Position paper on advancing sickle cell disease management in France by bridging the clinical practices and guidelines through expert insights.

In France, sickle cell disease (SCD) is the most common rare disease and represents the most prevalent genetic disorder, with 19,800 to 32,400 patients diagnosed in 2016 and 1:714 newborns affected in 2019. SCD is caused by a single mutation in the ß-globin gene, resulting in the production of abnormal hemoglobin (called HbS), chronic hemolytic anemia, and impaired red blood cell rheology. SCD patients face several severe acute and chronic complications, including stroke, acute chest syndrome (ACS), painful vaso-occlusive crisis (VOC), organ failure, and a high risk of infections. As patients' care pathway remains unclear in France, a roundtable advisory board meeting was organized in the country to provide insights into the management of SCD in alignment with clinical guidelines. The meeting brought together a panel of esteemed key opinion leaders (KOLs) in SCD management, encompassing both clinical practice and research. During the meeting, the KOLs discussed clinical practices and their alignment with French guidelines, identifying areas of concordance and discrepancy. They also addressed disparities in SCD clinical practices across regions and medical centers. The KOLs discussed the prophylactic and therapeutic options currently available for SCD patients in France, with a focus on transfusion therapies, especially automated red blood cell exchange (aRBCX). The results of this advisory board meeting provide a valuable platform for gathering expert perspectives on SCD management, clinical practices, guideline alignment, and the potential for contributions to guideline updates.

[1-14C]Arachidonic acid incorporation into glycerolipids and prostaglandin synthesis in peritoneal macrophages: effect of chloramphenicol.

Peritoneal macrophages from normal mice were labelled with [1-14C]arachidonic acid after 2 h culture. The uptake of arachidonic acid into cellular lipids was rapid, time-dependent and can be represented within the limit of the studied times by a parabolic regression. Indomethacin decreased the kinetics of uptake; this inhibition is dose-dependent. Chloramphenicol had no effect on macrophage [1-14C]arachidonic acid uptake. After 3 h, the radioactivity was recovered in phosphatidylcholine (38.6%), phosphatidylserine-phosphatidylinositol (8.5%), phosphatidylethanolamine (22.1%), diacylglycerol (2.9%), triacyglycerol (2%) and cholesteryl ester (11.8%). Chloramphenicol and indomethacin inhibited the labelling of phospholipids and stimulated the labelling of neutral lipids and cholesteryl ester. Studies on arachidonic acid release from glycerolipids of prelabelled 2-h cultured macrophages showed that phosphatidylcholine and phosphatidylserine-phosphatidylinositol are the major source of arachidonic acid in prostaglandin synthesis in macrophages stimulated or not by zymosan. Chloramphenicol inhibited release of fatty acid from phosphatidylcholine and phosphatidylserine-phosphatidylinositol; indomethacin had no effect. Both drugs inhibited prostaglandin synthesis in stimulated or non-stimulated macrophages. In the culture medium, indomethacin increased the release of free arachidonic acid by stimulated macrophages. Possible explanations for the mechanisms underlying these effects are presented. It is concluded that indomethacin and chloramphenicol exert profound effects on the metabolism of phospholipids and its zymosan activation. Chloramphenicol appears to impair prostaglandin synthesis through several mechanisms and especially through phospholipase inhibition.

IL-13 increases the cPLA2 gene and protein expression and the mobilization of arachidonic acid during an inflammatory process in mouse peritoneal macrophages.

Pretreatment of mouse peritoneal macrophages with interleukin-13 (IL-13) potentiates the mobilization of arachidonic acid (AA) and the production of HETEs but does not affect the production of cyclooxygenase metabolites triggered by the suboptimal concentration of an inflammatory agonist (opsonized-zymosan). Cycloheximide suppresses these effects of IL-13 suggesting that de novo protein synthesis is involved. Indeed, IL-13 induces a time-dependent increase in the levels of cytosolic PLA2 (cPLA2) protein and mRNA. This study demonstrates a new pathway for IL-13 to modulate the inflammatory process in macrophages via modifications of cPLA2 expression and subsequent AA mobilization.

Effect of interleukin-4 on allergen-induced arachidonic acid metabolism of rat peritoneal macrophages during immediate hypersensitivity reactions.

The aim of this study was to investigate the [3H]arachidonic acid metabolism of rat peritoneal macrophages, induced by allergen (ovalbumin) and the impact of interleukin-4 on this process. We established that ovalbumin induces an increase of [3H]arachidonic acid mobilisation from membrane lipids and of [3H]arachidonic acid catabolism, principally by the 5-lipoxygenase pathway, when the macrophages are sensitized and when serum is present. The allergen effect is not modified by the presence of interleukin-4 in the culture medium of macrophages 15 h before the allergen challenge. We also showed that, whereas the basal [3H]arachidonic acid metabolism of macrophages from control and actively sensitized rats is not different, interleukin-4 increases the [3H]arachidonic acid mobilisation and catabolism by cyclooxygenase and 5-lipoxygenase pathways in macrophages from control rats although it does not in macrophages from actively sensitized rats. In macrophages from control rats, the interleukin-4 effect is diminished by the addition of IgEs to their culture medium. In summary, interleukin-4 has an enhancer effect on the macrophage arachidonic acid catabolism that depends on the sensitization condition of the cell but that has no consequences on the further increased arachidonic acid metabolism induced by the allergen.

Distribution of radioactivity following oral administration of carcinogenic 14CH3-labelled nitrosocarbaryl in the rat.

After a single intragastric administration of 14C-labelled carcinogenic nitrosocarbaryl, a nitrosated pesticide, the distribution of radioactivity was investigated in the blood and a number of organs in male rats. The animals received 0.25 mg/kg of labelled nitrosamine and were killed following administration at timed intervals between 0.5 h and 24 h. Our results show that the greatest amount of the 14CH3--group was associated with the forestomach, tumor-susceptible tissue; the level of radioactivity is noteworthy but less important in the glandular stomach. There are also sites of radioactivity accumulation mainly in the liver. Moreover, [14C]nitrosocarbaryl was revealed in the blood suggesting that nitrosamine itself rapidly (0.5 h) crosses the intestinal barrier and in a significant quantity (13%). These facts constitute a potential carcinogenic risk.

The effects of locally injected antibiotic on carrageenan-induced granuloma in rats.

Indomethacin (0.5 mg/100 g b.w./day) and chloramphenicol (0.5 mg or 15 mg/100 g b.w./day) were tested for their anti-inflammatory effects on 7th day carrageenan-induced granuloma formation. Neither of the drugs modified granuloma or pouch wall weight but they decreased the exudate and the cluster of dead cells. Indomethacin and chloramphenicol decreased glucosamine in the dead cell granuloma fraction and increased the level of collagen in the pouch wall. The drugs differed in their inhibitory effect on lysozyme and prostaglandin E2 accumulation in the exudate. The increase in collagen was related to a drop in the level of prostaglandin E2 which seems to regulate collagen deposition in the granuloma. However, the prostaglandin E2-lysozyme correlation--which was only significant with chloramphenicol--suggests a mode of action for chloramphenicol different from that of indomethacin. Chloramphenicol could act by a myelodepressive and/or chemotactic effect. The effects of chloramphenicol on the macrophages are discussed.

Contrasting effect of interleukin-4 on the expression of cytosolic phospholipase A2, 5-lipoxygenase and 5-lipoxygenase-activating protein in peritoneal macrophages from control and ovalbumin-sensitized rats.

Interleukin-4 (IL-4), which has been widely described as an anti-inflammatory cytokine, can also exert proinflammatory effects. In this study, we extend these findings to demonstrate, in an allergic model, the dual effect of IL-4 on arachidonic acid (AA) metabolism in macrophages. In peritoneal macrophages from control rats (cPM), IL-4 had no effect on cPLA2 and 5-LO expression, but increased FLAP expression without affecting basal and A23187- or PMA-challenged arachidonic acid (AA) metabolism. In contrast, in peritoneal macrophages from ovalbumin-sensitized rats (sPM), IL-4 decreased cPLA2, 5-LO and FLAP expression and PMA-challenged eicosanoid production. A23187-challenged AA metabolism of sPM was not affected by IL-4 pretreatment. Thus, IL-4 acts differently on cPLA2, 5-LO and FLAP expression and AA metabolism in peritoneal macrophages depending on their resident or sensitization-induced differentiated status.

Lipid peroxidation in rat liver microsomes: influence of carcinogenic N-nitrosocarbaryl.

The reactivities of carbaryl, N-methyl 1-naphthylcarbamate insecticide and its N-nitrosated derivative carcinogenic, N-nitrosocarbaryl, were investigated on the microsomal hepatic lipid peroxidation and NADPH-dependent reductase activities. The in vivo treatment by N-nitrosocarbaryl produced a reduction in lipoperoxidative degradation induced in vitro by NADPH with regard to the formation of malonaldehyde and conjugated dienes. Carbaryl, its precursor did not affect lipid peroxidation under the same in vivo conditions. Moreover, following administration of the 2 compounds, the activities of NADPH-cytochrome c reductase as well as NADPH-neotetrazolium reductase were significantly decreased by N-nitrosocarbaryl but not influenced by carbaryl. Correspondingly, in vitro studies were performed; different action patterns of the 2 tested xenobiotics were also noted after treatment of rat liver microsomes in vitro by carbaryl and N-nitrosocarbaryl especially in their ability to cope with microsomal oxygen activation. N-Nitrosocarbaryl proved to have a potent inhibitor concentration effect on NADPH-dependent chemiluminescence response in vitro; carbaryl was virtually ineffective on this parameter. No significant difference appeared in the affinity of N-nitrosocarbaryl and carbaryl for the microsomal phospholipids. From the in vitro explorations, it was suggested that carcinogenic N-nitrosocarbaryl may be involved in the inhibition mechanism of microsomal lipid peroxidation through decreases in both NADPH-dependent reductase activities and superoxide generation.

Inhibition of two rat hepatic microsomal drug-metabolizing enzymes by a carcinogenic N-nitrosated pesticide: N-nitrosocarbaryl.

N-Nitrosocarbaryl (N-methyl-1-naphthyl N-nitrosocarbamate) was intraperitoneally administered to male and female rats on four consecutive days at the following doses: 6.25 mg, 12.5 mg, 25 mg and 50 mg/kg body weight/day in olive oil solution; the controls received just the oil. In a second experiment, a daily intraperitoneal dose of 25 mg/kg of N-nitrosocarbaryl was given for 1, 2, 3 or 4 days; the animals were killed 24 h after the last treatment. The two following microsomal enzymatic activities were assayed: aniline aromatic hydroxylase and p-nitroanisole O-demethylase; the levels of cytochrome P-450, proteins and RNA were measured in the hepatic microsomal fraction. N-Nitrosocarbaryl is an inhibitor of the two investigated microsomal monooxygenases at doses of 25 and 50 mg/kg when administered on 4 consecutive days. During the daily administration, enzyme inhibition is seen in females after one day of treatment whereas cytochrome P-450 only becomes lowered after 4 days of administration. In males, no modification of this parameter is observed whereas the activities of microsomal monooxygenases are inhibited. These results suggest that N-nitrosocarbaryl could act on the active sites of the enzymes which metabolize aniline and p-nitroanisole.

Interaction of carbaryl and N-nitrosocarbaryl with microsomal monooxygenase activities.

The in vitro interactions of carbaryl and carcinogenic N-nitrosocarbaryl with rat liver microsomal monooxygenase activities are compared. The inhibitory effect of the nitroso-compound is demonstrated to be non-competitive on aminopyrine N-demethylase, p-nitroanisole O-demethylase and aniline hydroxylase. The nature of the inhibition induced by the parent amide is found to be competitive on aminopyrine N-demethylase and p-nitroanisole O-demethylase. Correspondingly, in vitro studies of the metabolism of the two compounds were carried out: they both yield formaldehyde. Moreover, N-nitrosocarbaryl is denitrosated through a NO-cytochrome P-450 complex during microsomal metabolism. The toxic effects and biological activities of the two compounds are discussed on the basis of data of metabolic studies and different patterns of enzyme inhibition.

[The influence of dietary regimen on the stimulation of microsomal monooxygenases in rat liver induced by 2,3-dimethylquinoxaline]. / Influence des régimes alimentaires sur la stimulation des monooxygénases microsomales du foie de rat induite par la diméthyl-2,3 quinoxaline

Activities, on the hepatic microsomal fraction, of the following enzymes: aniline aromatic hydroxylase, p-nitroanisole O-demethylase, aminopyrine N-demethylase, N-methylaniline N-demethylases, as well as the P-450 cytochrome level, have been evaluated on female rats. During a period of 26 days, they are fed on 4 diets including either: 20% or 30% proteins, or: 1% or 30% lipids. A parallel study is carried out: animals fed on the same diets are given, orally on the 22nd day and for 4 days, a bactericide, the dimethyl-2.3 quinoxaline; the dose is 500 mg/kg/day, with an interval of 24 hours. 1) A stimulation of the microsomal monooxygenase activity and an increase in the P-450 cytochrome level are induced by the dimethyl-2.3 quinoxaline on all animals regardless the diet they are on. 2) The hyperlipidic diet is the only one to induce an increased activity of the same enzymes, but the P-450 cytochrome level remains unchanged. 3) In rats fed on the hyperlipidic diet and treated with dimethyl-2.3 quinoxaline, the effects of this substance and of this diet, on the drug-metabolizing enzymes, become additive.

[Effect of ingestion of derivatives of quinoxaline on the enzymatic detoxication activity of hepatic microsomes in the rat]. / Effet de l'lngestion de dérivés de la quinoxaline sur les activités de détoxication enzymatique des microsomes hépatiques du rat

Female rats received quinoxaline, 2,3-dimethyl quinoxaline, 2,3-diphenyl quinoxaline and 2,3-dithiol quinoxaline by oral intubation for 4 days (500 mg/kg/day) and drug-metabolizing enzymes were investigated. A significant stimulation in liver microsomal enzymes occurred after 2,3-dimethyl quinoxaline and 2,3-diphenyl quinoxaline treatments; these changes were associated with increases in cytochrome P-450 level and in RNA and protein contents of the microsomal fraction. Quinoxaline had no effect on the metabolic rate of the studied substrates but increased the cytochrome P-450 and the RNA and protein levels; whereas, 2,3-dithiol quinoxaline inhibited the aminopyrine and N-methylaniline N-demethylations and decreased the cytochrome P-450 per mg microsomal protein.

[Relationship between the phagocytic inhibition of the rat reticuloendothelial system and the anticholinesterasic effect of an insecticide, Carbaryl (author's transl)]. / Relation entre l'inhibition phagocytaire du système réticuloendothélial du rat et l'effet anticholinestérasique d'un insecticide, le carbaryl.

Phagocytic activity of the reticuloendothelial system (RES) and blood cholinesterase activity were determined in male rats after veinous administrations of carbaryl and 1-naphthol, a carbaryl metabolite. The various parameters were measured 1, 24, 48 and 72 hours after administration of the following four doses per 100 g body weight : 1.875, 3.75, 7.5 and 15 mumol. 1. Results showed an inhibition of the RES phagocytic activity (clearance of colloidal carbon) after carbaryl administration; although 1.875 mumol/100 g had no effect, the other doses inhibited RES activity, blockade time being a function of the dose given. The phagocytic function had returned to normal 72 hr after carbaryl administration. 2. Reductions in spleen weight and protein content were observed together with the RES blockade. 3. At all four doses, the anticholinesterase effect was already apparent one hour after carbaryl administration. 4. 1-naphthol, one of carbaryl's chief metabolites, had no effect either on the RES or on the different parameters studied. These results show a relationship between the phagocytic inhibition of the reticuloendothelial system and the anticholinesterasic effect by carbaryl. They suggest an inhibition of some esterases of macrophages interfering with the phagocytosis.

[Phagocytic activity of the reticuloendothelial system in the rat after administration of an anticholinesterasic pesticide, carbaryl (author's transl)]. / Activité phagocytaire du système réticuloendothélial du rat après administration d'un anticholinestérasique, le carbaryl.

The effects of 4 carbaryl doses (0.375, 0.75, 1.50 and 3 mg/100 g) on the reticuloendothelial system (RES) phagocytic activity were studied 1 h after their administration to male rats. Carbaryl reduced RES phagocytic activity. Results showed a dose-dependent drop in RES phagocytic activity. Carbaryl might act as an inhibitor of phagocytes by saturing them to greater or lesser degree, depending on the dose administered.

[Effects of chloramphenicol on microsomal reductases associated with lipoperoxidation (author's transl)]. / Effets du chloramphenicol sur les réductases microsomales liées aux péroxydations lipidiques.

The effects of chloramphenicol treatment to rats, by intraperitoneal way on three consecutive days were studied on various enzymatic systems of hepatic microsomes, implicated in the detoxication reactions non induced or induced by phenobarbital. Chloramphenicol increases the activity of reductases like NADPH cytochrome c reductase and neotetrazolium reductases. It does not appear to modify the activity of the N-demethylase systems or of the cytochrome P 450 level contrarily to phenobarbital. Chloramphenicol decreases the activity of microsomal nitro reductases, the production of malonaldehyde and conjugated dienes in the microsomes currently considered as a reliable estimate of the extent of the lipoperoxidation process. These results suggest that chloramphenicol could act like an antioxdant derivative.

Phagocytic activity of the rat reticuloendothelial system and the pharmacokinetics of an anticholinesterasic insecticide: carbaryl.

1. The pharmacokinetics of [14C]carbaryl administered intravenously and orally were studied in male rats whose reticuloendothelial system (RES) was inhibited by colloidal carbon or activated by glyceryl trioleate. 2. A time course for [14C]carbaryl blood concn. was fitted to a two-compartment open model following single intravenous administration. A single exponential decay was noted following intragastric administration. 3. The constant blood elimination of [14C]carbaryl decreased significantly in animals with the RES inhibited and increased in those whose RES was activated compared to control animals. 4. There was an increase in carbaryl concn. in the tissue compartment in animals with the RES activated, but no change in animals with the RES inhibited. 5. The equivalent [14C]carbaryl concn. of liver and lungs were decreased or increased in animals with the RES inhibited or activated respectively.

Evaluation of cellular and humoral mechanisms of carbaryl-induced reticuloendothelial phagocytic depression.

The simultaneous injection of carbaryl and colloidal carbon phagocytized by the reticuloendothelial cells results in competition between the two substances in favor of the carbon particles. Experiments with opsonized carbaryl suggest that the decrease in carbaryl blood clearance by the colloid is mediated by a depletion of serum opsonins. Following blockade, the liver carbaryl uptake was depressed in the control group (17%), while it was increased in the opsonized group (12%). With all preparations of carbaryl, opsonized or non-opsonized, colloidal carbon produced a slight and variable increase in carbaryl uptake by the spleen and lungs. These results indicate that, besides the uptake of carbaryl by the hepatocytes, other clearance sites must also be considered such as the Kupffer cells and other liver sinusoidal cells. Moreover our results show that intravenous administration of carbaryl induces a state of phagocytic depression as indicated by impaired intravascular phagocytosis and depressed hepatic uptake of the reticuloendothelial (RE)-test colloidal suspension. The results obtained from injection of opsonized colloidal particles during carbaryl-induced RE-depression, and the fact that carbaryl and carbon are both opsonized by the same serum factor, suggest that the mechanisms of RE-blockade involve selective hepatic and splenic macrophage failure and depletion of serum opsonins. According to our enzymatic investigation, this failure of the RE system to incorporate colloids during carbaryl--RE-blockade could be due to a defect in the activity of macrophage membrane-bound serine esterase.

The effect of carbaryl on the arachidonic acid metabolism and superoxide production by mouse resident peritoneal macrophages challenged by zymosan.

Carbaryl, a broad spectrum insecticide with anticholinesterase activity, was tested for its ability to disturb resident peritoneal macrophages stimulated by opsonized zymosan. The effect of carbaryl on superoxide production and on the release of [1-14C] arachidonic acid and 14C-labelled prostaglandins was dose-dependent. For 2.5 X 10(-6) M of carbaryl, superoxide production and prostaglandin release were not significantly inhibited. At 12.5 X 10(-6) M, the inhibitory effect was apparent for superoxide production (33%) and for the release of 6 KPGF1 alpha (60%), PGE2 (42%), PGF2 alpha (38%), PGD2 (33%). Carbaryl had no effect on the level of free arachidonic acid. Insecticide at 12.5 X 10(-6) M significantly decreased the deacylation of the phosphatidylcholine (20%). Incubation of resident peritoneal macrophages with indomethacin studied conjointly decreased only the prostaglandin release. These results suggest that carbaryl decreases the sequence of events following the binding of a particulate agent to its receptor and leading to the induction of phospholipase activity and the subsequent release of 20:4 and the oxidative burst in the cells. The effect of this pesticide on phospholipid metabolism and its consequences on macrophage stimulation are discussed. Ecto-serine esterase inhibition in the effect mechanism of the pesticide was suggested.

Chemiluminescence response and arachidonic acid metabolism of macrophages induced by gamma-hexachlorocyclohexane (lindane).

The influence of gamma-hexachlorocyclohexane (gamma-HCCH) on arachidonic acid (AA) metabolism and oxygen metabolite production was investigated on mouse peritoneal macrophages gamma-HCCH stimulated 6KPGF1 alpha, PGE2, LTC4, LTB4 and HETEs production and increased the luminol-dependent chemiluminescence (CL). Lindane acted synergistically with phorbol ester on prostaglandins-leukotrienes (PGs-LTs) and CL production. Similar stimulation of CL and PGs-LTs production was found after challenge by the calcium ionophore A23187. The implication of calcium mobilization in lindane effects was proposed.