Etude de la simulation des réactions de polycondensation par la méthode de Monte-Carlo

Simulation of polycondensation reactions using a Monte-Carlo type method is discussed in detail. The process applies to reactions carried out in homogeneous isotropic phase and it is valid regardless of the number of constituents involved, and whatever the reactivity of the individual functions may be. However, it does not include cyclization, and cannot take into account the variation of the reactivity of a given function with conversion. The conditions that should be satisfied are discussed thoroughly; in particular, duplication procedures are introduced at various stages of the reaction to prevent the system from becoming too small at high conversion. The method was tested on two rather simple cases: the self condensation of AB-type monomer molecules, and chain extension involving AA-type macromolecules and BB-type micromolecular coupling agents. The agreement between the simulation and the data of the classical polycondensation theory is very satisfactory.     

Assessment of muscular metabolism in peripheral arterial occlusive disease using31P nuclear magnetic resonance spectroscopy

Summary The behaviour of muscular metabolism was investigated in 10 patients with peripheral arterial occlusive disease stage II at rest and after maximum erometric calf exercise. The intracellular concentrations of phosphocreatine, inorganic phosphate and adenosine triphosphate as well as muscle pH were measured by means of³¹P magnetic resonance spectroscopy and compared with those from a control group. In addition, arteriovenous differences in concentrations of lactate, pyruvate, ammonia, hypoxanthine and alanine in the femoral blood were determined. The fall in intracellular phosphocreatine concentration during exercise was significantly greater in the calf muscles of patients with arterial occlusion than in controls and correlated linearly with the increase in femoral arteriovenous differences in lactate, ammonia and alanine. A significant fall in intracellular pH occurred during muscular activity only in the patient group, but not in the identically exercised control group. The fall in pH correlated closely with the rise in arteriovenous lactate difference in the femoral blood. The intramuscular ATP concentration remained constant throughout the exercise procedure. The behaviour of both the directly and indirectly measured metabolites permits the deduction of activation of the creatine kinase reaction, glycolysis, myokinase reaction and the purine nucleotide cycle during exercise-induced hypoxia in the presence of arterial occlusive disease. The anaerobic production of energy is sufficient to maintain the ATP concentration even during claudication pain. Magnetic resonance spectroscopy proved to be a useful tool for non-invasive assessment of the metabolic changes in peripheral arterial occlusive disease.Abbreviations ADP adenosine diphosphate - AMP adenosine monophosphate - ATP adenosine triphosphate - AVD arteriovenous difference - (P)AOD (peripheral) arterial occlusive disease - IMP inosine monophosphate - NAD⁺ nicotinamide adenine dinucleotide - PCr phosphocreatine - P₁ inorganic phosphate - ³¹P-MRS ³¹phosphorus magnetic resonance spectroscopy - NMR nuclear magnetic resonance - PNC purine nucleotide cycle     

Light chain heterogeneity of type lambda anti-streptococcal group A-variant polysaccharide antibodies in rabbits.

BASILEA rabbits lack the expression of chi-polypeptide chains and compensate for this lack with expression of lambda polypeptide light chains. These rabbits were immunized with streptococcal group A-variant vaccines. The heterogeneity of the lambda polypeptide chains of specific antibodies was analyzed and compared with that of chi light chains. No significant difference was found by SDS-PAGE and IEF in the number of light chain bands of high affinity antibody expressed. This suggests that in the rabbit the size of the variable region repertoire is similar for lambda and chi light chains.     

Gentle vs. aversive handling of pregnant ewes: II. Physiology and behavior of the lambs

We compared the effects of aversive and gentle handling in late pregnant ewes on fearfulness, heart rate variability and spatial learning in lambs. Twenty-four Norwegian-Dala ewes were studied. Ewes were subjected to gentle (i.e. soft talking and calm behavior) or aversive handling (i.e. swift movements and shouting) for 10 min twice a day during the last five weeks of pregnancy. Lambs from aversively (AVS) or gently (GEN) treated ewes were tested at 4 weeks of age. Lamb behavior was recorded during a) a human approach test, composed of 4 min of isolation and 4 min of exposure to an unfamiliar human, b) an umbrella startle test followed by 5-min recording, and c) two repetitions of a maze test. In addition, heart rate variability was recorded telemetrically before and after the human and startle tests. The baseline heart rate variability measures suggested a lower influence of vagal stimulation in AVS lambs. In the human approach test, AVS lambs vocalized and explored the environment less, and were slower to approach the human. They also tended to have higher flight distances during the startle test than the GEN lambs. The prenatal treatment had no significant effect in the maze test. In conclusion, we showed that aversive handling of pregnant ewes increased fearfulness and reduced vagal tone in their progeny compared to GEN lambs. These effects can have consequences for how lambs cope with rearing conditions.     

Development of l-CDB-4022 as a nonsteroidal male oral contraceptive: induction and recovery from severe oligospermia in the adult male cynomolgus monkey (Macaca fascicularis)

The present study was undertaken to examine the antispermatogenic effect of l-CDB-4022 in the adult male cynomolgus monkey. Monkeys (four per group) were dosed via nasogastric tube for 7 d with l-CDB-4022 at 12.5 mg/kg.d or vehicle (d 0=first day of dosing). Plasma levels of l-CDB-4022 and its deesterified metabolite were nondetectable prior to treatment and in all vehicle-treated monkeys. Peak levels of l-CDB-4022 and its metabolite were observed at 4 h after dosing with steady-state levels apparent around d 4. Sperm concentration and total sperm per ejaculate were decreased to levels below 1x10(6) sperm/ml or sperm/ejaculate in l-CDB-4022-treated monkeys by d 17 and remained suppressed through wk 6. Sperm motility also declined to 0% for 6 wk. Testicular volume was reduced in l-CDB-4022-treated monkeys through d 21. The left testis and epididymis were removed from all monkeys on d 24. At this time, the most mature germ cells in the seminiferous tubules of testes from l-CDB-4022-treated monkeys were either spermatocytes or round spermatids. Immature germ cells, but not mature sperm, were found in the efferent ducts and collapsed epididymal lumen of l-CDB-4022-treated monkeys. A steady recovery in sperm motility, concentration, and total sperm per ejaculate was observed in l-CDB-4022-treated monkeys such that these parameters were not different from those of vehicle-treated monkeys by wk 16. Volume of the remaining testis increased in vehicle- and l-CDB-4022-treated monkeys after hemicastration; however, the increase in l-CDB-4022-treated monkeys was delayed compared with that observed in the vehicle-treated monkeys. The morphology of the remaining testis and epididymis, which were removed on wk 17, was normal. Serum inhibin B levels were increased in l-CDB-4022-treated monkeys during the dosing interval; thereafter serum inhibin B levels declined such that there was no difference between the groups by wk 3. l-CDB-4022 treatment did not affect circulating levels of testosterone, LH, FSH, or estradiol. In conclusion, these data indicate that in the cynomolgus monkey, a representative higher primate, l-CDB-4022 exerts a selective antispermatogenic action, which was reversible under the conditions of this study and thus has potential as a nonhormonal oral male contraceptive.     

The effects of the NMR shift-reagents Dy(PPP)2, Dy(TTHA) and Tm(DOTP) on developed pressure in isolated perfused rat hearts. The role of shift-reagent calcium complexes

The 23Na NMR shift-reagent complexes (Dy(PPP)2, Dy(TTHA), and Tm(DOTP)) bind stoichiometric amounts of Ca2+. Thus, in perfused rat heart systems, a supplementation of Ca2+ is required to maintain the requisite extracellular free calcium concentration ([Ca(o)]f) and to approximate a physiological level of contractile function. The amount of reagent-bound Ca2+ in a heart perfusate that contains a shift-reagent depends on: (1) Ca2+ binding by excess ligand used during the preparation of the shift-reagent; and (2) the Ca2+ binding affinity of the shift-reagent. To address point 1), we introduced a 1H and 31P NMR spectroscopic titration method to quantify directly the concentration of the excess ligand. We also used this method to minimize the amount of excess ligand (L) and thus the amount of Ca*L complex. To address point (2), we determined the stepwise Kd (microm) values of the Ca complexes of the three shift-reagents.: Dy(PPP)2, Kd=0.09, Kd2=7.9; Dy(TTHA), Kd1=10.66, Kd2=10.12; and Tm(DOTP), K(d1)=0.502, Kd2=4.98. The Kd values of the Ca complexes of the phosphonate and triphosphate based shift-reagents, Tm(DOTP) and Dy(PPP)2, respectively, are lower than those of the polyaminocarboxylate-based Dy(TTHA), indicating stronger Ca binding affinities for the former two types of complexes. We have also shown a positive correlation between [Ca(o)]f and left ventricular developed pressure (LVDP) in perfused rat hearts. Dy(TTHA) has shown no effect on LVDP v[Ca(o)]f. The LVDP values in the presence of the phosphonate and triphosphate based shift-reagents, however, were significantly higher than expected from the [Ca(o)]f levels alone. Thus a positive inotropic effect, independent of [Ca(o)]f, is evident in the presence of Tm(DOTP) or Dy(PPP)2.     

Tissue engineered bone grafts based on biomimetic nanocomposite PLGA/amorphous calcium phosphate scaffold and human adipose-derived stem cells

For tissue engineering of critical size bone grafts, nanocomposites are getting more and more attractive due to their controllable physical and biological properties. We report in vitro and in vivo behaviour of an electrospun nanocomposite based on poly-lactic-co-glycolic acid and amorphous calcium phosphate nanoparticles (PLGA/a-CaP) seeded with human adipose-derived stem cells (ASC) compared to PLGA. Major findings were that cell attachment, three-dimensional ingrowth and proliferation were very good on both materials. Cell morphology changed from a spindle-shaped fibroblast-like form to a more roundish type when ASC were seeded on PLGA, while they retained their morphology on PLGA/a-CaP. Moreover, we found ASC differentiation to a phenotype committed towards osteogenesis when a-CaP nanoparticles were suspended in normal culture medium without any osteogenic supplements, which renders a-CaP nanoparticles an interesting osteoinductive component for the synthesis of other nanocomposites than PLGA/a-CaP. Finally, electrospun PLGA/a-CaP scaffold architecture is suitable for a rapid and homogenous vascularisation confirmed by a complete penetration by avian vessels from the chick chorioallantoic membrane (CAM) within one week.     

Steroid hormonal regulation of growth, prostate specific antigen secretion, and transcription mediated by the mutated androgen receptor in CWR22Rv1 human prostate carcinoma cells

CWR22Rv1 (22Rv1) is an androgen-responsive human prostate carcinoma cell line derived from a primary prostate tumor that expresses mutant (H874Y) androgen receptors (AR) and secretes low levels of prostate specific antigen (PSA). In this study, we examined the effects of various androgens and other steroid hormones on proliferation of 22Rv1 cells, PSA secretion, and transactivation. Incubation of 22Rv1 cells with various concentrations of testosterone resulted in a dose-dependent 50-80% increase in growth over 72 h. PSA release and transactivation of PRE2-tk-LUC in 22Rv1 cells were stimulated by low concentrations of natural and synthetic androgens (EC(50)s = 10(-10) to 10(-9)M) and a broad range of other classes of steroid hormones, albeit with lower potency. Uniform positive immunocytochemical staining was observed in 22Rv1 cell nuclei with mouse monoclonal antibodies to human AR. Competitive binding assays indicated that the mutant AR in 22Rv1 cytosol is more promiscuous than a wild-type AR (ARLBD: rat AR ligand binding domain). Testosterone (10(-8)M)-induced PSA release and transactivation were blocked by both antiandrogens and antiprogestins with IC(50)s of 10(-7) to 10(-6)M. At high concentration (10(-6)M), these antagonists showed partial agonist activity in terms of PSA secretion but not transactivation. In conclusion, the mutant AR in 22Rv1 cells binds and responds to low levels of androgens and a wide spectrum of other natural and synthetic steroid hormones, mechanisms proposed to contribute to tumor progression following androgen ablation.     

Proliferation of ASC-derived endothelial cells in a 3D electrospun mesh: Impact of bone-biomimetic nanocomposite and co-culture with ASC-derived osteoblasts

Background

Fractures with a critical size bone defect are associated with high rates of delayed- and non-union. The treatment of such complications remains a serious issue in orthopaedic surgery. Adipose derived stem cells (ASCs) combined with biomimetic materials can potentially be used to increase fracture healing. Nevertheless, a number of requirements have to be fulfilled; in particular, the insufficient vascularisation of the bone constructs. Here, the objectives were to study the impact of ASC-derived osteoblasts on ASC-derived endothelial cells in a 3D co-culture and the effect of 40 wt% of amorphous calcium phosphate nanoparticles on the proliferation and differentiation of ASC-derived endothelial cells when present in PLGA.

Materials and methods

Five primary ASC lines were differentiated towards osteoblasts (OBs) and endothelial cells (ECs) and two of them were chosen based on quantitative PCR results. Either a mono-culture of ASC-derived EC or a co-culture of ASC-derived EC with ASC-derived OB (1:1) was seeded on an electrospun nanocomposite of poly-(lactic-co-glycolic acid) and amorphous calcium phosphate nanoparticles (PLGA/a-CaP; reference: PLGA). The proliferation behaviour was determined histomorphometrically in different zones and the expression of von Willebrand Factor (vWF) was quantified.

Results

Independently of the fat source (biologic variability), ASC-derived osteoblasts decelerated the proliferation behaviour of ASC-derived endothelial cells in the co-culture compared to the mono-culture. However, expression of vWF was clearly stronger in the co-culture, indicating further differentiation of the ASC-derived EC into the EC lineage. Moreover, the presence of a-CaP nanoparticles in the scaffold slowed the proliferation behaviour of the co-culture cells, too, going along with a further differentiation of the ASC-derived OB, when compared to pure PLGA scaffolds.

Conclusions

This study revealed significant findings for bone tissue-engineering. Co-cultures of ASC-derived EC and ASC-derived OB stimulate each other's further differentiation. A nanocomposite with a-CaP nanoparticles offers higher mechanical stability, bioactivity and osteoconductivity compared to mere PLGA and can easily be seeded with pre-differentiated EC and OB.