PKC-δ is an important molecule for B-cell proliferation and tolerance. B cells have long been recognized to play a part in osteoimmunology and pathological bone loss. However, the role of B cells with PKC-δ deficiency in bone homeostasis and the underlying mechanisms are unknown. We generated mice with PKC-δ deletion selectively in B cells by crossing PKC-δ-loxP mice with CD19-Cre mice. We studied their bone phenotype using micro-CT and histology. Next, immune organs were obtained and analyzed. Western blotting was used to determine the RANKL/OPG ratio in vitro in B-cell cultures, ELISA assay and immunohistochemistry were used to analyze in vivo RANKL/OPG balance in serum and bone sections respectively. Finally, we utilized osteoclastogenesis to study osteoclast function via hydroxyapatite resorption assay, and isolated primary calvaria osteoblasts to investigate osteoblast proliferation and differentiation. We also investigated osteoclast and osteoblast biology in co-culture with B-cell supernatants. We found that mice with PKC-δ deficiency in B cells displayed an osteopenia phenotype in the trabecular and cortical compartment of long bones. In addition, PKC-δ deletion resulted in changes of trabecular bone structure in association with activation of osteoclast bone resorption and decrease in osteoblast parameters. As expected, inactivation of PKC-δ in B cells resulted in changes in spleen B-cell number, function, and distribution. Consistently, the RANKL/OPG ratio was elevated remarkably in B-cell culture, in the serum and in bone specimens after loss of PKC-δ in B cells. Finally, in vitro analysis revealed that PKC-δ ablation suppressed osteoclast differentiation and function but co-culture with B-cell supernatant reversed the suppression effect, as well as impaired osteoblast proliferation and function, indicative of osteoclast–osteoblast uncoupling. In conclusion, PKC-δ plays an important role in the interplay between B cells in the immune system and bone cells in the pathogenesis of bone lytic diseases.

PKC-δ 是 B 细胞增殖和耐受的重要分子。B 细胞长期以来被认为在骨免疫学和病理性骨丢失中发挥作用。然而，PKC-δ 缺乏的 B 细胞在骨稳态中的作用及其潜在机制尚不清楚。我们通过将 PKC-δ-loxP 小鼠与 CD19-Cre 小鼠杂交，在 B 细胞中选择性地产生了具有 PKC-δ 缺失的小鼠。我们使用显微 CT 和组织学研究了他们的骨表型。接下来，获得并分析免疫器官。使用蛋白质印迹法测定体外 B 细胞培养物中的 RANKL/OPG 比率，使用 ELISA 测定和免疫组织化学分别分析血清和骨切片中的体内 RANKL/OPG 平衡。最后，我们利用破骨细胞生成通过羟基磷灰石再吸收测定研究破骨细胞功能，和分离的原代颅骨成骨细胞以研究成骨细胞的增殖和分化。我们还研究了与 B 细胞上清液共培养的破骨细胞和成骨细胞生物学。我们发现 B 细胞中 PKC-δ 缺乏的小鼠在长骨的小梁和皮质区室中表现出骨质减少表型。此外，PKC-δ 缺失导致骨小梁结构的变化与破骨细胞骨吸收的激活和成骨细胞参数的降低相关。正如预期的那样，B 细胞中 PKC-δ 的失活导致脾脏 B 细胞数量、功能和分布的变化。一致地，在 B 细胞中失去 PKC-δ 后，B 细胞培养物、血清和骨标本中的 RANKL/OPG 比率显着升高。最后，体外分析表明，PKC-δ 消融抑制破骨细胞分化和功能，但与 B 细胞上清液共培养逆转了抑制作用，以及成骨细胞增殖和功能受损，表明破骨细胞 - 成骨细胞解偶联。总之，PKC-δ在免疫系统中的B细胞与骨细胞在溶骨性疾病的发病机制中的相互作用中起重要作用。