Elimination of nurse cell nuclei that shuttle into oocytes during oogenesis

Drosophila oocytes develop together with 15 sister germline nurse cells (NCs), which pass products to the oocyte through intercellular bridges. The NCs are completely eliminated during stages 12–14, but we discovered that at stage 10B, two specific NCs fuse with the oocyte and extrude their nuclei through a channel that opens in the anterior face of the oocyte. These nuclei extinguish in the ooplasm, leaving 2 enucleated and 13 nucleated NCs. At stage 11, the cell boundaries of the oocyte are mostly restored. Oocytes in egg chambers that fail to eliminate NC nuclei at stage 10B develop with abnormal morphology. These findings show that stage 10B NCs are distinguished by position and identity, and that NC elimination proceeds in two stages: first at stage 10B and later at stages 12–14.     

Impact of the type of microcarrier and agitation modes on the expansion performances of mesenchymal stem cells derived from umbilical cord

The present study proposed to compare the impact of agitation mode (static, orbital, and mechanical) on the culture of mesenchymal stem cells extracted from the Wharton's jelly of umbilical cords (WJ-MSC), in a clinical grade culture medium, using human platelet lysate and different xeno-free microcarriers. Attachment, expansion, and detachment performances were characterized by a new dedicated tool of microscopic image posttreatment, allowing an in situ cell counting without detachment step. Results showed that performances in static mode were not necessarily representative of those obtained in dynamic mode. Moreover, impacts on nutrient consumptions and metabolite productions were identified, such as a higher glutamine consumption when Cytodex-1 microcarriers were used. The detachment strategy used was relatively efficient for Star-Plus, Plastic-Plus, and Hillex II, but not sufficient for Cytodex-1. Despite Cytodex-1 presented promising attachment and expansion performances, Star-Plus and Plastic-Plus showed a better compromise, respectively, for the orbital and the mechanical agitation modes.     

Polyamine Sharing between Tubulin Dimers Favours Microtubule Nucleation and Elongation via Facilitated Diffusion

We suggest for the first time that the action of multivalent cations on microtubule dynamics can result from facilitated diffusion of GTP-tubulin to the microtubule ends. Facilitated diffusion can promote microtubule assembly, because, upon encountering a growing nucleus or the microtubule wall, random GTP-tubulin sliding on their surfaces will increase the probability of association to the target sites (nucleation sites or MT ends). This is an original explanation for understanding the apparent discrepancy between the high rate of microtubule elongation and the low rate of tubulin association at the microtubule ends in the viscous cytoplasm. The mechanism of facilitated diffusion requires an attraction force between two tubulins, which can result from the sharing of multivalent counterions. Natural polyamines (putrescine, spermidine, and spermine) are present in all living cells and are potent agents to trigger tubulin self-attraction. By using an analytical model, we analyze the implication of facilitated diffusion mediated by polyamines on nucleation and elongation of microtubules. In vitro experiments using pure tubulin indicate that the promotion of microtubule assembly by polyamines is typical of facilitated diffusion. The results presented here show that polyamines can be of particular importance for the regulation of the microtubule network in vivo and provide the basis for further investigations into the effects of facilitated diffusion on cytoskeleton dynamics.     

“Escherichia coli‐milk” biofilm removal from stainless steel surfaces: Synergism between ultrasonic waves and enzymes

Three different methods to standardize biofilm removal for in situ sanitary control of closed surfaces in the food industry have been developed and compared, i.e. sonication, enzymatic treatment and a combined treatment which involved the application of ultrasound to enzyme preparations. The biofilm studied was an Escherichia coli model biofilm, made with milk on stainless steel sheets. Plate counting and epifluorescence microscopy were used to assess the efficiency of each treatment. The results are expressed in percentages, 100% denoting total removal, obtained with a flat ultrasonic transducer (T1) developed and presented in a previous study. The application of ultrasound by a patented curved transducer, T2 (10 s, 40 kHz), specifically devised for closed surfaces, was not sufficient to completely remove the biofilm (30 ± 7%). This biofilm was dislodged by two proteolytic enzyme preparations tested by immersion, viz. a 15‐min application of protease (84±1%) and a 30‐min trypsin application (95±8%). Using a combined treatment, the results showed a synergism between ultrasonic waves and proteolytic or glycolytic enzyme preparations, with removal of a significant amount of biofilm, i.e. 61–96% depending on the conditions tested, i.e. two to three times greater compared to sonication alone (30%). This application was in agreement with an industrial control, i.e. a good reproducible recovery of the biofilm in 10 s compared with 30 or 15 min with the enzyme alone.     

Characterization and localization of laccase forms in stem and cell cultures of sycamore

A laccase-type polyphenoloxidase (EC 1.10.3.2.), abundantly secreted by suspension-cultured sycamore (Acer pseudoplatanus) cells was purified to homogeneity. This laccase form is a glycoprotein (molecular weight 110000) with high mannose and complex glycans. The polypeptide moiety has a molecular weight of 66 000, indicating that the glycoprotein is 40% carbohydrate. Laccase is abundantly present in both the cell wall and the culture medium of suspension-cultured sycamore cells, but it is not detected in the cytoplasm, indicating that this large protein is efficiently secreted by the cells. Polyclonal rabbit antiserum was raised against the deglycosylated protein and was used to probe extracts of sycamore stem tissues. A second laccase form (molecular weight 56 000), antigenically related to laccase from cell cultures, is abundant in the epidermis of sycamore stems. In addition, this 56 kDa laccase form co-localizes with lignin precursors on tissue prints from sycamore stems. A polypeptide (molecular weight 50 000-56 000), antigenically related to sycamore laccase, was also immunodetected in most plant organs previously described in the literature as polyphenoloxidase-rich.