Anomalies des autosomes et infertilité masculine

Spermatogenesis impairment is frequently associated with autosomal rearrangements. A meiotic study was performed on testicular biopsies of men ascertained through primary fertility and found to be heterozygous for reciprocal translocation, Robertsonian translocations, inversions and extra chromosomes. Pairing failure at meiosis as well as intimate associations between rearranged autosomes and the XY bivalent have been discussed as a cause of gametogenic arrest.     

Ferritin conjugates as specific magnetic labels. Implications for cell separation

Concanavalin A coupled to the naturally occurring iron storage protein ferritin is used to label rat erythrocytes and increase the cells' magnetic susceptibility. Labeled cells are introduced into a chamber containing spherical iron particles and the chamber is placed in a uniform 5.2 kG (gauss) magnetic field. The trajectory of cells in the inhomogeneous magnetic field around the iron particles and the polar distributions of cells bound to the iron particles compare well with the theoretical predictions for high gradient magnetic systems. On the basis of these findings we suggest that ferritin conjugated ligands can be used for selective magnetic separation of labeled cells.     

Colorimetric determination of N-hydroxylated metabolites in microsomal studies

A colorimetric method is described which can be used for the routine determination of primary and secondary N-hydroxylamino compounds in drug metabolism studies using microsomal preparations. The assay is carried out on the supernatant obtained after protein precipitation of the incubation mixture. The method is based on the reduction of ferric ion by the hydroxylamino moiety with the resulting ferrous ion being quantitated by coupling with 2,4,6-tripyridyl-s-triazine to form a purple color with a maximum absorbance at 595 nm. The method is specific to primary and secondary hydroxylamino compounds and calibration curves can be obtained in the range of concentrations of 1 to 20 μg/ml. Hydroxamic acid, amido, amino, phenolic, nitro, nitroso, oxime, nitrone, aldehyde, and ketone compounds do not produce any color reaction when analyzed by the same method. The method is simple, rapid, and many samples can be analyzed in a short period of time.     

Pulsed magnetohydrodynamic blood flow in a rigid vessel under physiological pressure gradient

Blood flow in a steady magnetic field has been of great interest over recent years. Many researchers have examined the effects of magnetic fields on velocity profiles and arterial pressure, and major studies have focused on steady or sinusoidal flows. In this paper, we present a solution for pulsed magnetohydrodynamic blood flow with a somewhat realistic physiological pressure wave obtained using a Windkessel lumped model. A pressure gradient is derived along a rigid vessel placed at the output of a compliant module which receives the ventricle outflow. Then, velocity profile and flow rate expressions are derived in the rigid vessel in the presence of a steady transverse magnetic field. As expected, results showed flow retardation and flattening. The adaptability of our solution approach allowed a comparison with previously addressed flow cases and calculations presented a good coherence with those well established solutions.     

Aberrant heteroplasmic transmission of mtDNA in cloned pigs arising from double nuclear transfer

Double nuclear transfer begins with the transfer of nuclear DNA from a donor cell into an enucleated recipient oocyte. This reconstructed oocyte is allowed to develop to the pronuclear stage, where the pronuclei are transferred into an enucleated zygote. This reconstructed zygote is then transferred to a surrogate sow. The genetic integrity of cloned offspring can be compromised by the transmission of mitochondrial DNA from the donor cell, the recipient oocyte and the recipient zygote. We have verified through the use of sequence analysis, restriction fragment length polymorphism analysis, allele specific PCR and primer extension polymorphism analysis that following double nuclear transfer the donor cell mtDNA is eliminated. However, it is likely that the recipient oocyte and zygote mitochondrial DNA are transmitted to the offspring, indicating bimaternal mitochondrial DNA transmission. This pattern of mtDNA inheritance is similar to that observed following cytoplasmic transfer and violates the strict unimaternal inheritance of mitochondrial DNA to offspring. This form of transmission raises concerns regarding the genetic integrity of cloned offspring and their uses in studies that require metabolic analysis or a stable genetic environment where only one variable is under analysis, such as in knockout technology.     

A catalogue of Clostridium thermocellum endoglucanase, β-glucosidase and xylanase genes cloned in Escherichia coli

Abstract Two independent collections of clones containing Clostridium thermocellum genes involved in cellulose have been previously obtained at IAPGR, Cambridge, and at the Pasteur Institute, Paris. The two collections were compared for cross-hybridization, restriction maps and enzyme phenotypes. Truly distinct genes were one β-glucosidase gene, two xylanase genes, and fifteen endogluconase genes. Two of the cloned fragments contained extraneous DNA which was absent from their respective counterparts isolated in the other collection. The dicrepancies resulted from in vivo rearrangements which had occurred in either of the C. thermocellum NCIB 10682 stocks used to generate the two gene banks.     

Inhibition of human pancreatic proteinases by human plasma alpha2-antiplasmin and antithrombin

Human plasma alpha1-antitrypsin inhibits human pancreatic trypsin, chymotrypsin and elastase, which are massively released into the blood stream during acute pancreatitis. To examine whether the plasma proteins of individuals with genetic deficiency of alpha1-antitrypsin are protected against the deleterious action of these enzymes by other inhibitors, we have tested their inhibition by alpha2-antiplasmin and antithrombin. We have determined the inhibition rate constants kass and calculated d(t), the in vivo inhibition time. Surprisingly, trypsin is inhibited faster by alpha2-antiplasmin [kass=2.5 x 10(6) M(-1)S(-1), d(t)=2.3 s] and antithrombin [kass=1.7 x 10(5) M(-1)s(-1), d(t)=5.8 s] than by alpha1-antitrypsin [d(t)=17 s or 116 s in alpha1-antitrypsin-sufficient or alpha1-antitrypsin-deficient individuals, respectively]. Low molecular weight heparin accelerates the inhibition of trypsin by antithrombin by a factor of 16 [d(t)=0.36 s]. Antithrombin and alpha2-antiplasmin are not physiological inhibitors of chymotrypsin and elastase. These enzymes are, however, physiologically inhibited by alpha1-antitrypsin and alpha1-antichymotrypsin even in alpha1-antitrypsin-deficient individuals. We conclude that (i) low molecular weight heparin may be helpful in the management of acute pancreatitis, and (ii) genetically determined alpha1-antitrypsin deficiency probably does not lead to a significantly increased risk of plasma protein degradation during this disease.     

Nature of DNA damage in ejaculated human spermatozoa and the possible involvement of apoptosis

Numerous studies have shown the presence of DNA strand breaks in human ejaculated spermatozoa. The nature of this nuclear anomaly and its relationship to patient etiology is however poorly understood. The aim of this study was to investigate the relationship between nuclear DNA damage, assessed using the TUNEL assay and a number of key apoptotic markers, including Fas, Bcl-x, and p53, in ejaculated human spermatozoa from men with normal and abnormal semen parameters. We also determined the nature of the DNA damage by examining the percentage of ejaculated spermatozoa exhibiting DNA damage using the comet assay and by challenging sperm chromatin to attack by micrococcal nuclease S7 and DNase I. We show that TUNEL positivity and apoptotic markers do not always exist in unison; however, semen samples that had a low sperm concentration and poor morphology were more likely to show high levels of TUNEL positivity and Fas and p53 expression. In addition, the DNA damage in ejaculated human sperm is represented by both single- and double-stranded DNA breaks, and access to the DNA is restricted by the compacted nature of ejaculated spermatozoa. This DNA protection is poorer in men with abnormal semen parameters. We propose that the presence of DNA damage is not directly linked to an apoptotic process occurring in spermatozoa and arises due to problems in the nuclear remodeling process. Subsequently, the presence of apoptotic proteins in ejaculated spermatozoa may be linked to defects in cytoplasmic remodeling during the later stages of spermatogenesis.