Promoter methylation of argininosuccinate synthetase-1 sensitises lymphomas to arginine deiminase treatment, autophagy and caspase-dependent apoptosis.

Tumours lacking argininosuccinate synthetase-1 (ASS1) are auxotrophic for arginine and sensitive to amino-acid deprivation. Here, we investigated the role of ASS1 as a biomarker of response to the arginine-lowering agent, pegylated arginine deiminase (ADI-PEG20), in lymphoid malignancies. Although ASS1 protein was largely undetectable in normal and malignant lymphoid tissues, frequent hypermethylation of the ASS1 promoter was observed specifically in the latter. A good correlation was observed between ASS1 methylation, low ASS1 mRNA, absence of ASS1 protein expression and sensitivity to ADI-PEG20 in malignant lymphoid cell lines. We confirmed that the demethylating agent 5-Aza-dC reactivated ASS1 expression and rescued lymphoma cell lines from ADI-PEG20 cytotoxicity. ASS1-methylated cell lines exhibited autophagy and caspase-dependent apoptosis following treatment with ADI-PEG20. In addition, the autophagy inhibitor chloroquine triggered an accumulation of light chain 3-II protein and potentiated the apoptotic effect of ADI-PEG20 in malignant lymphoid cells and patient-derived tumour cells. Finally, a patient with an ASS1-methylated cutaneous T-cell lymphoma responded to compassionate-use ADI-PEG20. In summary, ASS1 promoter methylation contributes to arginine auxotrophy and represents a novel biomarker for evaluating the efficacy of arginine deprivation in patients with lymphoma.

Effect of vitamin A content in cafeteria diet on the expression of nuclear receptors in rat subcutaneous adipose tissue.

The aim of this study was to determine the effects of cafeteria diet containing control or elevated level of vitamin A on the expression of nuclear receptors in adipose tissue. Male Wistar rats were submitted to 3 experimental diets during 8 weeks, a standard diet and two hyper-energetic, hyperlipidic "cafeteria" diets containing normal (Caf) or higher (Caf+) vitamin A level. During the experiment, body weights and energy intakes were measured. At the end of the experimental period, subcutaneous adipose tissue (Swat) and all the fat mass were removed and weighted. Nuclear receptors mRNA levels of RARalpha, RARgamma, RXRalpha, PPARgamma were measured in the Swat by a real-time semi-quantitative RT-PCR method. We observed that energy intake of Caf+ and Caf groups was significantly higher than that of the control group. Despite a higher increase of the energy intake in the Caf group compared to the Caf+ group, no significant difference was observed in the body weight gain of the Caf+ compared to the Caf group. The Caf+ and Caf diets led to a significant increase of adipose tissue in cafeteria groups as observed in the Swat depot. The mRNA levels of PPARgamma and RXRalpha were significantly increased in the Caf+ group as compared to control group, with a significant positive correlation between these two parameters. Expressions of RARalpha and RARgamma were not modified in experimental groups compared to controls. In conclusion, 8-week exposure to cafeteria diets with normal and higher levels of vitamin A led to an increase of adiposity in rats, associated, only in the group fed with the higher vitamin A level cafeteria diet, with an increase of PPARgamma and RXRalpha expressions in subcutaneous adipose tissue.

Effect of vitamin A content in cafeteria diet on the expression of nuclear receptors in rat subcutaneous adipose tissue

The aim of this study was to determine the effects of cafeteria diet containing control or elevated level of vitamin A on the expression of nuclear receptors in adipose tissue. Male Wistar rats were submitted to 3 experimental diets during 8 weeks, a standard diet and two hyper-energetic, hyperlipidic "cafeteria" diets containing normal (Caf) or higher (Caf+) vitamin A level. During the experiment, body weights and energy intakes were measured. At the end of the experimental period, subcutaneous adipose tissue (Swat) and all the fat mass were removed and weighted. Nuclear receptors mRNA levels of RARa, RARg, RXRa, PPARg were measured in the Swat by a real-time semi-quantitative RT-PCR method. We observed that energy intake of Caf+ and Caf groups was significantly higher than that of the control group. Despite a higher increase of the energy intake in the Caf group compared to the Caf+ group, no significant difference was observed in the body weight gain of the Caf+ compared to the Caf group. The Caf+ and Caf diets led to a significant increase of adipose tissue in cafeteria groups as observed in the Swat depot. The mRNA levels of PPARg and RXRa were significantly increased in the Caf+ group as compared to control group, with a significant positive correlation between these two parameters. Expressions of RARa and RARg were not modified in experimental groups compared to controls. In conclusion, 8-week exposure to cafeteria diets with normal and higher levels of vitamin A led to an increase of adiposity in rats, associated, only in the group fed with the higher vitamin A level cafeteria diet, with an increase of PPARg and RXRa expressions in subcutaneous adipose tissue

El objetivo del presente trabajo consiste en determinar las consecuencias de un alto contenido en vitamina A en dieta de cafetería sobre la expresión de receptores nucleares en el tejido adiposo. Así, ratas macho Wistar se dividieron en tres grupos : Durante 8 semanas, el grupo control se alimentó con pienso estándar, mientras que los grupos tratados recibieron una dieta rica en grasa (dieta de cafetería) enriquecida (Caf+) o no (Caf) con vitamina A. El peso corporal y la ingesta se determinaron durante todo el experimento. Al final del tratamiento, se pesó el tejido adiposo subcutáneo (Swat) y las otras reservas de grasa. Los niveles de ARNm de los receptores nucleares RARa, RARg, RXRa, PPARg se determinaron en el Swat con un método semi-cuantitativo de RT-PCR en tiempo real. Las ingestas energéticas de los grupos Caf+ y Caf fueron significativamente mayores que las del grupo control. A pesar del aumento en la ingesta del grupo Caf respecto del Caf+, no se observaron diferencias significativas en el aumento de peso corporal entre ambos grupos. Además, las dietas de los grupos Caf+ y Caf provocaron un claro aumento del tamaño de las reservas de grasa, incluido el peso del Swat. Los niveles de ARNm de PPARg yRXRa se incrementaron significativamente en el grupo Caf+ respecto del control, con correlación positiva entre ambos. En cambio, no se modificó la expresión de RARa y RARg. En suma, 8 semanas de alimentación con dieta de cafetería con niveles normales o elevados de vitamina A dan lugar a aumento de la adiposidad en la rata, asociado con aumento de la expresión de PPARg y RXRa en el tejido adiposo subcutáneo solo en el grupo que recibió suplemento de vitamina A

Hyperlipidic diets induce early alterations of the vitamin A signalling pathway in rat colonic mucosa.

OBJECTIVE: Dietary factors can be associated with colorectal cancer. Fatty acids modulate gene expression in various tissues, mediated by activation of the peroxisome proliferator activated receptor: PPAR. Vitamin A signalling is mediated by retinoic acid (RA) receptors (RAR) and retinoid X receptors (RXR). The steroid nuclear receptors PPAR, RAR, RXR, are DNA-binding proteins and they induce gene transcription upon activation by specific ligands and interacting with distinct promoter sequences in the target genes. The aim of this study was to investigate the impact of hyperlipidic diets on the expression of PPARg, RXRa and RARb mRNA in rat colon. METHODS: Rats were fed during 4 weeks with the following diets: a cafeteria diet where 60% of the energy was supplied as lipids and a high fat diet (HFD) represented by 25% of a safflower oil (w/w) rich in polyunsaturated fatty acids, mainly n-6. Nuclear receptors mRNA were quantified by realtime RT-PCR with TaqMan probe process or SYBRGreen I chemical. RESULTS: The cafeteria diet and the HFD induced a significant decrease in RARb mRNA: -36% (p<0.02) and -64% (P<0.001) respectively. Simultaneously, an increased expression of PPARg mRNA was observed for cafeteria diet +35% (P<0.05) and for HFD +45% (P<0.05). The level of RXRa mRNA was significantly increased for cafeteria diet: +53% (P<0.0002), while no significant difference in RXRa mRNA was observed in colonic mucosa rats whose fed with the 25% HFD. CONCLUSIONS: These results showed that an hyperlipidic diet could induce early modifications in the pattern of expression of nuclear receptors in rat colon. Many mechanisms could be probably involved but one hypothesis is that a modification of the balance between the nuclear receptors, resulting from an increased expression of PPARg, could induce a decreased expression of RARb in rat colon.

Discrimination of osteoarthritic and rheumatoid human synovial cells in culture by nuclear image analysis.

Rheumatoid arthritic (RA) and osteoarthritic (OA) synovial cells in culture differ in their metabolic and proliferative behaviour. To assess links between these properties and nuclear changes, we used image analysis to study chromatin texture, together with nuclear morphometry and densitometry of OA and RA cells in primary culture. Chromatin pattern at the third day (D3) was heterogeneous and granular with chromatin clumps whereas at the final stage (D11) of culture a homogeneous and finely granular chromatin texture was observed. This evolution indicates global chromatin decondensation. These characteristics were more marked for RA than for OA nuclei. At each culture time, RA nuclei could be discriminated with high confidence from OA ones from parameters evaluating the organization of the chromatine texture. Nuclear image analysis is thus a useful tool for investigating synovial cell biology.

Methodological considerations on Feulgen-staining applied to cells in primoculture: the model of osteoarthritic synovial cells.

Human synovial cells in primoculture are an interesting model for the study of articular joint diseases and anti-rheumatic drugs. Based on results obtained by image cytometry of Feulgen-stained nuclei, we describe the heterogeneity of synovial cell populations and their progression during culture time in primoculture. Using the hydrolysis properties of the Feulgen reaction and their variations dependent on fixatives, we demonstrate the high acid-lability of the condensed chromatin observed in short term cultured nuclei compared to the acid-resistance of decondensed chromatin in long term cultured nuclei; these variations being probably induced by modifications in the molecular supra-organisation of chromatin during the aging of a culture. Finally, due to the cellular heterogeneity of the biological model and its evolution during culture progression, technical compromises are proposed to obtain optimal Feulgen staining, using Böhm-Sprenger fixative and a 1 h hydrolysis by 6 M HCl at 20 degrees C.

Influence of the molecular structure of steroids on their ability to interrupt gap junctional communication.

17 beta-estradiol propionate was found to reduce the gap junctional communication in a concentration range similar to that of testosterone propionate, in primary cultures of rat Sertoli cells and cardiac myocytes. Uncoupling was reversible on washing out and occurred without concomitant rise in the intracellular calcium concentration. Esterification was prerequisite for the activity of extracellularly applied steroid compounds (for example, testosterone was ineffective even at external concentrations up to 100 microM, whereas its intracellular application at 1 microM totally interrupted intercellular communication), but their uncoupling efficiency did not depend on the nature of the ester chain nor on its position on the steroid nucleus. The derivatives of two other androgen hormones (derivatives of the androstane nucleus) were also efficient as junctional uncouplers. Among five steroid molecules belonging to the pregnane family, only one (pregnanediol diacetate) interrupted the junctional communication. Neither cholic acid nor cholesteryl acetate or ouabain showed this effect. Altogether, no correlation with the presence or position of double bonds nor with the trans- or cis-fusion of the A and B rings could be recognized. These results suggest that this reversible, nondeleterious uncoupling effect of steroids is independent of the shape of the molecules and is more probably related to their size and liposolubility, that condition their insertion into the lipid bilayer. Their incorporation into the membrane could disturb the activity of the membrane proteins by a physical mechanism.

[Isolated primary dissection of the right coronary artery]. / Dissection primitive isolée de l'artère coronaire droite.

A 59 year old female had unstable angina threatening infarction, together with Prinzmetal's syndrome in the postero-diaphragmatic area, and a transient second degree atrio-ventricular block. Selective arteriography revealed complete dissection, as an isolated and primary finding, in the right coronary artery. Operation consisted of dividing the coronary artery at the superior limit of the dissection, of suturing together the separated layers of the artery, and finally an end-to-end aorto-coronary bypass. The patient is living without any functional symptoms 15 months after the initial episode, but a follow-up arteriogram showed that separation of the two layers of the dissection was persisting. This condition has caused rapid or sudden death in 47 of the 51 reported cases, often occurring in females at the beginning of pregnancy. This mortality rate justifies early operation as soon as the diagnosis is made by coronary arteriography.

[Phonomechanographic evaluation of the severity of pulmonary stenosis]. / Evaluation phonomécanographique de la sévérité des rétrécissements pulmonaires.

External phonomechanographic methods allow us to obtain a fairly precise estimate of the degree of obstruction to the pulmonary outflow, whether such an obstruction is a single entity or associated with another intracardiac malformation. The greatest precision in this field is obtained with the cases of pure pulmonary stenosis. The findings on phonomechanographic investigation of 54 cases with pulmonary stenosis have been compared with the results of the haemodynamic tests (catheterisation and angiography) as well as with the anatomical findings after a surgical cure had been effected in a certain number of cases. This study has allowed us to pick out five phonomechanographic criteria of severity of pulmonary stenosis: an increased reduplication of the second sound, lenghening of the interval between the Q wave of the electrocardiogram and the maximum portion of the systolic murmur, an increase in Furuta's ratio and also in the amplitude of the jugular venous a-wave as a function of the total height t (the a/t ratio), but most especially the "corrected" Furuta ratio as a function of the length of the ejection phase of the right ventricle, as this has been shown to possess the best correlation with the right ventricular systolic pressures.

[Coronary arteriography in the threatened infarction syndrome]. / La coronarographie dans le syndrome de menace d'infarctus.

Although greater than in patients with stabilised coronary insufficiency, the risks of carrying out coronary arteriography during the period of a threatened infarction are still modest in the hands of an experienced team. In a series of 66 such investigations under these conditions over an 18 month period, there were no deaths as immediate sequelae. Three myocardial infarctions occurred, one of which was probably encouraged by ceasing the propranolol prior to arteriography. Analysis of the results confirms the frequency of lesions affecting all three trunks (47% of cases), the anterior descending branch being the most commonly affected. A collateral circulation was established in 23% of cases. Ventriculography was normal in one third of cases. The mean value for the ventricular ejection fraction was 0.57 over the entire series. In 58% of cases, all the arteriographic and ventriculographic criteria of operability were satisfied. It therefore appears that coronary arteriography is justifiable during the period of threatened infarction; it may be undertaken at an early stage if the angina does not respond to medical treatment. It is always best to carry it out several days after cessation of the pain where possible. The investigation may lead to urgent surgical intervention.

[Radioisotope measurement of left ventricular ejection fraction. Comparison with results of cineangiocardiography]. / Mesure radioisotopique de la fraction d'éjection ventriculaire gauche. Comparaison avec les résultats de la cinéangiocardiographie

The combination of radiocardiography with scintigraphy of the cavities of the heart allows us to calculate the effective systemic ejection fraction. The systolic volume effectively ejected into the systemic circulation is obtained from radiocardiography using Stewart-Hamilton's principle. It is correlated with the left ventricular end-diastolic volume as calculated by planimetry on the scintigraphic record of the cavities of the heart, making due allowance for the geometrical axis of the heart. The ejection fraction measured by this isotope method has been compared with the ejection fraction measured by selective left ventricular cineangiography, both measurements being carried out at an interval of less than 48 hours in 100 patients without valvular disease. The results show an excellent correlation between the two methods (r = 0.94). In the 15 cases where the haemodynamic status of the ventricule appeared to be normal, the ejection fraction meastured by the two techniques was 0.65 +/- 0.04 and 0.66 +/- 0.04 respectively. In those patients with valve disorders with incompetence, the effective systemic ejection fraction is always lower than the total ejection fraction given by cineangiography. The difference corresponds with the fraction of the blood volume which is regurgitated.

[Phonomecanography in recent myocardial infarction. Ventricular mechanic curve]. / Phonomécanographie de l'infarctus du myocarde récent. la courbe mécanique ventriculaire

Repeated recordings were made of the apexcardiogram throughout the first month after myocardial infarction in 30 patients. The classical timed intervals of the systolic wave are open to some criticism. The systolic waveforms are important. In the majority of transmural anterior infarctions there is a rounded appearance to the beginning of the wave which seems to prolong the electromechanical latency, followed by a late systolic bulge, or a domed waveform. This signifies a non-contractile area, and not neccessarily an ectasia. The early diastolic "peaktrough" appearance, found very frequently wherever the necrosis is situated, is indicative of asynergic contraction of the left ventricle. All of the diastolic phases are altered, probably by increased parietal stiffness: the TRI is lengthened; the "F" wave is flattened (and often absent later on in the condition), its duration is shortened over the anterior positions, and it may contain a shallow dip if there is LVF; the stasis wave is very feeble; the "a" wave is large when the infarct is extensive, or when there is LVF, or when there is longstanding hypertension. Enlargement of the "a" wave is especially indicative of a lowering of the performance of the left ventricle.

[Phonomechanography of recent myocardial infarct: the jugulogram and carotidogram]. / Phonomécanographie de l'infarctus du myocarde récent: Le jugulogramme, le carotidogramme

The mechanographic tracings of 30 patients with recent myocardial infarction, usually uncomplicated and without mitral incompetence, were compared with those of two control groups. The jugular venogram was unhelpful; full measurements can be made in only 40% of cases, and the scatter of normal values is too great. The carotid arteriogram is usually of the "arterial" type. Transmission time and the duration of the rising phase were both shortened. These findings result from the high level of peripheral resistance. The mean left systolic time was, except for qB2, abnormal, and remained so at the end of the first month. In this series there were neither maximal changes in J3, nor significant improvement from J12 nor J30. The largest increases in PPE and PPE/LVET were found in cases with large infarcts, and the largest reduction of LVET was in a case with left ventricular failure. There was no particular peculiarity corresponding with the site of the necrosis. Alteration in the ratio PPE/LVET is often a lasting measure of depressed left ventricular function. It would be most useful to study this ratio in the long term and to use it as a measure of recovery of function and as a guide to advising the patient on his future life style.