The differentiation of human memory B cells into specific antibody-secreting cells is CD40 independent.

It is generally accepted that memory B cells can be defined by their ability to produce, upon antigenic challenge, somatically mutated antibody molecules characterized by an increased affinity and by the expression of a downstream heavy chain isotype. However, the inability to isolate this particular B cell compartment has precluded the study of memory B lymphocyte physiology in man. We previously reported on the identification of an IgD- B cell subset in human tonsils that we defined as CD38- B cells, whose phenotype is highly reminiscent of that of memory B lymphocytes from the splenic marginal zone of rodents. In the present study, we developed a model of the measles virus (MV)-specific secondary antibody response in vitro to assess the presence of memory B lymphocytes in different B cell subsets isolated from human tonsils and explore the activation requirements of human memory B cells. Our findings show that the memory B cell pool resides in the CD38- B cell subpopulation and that the differentiation of MV-activated memory B cells into antibody-secreting cells can be achieved upon co-stimulation with interleukin (IL)-2 and IL-10, but does not require engagement of CD40. Interestingly, the CD40-mediated signal was found to synergize with Ig-cross-linking agents for the proliferation of memory B cells, but strongly suppressed their capacity to differentiate along the plasmacytoid pathway. Collectively, our results suggest that the CD40 signaling pathway is instrumental for the clonal expansion of the memory B cell pool, but does not operate in the later phase of the response, which allows their maturation into antibody-secreting cells.

Establishment and characterization of photoautotrophic protoplast-derived cultures ofNicotiana plumbaginifolia.

A procedure is described for the rapid establishment of photoautotrophic protoplast-derived cultures ofNicotiana plumbaginifolia. Photoautotrophic growth was induced by lowering the glucose concentration to 2.5 g.l(-1) in the protoplast culture medium and by omitting glucose from the subsequent dilution medium. Four week-old highly viable suspensions were plated on an agar-medium without glucose in unsealed Petri dishes and kept in illuminated chambers flushed with 0.05 % or 2 % CO2. Air-grown calli had net photosynthesis rates of 1.8 and 17 µmoles CO2.g(-1) fresh wt.h(-1) in air at 0.034 % CO2 and in air enriched with 1 % CO2, respectively. Calli grown in 2 % CO2 exhibited lower rates of net photosynthesis at the two CO2 concentrations tested (0 and 7.5 µmoles CO2.g(-1) fresh wt.h(-1), respectively). The contribution of photosynthesis to growth was estimated to be 80 % in air-grown calli and more than 90 % in calli grown in 2 % CO2. The suitability of this photoautotrophic culture procedure is discussed with regard to the screening of photosynthetic mutants or transformants from protoplasts.

A clinical, biometrological and ultrastructural study of xerotic skin.

The purpose of this in vivo study was to investigate, non-invasively on human subjects, xerotic skin and its physiological evolution over time, compared to normal skin. Two groups of 17 female subjects were studied during the winter season, one made up of subjects with normal skin and the other subjects with xerotic skin. A clinical assessment and biometrological measurements of hydration and transepidermal water loss (TEWL) were performed on the same area of the external antero-lateral surface of the leg at the start of the study then after three weeks. At the end of the study, the ultrastructure of stratum corneum samples taken from the same area was examined by transmission electron microscopy. Subjects with xerotic skin were selected according to their impaired cutaneous barrier function, reflected in a TEWL higher than 12 g/m ; 2/h. Compared to normal subjects, they presented a hydration level more than 25% lower. After an interval of 21 days, no significant change in the hydration level or clinical appearance of the xerotic skin was observed. In contrast, the TEWL had decreased significantly (D _ 21- D _ 0=-3.6 g/m ; 2/h; p < 0.001) but still stayed higher than normal values. Changes in the ultrastructure of the stratum corneum were also observed in the subjects with xerotic skin. Unlike normal skin, corneosomes could be detected right up to the surface layers, accompanied by intercellular lipids in an amorphous form. These observations confirm the important roles played by both corneosomes and lipid organization in the cohesion/desquamation processes. In the subjects with normal skin, the hydration level and barrier function remained unchanged during the three week study but an onset of skin dryness was observed, the mean clinical score increasing by +1.3 (p = 0.01). These results confirm that there is no direct relationship between TEWL and the severity of skin dryness. It appears that a clinical evaluation is more sensitive than biometrological measurement for describing early state of cutaneous dryness. This study highlights the importance of a regular cosmetic or dermopharmaceutical treatment during the winter to prevent xerosis apparition on legs.

A role for the VLA-4 integrin in the activation of human memory B cells.

It is generally recognized that activation through membrane effector molecules such as CD40 or the B cell receptor (BCR) is mandatory to allow B cells to proliferate and differentiate into antibody (Ab)-secreting cells in response to cytokines. We show here that purified tonsillar B cells can be stimulated directly by a cytokine combination to proliferate and secrete immunoglobulins when cultures are performed at high cell density. The contact-mediated activation of B cells in this experimental system is strongly inhibited both by anti-very late antigen (VLA)-4 monoclonal Ab and by a peptide containing the LDV sequence specifically recognized by the alpha 4 integrin binding site. These reagents also significantly suppressed the B cell responses elicited by engagement of the BCR or CD40. Our data reveal that memory B cells but not virgin or germinal center B cells are sensitive to the direct stimulatory effect of cytokines in high-density cultures. Finally, we found that the dual expression of the alpha and beta chains of VLA-4 is a distinctive feature of the memory B cell population. Collectively, our findings support the notion that VLA-4-dependent homotypic B cell interactions can mediate a co-stimulatory signal to human memory B cells and might participate in the B cell activation triggered through the BCR and CD40.