follicle and, for selected follicles, during catagen. METHODS: We used immunohistochemistry and immunohistofluorescence to revisit the expression pattern of GAG chains and core proteins in human hair follicle. The studied epitopes included CD44v3, syndecan-1, perlecan, versican, aggrecan, biglycan, heparan sulphate (HS), chondroitin sulphate (CS), dermatan sulphate (DS) and keratan sulphate (KS). RESULTS: The membrane PGs syndecan-1 and CD44v3 were respectively detected in the epithelial part of whole hair and in the outer root sheath basal layer. The dermal part of the hair follicle contained high amounts of extracellular PGs such as perlecan, versican, aggrecan, biglycan and their saccharidic moieties, namely HS, CS, DS and KS. We also observed a variable distribution of these components along the hair follicle. Especially, we noted a PG impoverishment at the very bottom of the anagen bulb. Moreover, while type D chondroitin expression remained unaffected, 4C3-CS and PG4-CS/DS epitopes respectively decreased in the dermal papilla and the connective tissue sheath, at the onset of catagen. CONCLUSIONS: GAG and PG expression along the human anagen hair follicle was characterized by (i) discontinuities mainly affecting the basement membrane and (ii) disappearance of some epitopes at catagen onset. These results are discussed in term of functionalities in nutrient diffusion, cell proliferation and differentiation, and hair protection.

Protective effects of taurine on human hair follicle grown in vitro.

Taurine is a naturally occurring beta-amino acid produced by methionine and cysteine metabolism. It is involved in a variety of physiological functions, including immunomodulatory and antifibrotic. Taking advantage of the ability of human hair follicle grown in vitro to recapitulate most of the characteristic features of normal hair follicle in vivo, we studied (i) taurine uptake by isolated human hair follicles; (ii) its effects on hair growth and survival rate; and (iii) its protective potential against transforming growth factor (TGF)-beta1, an inhibitor of in vitro hair growth and a master switch of fibrotic program. We showed that taurine was taken up by the connective tissue sheath, proximal outer root sheath and hair bulb, promoted hair survival in vitro and prevented TGF-beta1-induced deleterious effects on hair follicle.

Human hair shape is programmed from the bulb.

BACKGROUND: Few biological data on curly hair follicles have been reported in the literature. OBJECTIVES: To investigate the growth and morphology of curly hair follicles. METHODS: Follicles were dissected from scalp skin samples from African, Guyanese and caucasian volunteers and were observed macroscopically, in culture in William's E medium, and by immunohistochemistry. RESULTS: Macroscopic study of scalp biopsies obtained from African volunteers showed that the dermal implantation of follicles was curved with a retrocurvature at the level of the bulb, as opposed to a straight shape in caucasian follicles. The bulb itself was bent, in the shape of a golf club, while both the outer root sheath (ORS) and the connective tissue sheath were dissymmetrical along the follicle. In vitro growth of curly hair follicles was slightly slower than that of caucasian follicles but, more importantly, the curvature was maintained in the hair shaft produced in vitro. As shown by immunohistochemistry, the proliferative matrix compartment of curly hair follicles was asymmetrical, Ki-67-labelled cells being more numerous on the convex side and extending above the Auber line. On the convex part of the follicle, the ORS was thinner and the differentiation programmes of the inner root sheath and hair shaft were delayed. Furthermore, some ORS cells expressed alpha-smooth muscle actin protein on the concave side of the curvature, reflecting a mechanical stress. CONCLUSIONS: Hair curliness is programmed from the bulb and is linked to asymmetry in differentiation programmes.

Tissue detection of diphenylchlorin sensitizer (SIM01) by fluorescence and high-performance liquid chromatography.

Cancer is today a major problem of public health. Unfortunately, the current treatments remain still too often impotent or too heavy compared to the gross national product of many countries. The use of PDT in the treatment of the malignant tumours currently raises great hopes. This physicochemical method is based on the combined action of a nontoxic drug given systematically to the patient and of the visible light delivered locally to the tumour using optical fibres. The radiation will activate the significant substance preferentially fixed on cancerous cells and will cause the death of the tumoral cells while releasing from the toxic ridicalizing species which then will deteriorate vital cellular targets. Tissue distribution and elimination kinetics of the SIM01 were analysed in biological samples from mice tissues by spectrofluorometry and HPLC. Measurements were performed 4, 6, 12, 24 and 48h after an intraperitoneal injection for SIM01 doses of 2, 5 and 15mgkg(-1). Elimination seemed to concern essentially gallbladder, liver and stools, where maximum fluorescence reached, respectively, 20,000, 2800 and 15,000cps for 5mgkg(-1), 6h after injection. Among the tissues examined with HPLC, the highest SIM01 levels were found in stools, urine, liver, gallbladder and spleen. Liver, gallbladder, and stool homogenates from drug-treated animals contained an additional peak (16, 7min) detectable only after injection of at least 15mgkg(-1). Our HPLC determinations and in vivo fluorescence detection of SIM01 gave comparable kinetic profiles. These techniques should be considered as complementary rather than exclusive for kinetic profiles determination.

Hair keratin pattern in human hair follicles grown in vitro.

The keratin family includes epithelial (soft) keratins and hair (hard) keratins, and can be divided into acidic type I and basic to neutral type II subfamilies. Recently, nine type I and six type II hair keratin genes have been characterized through the screening of a human PAC library. The expression of these genes in the hair follicle was determined in vivo and a combined catalog of acidic and basic hair keratins was established. In this study, we investigated the expression and localization of most of the human hair keratin members of both types in human hair grown in vitro. We show that in vitro growth of hair follicles for 10 days in complete William's E culture medium did not alter the expression pattern of hair keratins. Similarly to the in vivo situation, each hair keratin was localized in precise and discrete compartments of the follicle, ranging from the matrix to the upper cortex and/or the hair cuticle. This study shows that the increase in length of in vitro grown follicles was accompanied by the proper hair shaft keratinization process. It also shows that hair follicle integrity was maintained in vitro, both in terms of gross morphology and molecular organization despite the complexity of the keratin expression pattern.

Synthesis, and in vitro and in vivo evaluation of a diphenylchlorin sensitizer for photodynamic therapy.

This paper reports the synthesis of a new diphenylchlorin photosensitizer, 2,3-dihydro-5,15-di(3,5-dihydroxyphenyl)porphyrin (SIM01). The photodynamic properties, cell uptake and localization of SIM01 were compared with those of structurally related meso-tetra(hydroxyphenyl)chlorin (m-THPC). In vitro studies were conducted on rat glioma cells (C6) and human adenocarcinoma (HT-29), and in vivo studies on human colon adenocarcinoma cells (HT-29) and human prostate adenocarcinoma cells (PC3). Both dyes showed an absorption maximum at around 650 nm, with a molar extinction coefficient of 13017 M(-1) cm(-1) for SIM01 and 22718 M(-1) cm(-1) for m-THPC. Their capacity to generate singlet oxygen was identical, but differences in partition coefficients indicated that SIM01 was slightly more hydrophilic. In vitro, SIM01 was slightly more phototoxic than m-THPC for C6 cells (4.8 vs. 6.8 microg ml(-1)). However, phototoxicities were nearly identical for HT29 cells (0.45 microg ml(-1) for 5 h incubation followed by 300 mW, 20 J cm(-2)). Pharmacokinetics in vivo in mice, as determined by fibre spectrofluorimetry, showed that the SIM01 fluorescence signal in the tumor was maximal between 6 and 12 h after injection, as compared to 72 h for m-THPC. With a 2 mg kg(-1) dye dose and laser irradiation at 300 J cm(-2) (650 nm, 300 mW), the optimal PDT response occurred when the interval between injection and irradiation was 6 h for SIM01 and 24 h for m-THPC. For SIM01 with 5 mg kg(-1) injection, the optimal PDT response occurred with a 12 h delay and with the same irradiation parameters as described above, in this case the tumor response showing 40% growth. Considering the tumor volume doubling time, the value was 6.5 days in the control group and increased to 13.5 days with SIM01. Thus, SIM01 may be a powerful sensitizer characterized by strong in vitro and in vivo phototoxicity and faster tissue uptake and elimination than m-THPC.

PDT effects of m-THPC and ALA, phototoxicity and apoptosis.

The purpose of this study was to estimate the efficacy of an endogenous sensitizer (delta-aminolevulinic acid (or ALA) induced protoporphyrin IX (or PpIX)) and an exogenous sensitizer (meta(tetrahydroxyphenyl)chlorin or m-THPC) on two different cell lines, rat colon adenocarcinoma PROb cells and murine melanoma B16A45 (B16) cells, in apoptosis production. After sensitizer incubation, cells were irradiated with an argon dye laser. LD(50) with m-THPC was 2.8 microg/ml and 4.7 microg/ml under irradiation of 25 J/cm(2) respectively for PROb and B16 cells. With ALA, LD(50) was 150 microg/ml and 175 microg/ml under 25 J/cm(2) respectively for PROb and B16 cells. Apoptosis induction by m-THPC or ALA-PDT was detected by DNA gel electrophoresis and quantified using an ELISA assay 24 h after PDT. The maximal apoptosis enrichment factor (MAEF) was reached for 6 microg/ml m-THPC at 10 J/cm(2) for PROb and B16 cells and for 50 microg/ml ALA at 25 J/cm(2) for PROb or B16 cells. Both m-THPC and PpIX are efficient photosensitizers and apoptosis inducers. However, MAEF is obtained by sensitizer or laser doses inducing very different phototoxic effects: MAEF was obtained after m-THPC-PDT with LD(78) for PROb cells and LD(30) for B16 cells and after ALA-PDT with LD(22) for PROb cells and LD(18) for B16 cells. However the overall m-THPC/PDT apoptotic induction (under the curve surface analysis) was not different whatever the cell line for 10 and 25 J/cm(2). On the contrary, ALA-PpIX/PDT apoptotic induction was twice for 25 J/cm(2) as compared to 50 J/cm(2) (p < 0.01) for both the PROb and B16 cells. These results indicate that the apoptosis rate in PDT cell killing varies considerably according to cell type and sensitizer.

Effects of BAPTA-AM, Forskolin, DSF and Z.VAD.fmk on PDT-induced apoptosis and m-THPC phototoxicity on B16 cells.

As many types of cells exposed to photodynamic therapy (PDT) appear to undergo apoptosis, various apoptosis inhibitors have already been used in studies of PDT-induced apoptosis. Although these inhibitors decrease apoptosis, their real effect on the phototoxic efficacy of photosensitisers is unclear. The good phototoxicity of m-THPC was confirmed on murine melanoma B16-A45 cells. Toxicity and phototoxicity studies were then carried out using four apoptosis inhibitors: BAPTA-AM, Forskolin, DSF, and Z.VAD.fmk. Although all inhibitors tested blocked PDT-induced apoptosis, none produced a significant modification of the phototoxic effect of m-THPC on B16 cells. It has been suggested that apoptosis and necrosis share common initiation pathways and that the final outcome is determined by the presence of an active caspase. This implies that apoptosis inhibition reorients cells to necrosis, i.e. those cells sufficiently damaged by PDT appear to be killed, regardless of the mechanism involved.