Resveratrol addition to Chinese hamster ovary cell culture media: The effect on cell growth,monoclonal antibody synthesis,and its chemical modification

The effect of the addition of resveratrol to cell culture media during the production of monoclonal antibodies was investigated. Treatments of Chinese hamster ovary (CHO) cells expressing immunoglobulin G (IgG) with 25 and 50 μM resveratrol showed that resveratrol was capable of slowing cell growth while almost doubling cell-specific productivity to 4.7 ± 0.6 pg IgG/cell·day, resulting in up to a 1.37-fold increase of the final IgG titer. A resveratrol concentration of 50 μM slowed the progression through the cell cycle temporarily by trapping cells in the S-phase. Cation exchange chromatography showed no significant difference in the composition of acidic or basic IgG species and size exclusion chromatography indicated no change in fragmentation or aggregation of the recombinant IgG in the treatment groups. Resveratrol could be used as a chemical additive to CHO media where it would enhance IgG productivity and provide a degree of protection against hydroxyl and superoxide free radicals, expanding the range of options for process improvement available to monoclonal antibody manufacturers.     

The replicative helicase MCM recruits cohesin acetyltransferase ESCO2 to mediate centromeric sister chromatid cohesion

Chromosome segregation depends on sister chromatid cohesion which is established by cohesin during DNA replication. Cohesive cohesin complexes become acetylated to prevent their precocious release by WAPL before cells have reached mitosis. To obtain insight into how DNA replication, cohesion establishment and cohesin acetylation are coordinated, we analysed the interaction partners of 55 human proteins implicated in these processes by mass spectrometry. This proteomic screen revealed that on chromatin the cohesin acetyltransferase ESCO2 associates with the MCM2‐7 subcomplex of the replicative Cdc45‐MCM‐GINS helicase. The analysis of ESCO2 mutants defective in MCM binding indicates that these interactions are required for proper recruitment of ESCO2 to chromatin, cohesin acetylation during DNA replication, and centromeric cohesion. We propose that MCM binding enables ESCO2 to travel with replisomes to acetylate cohesive cohesin complexes in the vicinity of replication forks so that these complexes can be protected from precocious release by WAPL. Our results also indicate that ESCO1 and ESCO2 have distinct functions in maintaining cohesion between chromosome arms and centromeres, respectively.     

The Effect of Lactobacillus salivarius SGL03 on Clinical and Microbiological Parameters in Periodontal Patients

The destruction of periodontal tissues during periodontitis is the result of the immune-inflammatory reactions to the bacteria of dental biofilm. Probiotics may reduce dysbiosis by the modification of the dental microbiome, which can influence the immune-inflammatory mechanisms. The aim of this study was to estimate the clinical and microbiological parameters, before and after 30 days of application of the dietary supplement containing Lactobacillus salivarius SGL03 or placebo. The study was conducted in 51 patients with stage I or II periodontitis during the maintenance phase of treatment. The clinical parameters and the number of colony forming units (CFU) of bacteria in supragingival plaque were assessed before and after 30 days of the oral once daily administration of the dietary supplement in the form of suspension containing L. salivarius SGL03 or placebo. There were no changes in the PI scores between and within the groups. The value of BOP decreased in both groups. In the study group the significant reduction of the mean pocket depth was revealed (from 2.5 to 2.42, p = 0,027) but without the difference between the groups. There were no significant changes in the number of bacteria within the groups. In the control, but not the study group, positive correlations were observed between the clinical parameters (variables) and the number of bacteria. The use of the dietary supplement containing L. salivarius SGL03 may reduce pocket depth despite the lack of changes in other clinical parameters and the number of bacteria in supragingival plaque.Key words: probiotics, periodontal treatment, Lactobacillus salivarius     

Cytoskeletal changes in hepatocytes induced by Microcystis toxins and their relation to hyperphosphorylation of cell proteins.

The heptapeptide toxins produced by the blue-green alga (cyanobacterium) Microcystis aeruginosa are selectively hepatotoxic in mammals. The characteristic post-mortem pathology of the liver is extensive lobular disruption due to sinusoidal breakdown, leakage of blood into the tissue and hepatocyte disintegration. Isolated hepatocytes incubated with toxin show severe structural deformity and surface blebbing. This paper demonstrates the effects of Microcystis toxins on the contraction and aggregation of actin microfilaments, and on the relocation and breakdown of cytokeratin intermediate filaments, in cultured hepatocytes. Earlier work did not show changes in the assembly/disassembly of actin; however, this paper demonstrates the change in cytokeratin from intermediate filaments to distributed granules in the cytoplasm of toxin-affected cells. Acrylamide gel electrophoresis of cytoskeletal fractions from hepatocytes did not show changes in total cytokeratins; however, marked changes in the immunogenicity of cytokeratins at 52 and 58 kDa were seen on toxin exposure of cells. Measurement of 32P-phosphorylation of proteins in toxin-affected cells incubated with [32P]orthophosphate showed a dramatic increase compared to control incubations. This is in agreement with research elsewhere describing phosphatase inhibition in vitro by Microcystis toxins. The data indicate that phosphorylated cytokeratin is a major component of cytoplasmic fraction phosphorylated protein after toxin exposure to hepatocytes. It is concluded that the mechanism of Microcystis toxicity to the hepatocyte is through cytoskeletal damage leading to loss of cell morphology, cell to cell adhesion and finally cellular necrosis. The underlying biochemical lesion is likely to be phosphatase inhibition causing hyperphosphorylation of a number of hepatocyte proteins, including those cytokeratins responsible for microfilament orientation and intermediate filament integrity.     

Biomass recycling: a key to efficient foraging by fungal colonies

Using an existing fungal growth model that captures the physiological processes of vegetative growth and development of a fungal colony, and in particular incorporates, for the first time, a recycling of biomass mechanism, we explore the effects of recycling in various environmental contexts. Here we test whether resource density thresholds exist, below which finite colony expansion occurs, in three dimensions based on the number of randomly removed resource sites. We then test the effect of recycling on resource density thresholds. Modelled soil structure, derived from experiments, is combined with the fungal growth model. The effect of recycling on foraging efficiency is investigated for resource distributed homogeneously and heterogeneously throughout the modelled soil structure. The simulated results show that resource density thresholds do exist in three dimensions and that the recycling mechanism decreases the threshold value. Our results indicate that recycling promotes persistence and a recycling mechanism is crucial for those fungi that reside in a resource patchy and limited environment.     

Inhibition of fatty acid synthesis in rabbit mammary alveolar explants by progesterone and related steroids

The possible inhibitory effect of steroids related to progesterone on prolactin-stimulated fatty acid synthesis in mammary alveolar explants from 11-day pseudopregnant rabbits, after culture, was investigated. Like progesterone, 17 alpha-hydroxyprogesterone, 20 alpha-dihydroprogesterone, medroxyprogesterone acetate, ethinodiol diacetate, ORG 2058 were inhibitory whereas pregnenolone and 5 alpha-pregnan-3,20-dione were not.