Spermatozoa were collected from the rete testis and vas deferens of conscious rams. The endogenous oxygen uptake of the spermatozoa was unaffected by alpha-chlorohydrin added in vitro, although this compound abolished the stimulation of oxygen uptake caused by the addition of glycerol. The metabolism of [14C]glycerol by testicular and epididymal spermatozoa was markedly reduced by alpha-chlorohydrin, CO2 production and lactate accumulation being almost totally inhibited. These effects were dependent upon a period of preincubation of the spermatozoa with alpha-chlorohydrin alone, since the presence of glycerol protected the spermatozoa from its action. Longer exposure and a higher concentration of alpha-chlorohydrin were needed with testicular than with epididymal spermatozoa to achieve a maximal effect. The metabolism of [14C]glucose by both sperm types was also inhibited by alpha-chlorohyrin. Spermatozoa of the ram are therefore susceptible to the action of alpha-chlorohydrin throughout the epididymis, although more mature spermatozoa are more affected. It is suggested that alpha-chlorohydrin is converted to an intermediate which is the agent responsible for the inhibition of glycolysis in spermatozoa.
Glycosylation can be a critical quality attribute in biologic manufacturing. In particular, it has implications on the half-life, immunogenicity, and pharmacokinetics of therapeutic monoclonal antibodies (mAbs), and must be closely monitored throughout drug development and manufacturing. To address this, advances have been made primarily in upstream processing, including mammalian cell line engineering, to yield more predictably glycosylated mAbs and the addition of media supplements during fermentation to manipulate the metabolic pathways involved in glycosylation. A more robust approach would be a conjoined upstream-downstream processing strategy. This could include implementing novel downstream technologies, such as the use of Fc γ-based affinity ligands for the separation of mAb glycovariants. This review highlights the importance of controlling therapeutic antibody glycosylation patterns, the challenges faced in terms of glycosylation during mAb biosimilar development, current efforts both upstream and downstream to control glycosylation and their limitations, and the need for research in the downstream space to establish holistic and consistent manufacturing processes for the production of antibody therapies.
The rate of entry of alpha-chlorohydrin into rat rete testis fluid has been studied using the 14-C and 36-Cl-labelled compounds. The alpha-chlorohydrin crosses the blood-testis barrier and the concentration of radioactivity in rete testis fluid attained blood levels within 45 min. Within 3 hr of a single injection of [14-C] alpha-chlorohydrin, radioactivity was widely distributed in body fluids, and was present in the lipids of the brain, testis, epididymis and epididymal fat pads. No radioactivity was found in tissue lipids following the administration of [36-Cl] alpha-chlorohydrin, which suggests that dechlorination of this compound occurs before its incorporation. Neither a single high dose nor repeated low doses of alpha-chlorohydrin induced changes in the incorporation of [14-C] glycerol into lipids of the brain, testis, epididymis and epididymal fat pads.
Conversion of heavy-aggregate alveolar surfactant (H) to a light-aggregate, nonsurface active form (L) is believed to involve the activity of an enzyme, namely, convertase. This conversion can be reproduced in vitro by the surface-area cycling technique. The purpose of the present study was to use this technique to investigate the developmental aspects of convertase activity in fetal, newborn, and adult rabbits. H was isolated from alveolar lavage from term [31-day gestation (31d)] fetal rabbit pups, 1-, 4-, and 7-day-old newborns, and adults, and the percent conversion to L was determined. To assess lamellar bodies (LB) as a potential source of activity in this species, these structures were isolated from lung tissue of 27-day-gestation (27d) and 31d fetuses, 1-, 4-, and 7-day-old newborns, and adults and were cycled the same as for H. LB contained considerable activity at each developmental stage i.e., approximately 82% of a 27d LB preparation converted to L after 3 h of cycling. In the adult, this value was 78%. Very little conversion of H was obtained from fetal lung (i.e., <20% of the 31d fetal preparation converted to L), but, by postnatal day 4, this value was greatly increased (i.e., >80% conversion) and stayed elevated to adulthood. The activity for each H and LB fraction was temperature and concentration dependent and diminished with storage at 4 degreesC. These data suggest the LB as the source of convertase activity in the rabbit and demonstrate dramatic developmental changes in this activity after release of the LB contents to the alveoli.
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