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Human CD38hiCD138⁺ plasma cells can be generated in vitro from CD40-activated switched-memory B lymphocytes.

Maïga RI, Bonnaure G, Rochette JT, Néron S - J Immunol Res (2014)

Bottom Line: Furthermore, the generated CD38(hi)CD138(+) cells showed a higher proportion of CD31(+) cells than the CD38(hi)CD138(-) cells.Besides, analyses done with human blood and bone marrow plasma cells showed that in vivo and de novo generated CD38(hi)CD138(+) cells have a similar CD31 expression profile but are distinct according to their reduced CD39 expression level.Overall, we have evidences that in vitro generated plasma cells are heterogeneous and appear as CD39(+) precursors to the ones present in bone marrow niches.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Microbiology and Bio-Informatics, Laval University, 1045 Avenue de la Médecine, Québec, QC, Canada G1V 0A6 ; Hema-Quebec's Department of Research and Development, 1070 Avenue des Sciences-de-la-Vie, Québec, QC, Canada G1V 5C3.

ABSTRACT
B lymphocyte differentiation into long-lived plasma cells is the keystone event for the production of long-term protective antibodies. CD40-CD154 and CD27-CD70 interactions are involved in human B lymphocyte differentiation into CD38(hi)CD138(+) cells in vivo as well as in vitro. In this study, we have compared these interactions in their capacity to drive switched-memory B lymphocytes differentiation into CD38(hi)CD138(+) plasma cells. The targeted B lymphocytes were isolated from human peripheral blood, expanded for 19 days, and then submitted to CD70 or CD154 interactions for 14 days. The expanded B lymphocytes were constitutively expressing CD39, whereas CD31's expression was noticed only following the in vitro differentiation step (day 5) and was exclusively present on the CD38(hi) cell population. Furthermore, the generated CD38(hi)CD138(+) cells showed a higher proportion of CD31(+) cells than the CD38(hi)CD138(-) cells. Besides, analyses done with human blood and bone marrow plasma cells showed that in vivo and de novo generated CD38(hi)CD138(+) cells have a similar CD31 expression profile but are distinct according to their reduced CD39 expression level. Overall, we have evidences that in vitro generated plasma cells are heterogeneous and appear as CD39(+) precursors to the ones present in bone marrow niches.

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Generation of plasma cells from switched-memory B lymphocytes. Monitoring of plasma cell generation in the differentiation phase was done by flow cytometry and all analyses were done on viable cell populations. The results are presented as the mean ± S.E.M. of 6 independent samples: (a) CD38 and CD138 expression profiles on D0, 5 and 14. D0 being the day that the expanded memory B cells were thawed and were put in culture with CD70+ or CD154+ cell lines. (b) CD38hi cells frequency. The comparison of CD38hi cells frequency at D5 was determined using a Mann-Whitney test, P value = 0.0260. For the monitoring of CD138+ cells frequency (c), the Kruskal-Wallis test was used and no significant difference was observed. (d) Generated plasma cells morphology at D14 shown by fluorescence microscopy (100x immersion oil objective). Blue: nucleus; red: actin. Representative cells are shown.
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fig3: Generation of plasma cells from switched-memory B lymphocytes. Monitoring of plasma cell generation in the differentiation phase was done by flow cytometry and all analyses were done on viable cell populations. The results are presented as the mean ± S.E.M. of 6 independent samples: (a) CD38 and CD138 expression profiles on D0, 5 and 14. D0 being the day that the expanded memory B cells were thawed and were put in culture with CD70+ or CD154+ cell lines. (b) CD38hi cells frequency. The comparison of CD38hi cells frequency at D5 was determined using a Mann-Whitney test, P value = 0.0260. For the monitoring of CD138+ cells frequency (c), the Kruskal-Wallis test was used and no significant difference was observed. (d) Generated plasma cells morphology at D14 shown by fluorescence microscopy (100x immersion oil objective). Blue: nucleus; red: actin. Representative cells are shown.

Mentions: To verify whether CD40-CD154 and CD27-CD70 interactions were similarly competent for in vitro differentiation of switched-memory B lymphocytes, we used a cryopreserved expanded-memory B lymphocytes bank. In all following experiments, the day 0 (D0) is the first day the thawed cells were put in culture to induce their differentiation. For the differentiation phase in the presence of IL-6 and IL-10, adherent cell lines expressing CD70 (3H7) or CD154 (L4.5) were used to generate a low level of interaction stimulation (Figures 2 and 3).


Human CD38hiCD138⁺ plasma cells can be generated in vitro from CD40-activated switched-memory B lymphocytes.

Maïga RI, Bonnaure G, Rochette JT, Néron S - J Immunol Res (2014)

Generation of plasma cells from switched-memory B lymphocytes. Monitoring of plasma cell generation in the differentiation phase was done by flow cytometry and all analyses were done on viable cell populations. The results are presented as the mean ± S.E.M. of 6 independent samples: (a) CD38 and CD138 expression profiles on D0, 5 and 14. D0 being the day that the expanded memory B cells were thawed and were put in culture with CD70+ or CD154+ cell lines. (b) CD38hi cells frequency. The comparison of CD38hi cells frequency at D5 was determined using a Mann-Whitney test, P value = 0.0260. For the monitoring of CD138+ cells frequency (c), the Kruskal-Wallis test was used and no significant difference was observed. (d) Generated plasma cells morphology at D14 shown by fluorescence microscopy (100x immersion oil objective). Blue: nucleus; red: actin. Representative cells are shown.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4352507&req=5

fig3: Generation of plasma cells from switched-memory B lymphocytes. Monitoring of plasma cell generation in the differentiation phase was done by flow cytometry and all analyses were done on viable cell populations. The results are presented as the mean ± S.E.M. of 6 independent samples: (a) CD38 and CD138 expression profiles on D0, 5 and 14. D0 being the day that the expanded memory B cells were thawed and were put in culture with CD70+ or CD154+ cell lines. (b) CD38hi cells frequency. The comparison of CD38hi cells frequency at D5 was determined using a Mann-Whitney test, P value = 0.0260. For the monitoring of CD138+ cells frequency (c), the Kruskal-Wallis test was used and no significant difference was observed. (d) Generated plasma cells morphology at D14 shown by fluorescence microscopy (100x immersion oil objective). Blue: nucleus; red: actin. Representative cells are shown.
Mentions: To verify whether CD40-CD154 and CD27-CD70 interactions were similarly competent for in vitro differentiation of switched-memory B lymphocytes, we used a cryopreserved expanded-memory B lymphocytes bank. In all following experiments, the day 0 (D0) is the first day the thawed cells were put in culture to induce their differentiation. For the differentiation phase in the presence of IL-6 and IL-10, adherent cell lines expressing CD70 (3H7) or CD154 (L4.5) were used to generate a low level of interaction stimulation (Figures 2 and 3).

Bottom Line: Furthermore, the generated CD38(hi)CD138(+) cells showed a higher proportion of CD31(+) cells than the CD38(hi)CD138(-) cells.Besides, analyses done with human blood and bone marrow plasma cells showed that in vivo and de novo generated CD38(hi)CD138(+) cells have a similar CD31 expression profile but are distinct according to their reduced CD39 expression level.Overall, we have evidences that in vitro generated plasma cells are heterogeneous and appear as CD39(+) precursors to the ones present in bone marrow niches.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Microbiology and Bio-Informatics, Laval University, 1045 Avenue de la Médecine, Québec, QC, Canada G1V 0A6 ; Hema-Quebec's Department of Research and Development, 1070 Avenue des Sciences-de-la-Vie, Québec, QC, Canada G1V 5C3.

ABSTRACT
B lymphocyte differentiation into long-lived plasma cells is the keystone event for the production of long-term protective antibodies. CD40-CD154 and CD27-CD70 interactions are involved in human B lymphocyte differentiation into CD38(hi)CD138(+) cells in vivo as well as in vitro. In this study, we have compared these interactions in their capacity to drive switched-memory B lymphocytes differentiation into CD38(hi)CD138(+) plasma cells. The targeted B lymphocytes were isolated from human peripheral blood, expanded for 19 days, and then submitted to CD70 or CD154 interactions for 14 days. The expanded B lymphocytes were constitutively expressing CD39, whereas CD31's expression was noticed only following the in vitro differentiation step (day 5) and was exclusively present on the CD38(hi) cell population. Furthermore, the generated CD38(hi)CD138(+) cells showed a higher proportion of CD31(+) cells than the CD38(hi)CD138(-) cells. Besides, analyses done with human blood and bone marrow plasma cells showed that in vivo and de novo generated CD38(hi)CD138(+) cells have a similar CD31 expression profile but are distinct according to their reduced CD39 expression level. Overall, we have evidences that in vitro generated plasma cells are heterogeneous and appear as CD39(+) precursors to the ones present in bone marrow niches.

Show MeSH
Related in: MedlinePlus