Department of Biochemistry, Genetics and Immunology, University of Vigo, Vigo, Spain.
BMC Bioinformatics. 2013;14 Suppl 6(Suppl 6):S8. doi: 10.1186/1471-2105-14-S6-S8. Epub 2013 Apr 17.
The germinal center (GC) reaction leads to antibody affinity maturation and generation of memory B cells, but its underlying mechanisms are poorly understood. To assemble this puzzle, several key pieces of information are needed, one in particular being the number of participating B cell clones. Since this clonal diversity cannot be observed directly, earlier studies resorted to interpreting two types of available experimental data: Immunohistology of GCs containing two phenotypically distinct B-cell populations, and antibody gene sequences of small B-cell samples from GCs. Based on a simple model, investigators concluded that a typical GC was seeded by 2-8 B cells, endorsing the current notion that GCs are oligoclonal from the onset.
A re-evaluation of these data showed that the used simple model is not statistically consistent with the original data. From an analysis of the experimental system, we propose a new model for estimating GC clonal diversity, including the initially neglected sampling and measurement errors, and making more general assumptions. Consistency analysis with the new model yielded an estimation of sampling and measurement errors in the experimental data of 10-11% for one B-cell population and 62-64% for the other population, and an average number of 19-23 seeder B cells. An independent analysis of antibody gene sequences of small B-cell samples from GCs, using an adapted Yule estimator of diversity, yielded a minimum estimation of 20-30 GC founder B cells, confirming the previous results.
Our new experimental-based model provides a highly improved method to estimate the clonal diversity of GCs from immunohistochemistry data of chimeric animals. Calculations based on this model, and validated by an independent approach, indicate that GCs most likely contain broadly varying numbers of different B cell clones, averaging 5- to 10-fold more clones than previously estimated. These findings, in line with recent results showing that GC sizes and life times are also subject to high variability, dramatically change the picture of GC dynamics.
生发中心（GC）反应导致抗体亲和力成熟和记忆 B 细胞的产生，但其潜在机制尚不清楚。为了拼凑这个谜题，需要一些关键信息，其中一个特别重要的信息是参与的 B 细胞克隆的数量。由于这种克隆多样性无法直接观察到，早期的研究依赖于解释两种可用的实验数据：包含两种表型不同的 B 细胞群体的 GC 的免疫组织化学，以及来自 GC 的小 B 细胞样本的抗体基因序列。基于一个简单的模型，研究人员得出结论，一个典型的 GC 是由 2-8 个 B 细胞播种的，这支持了 GC 从一开始就是寡克隆的现有观点。
对这些数据的重新评估表明，所使用的简单模型在统计学上与原始数据不一致。从对实验系统的分析中，我们提出了一种新的模型来估计 GC 克隆多样性，包括最初被忽略的抽样和测量误差，并做出更一般的假设。用新模型进行一致性分析得出，对于一个 B 细胞群体，实验数据中的抽样和测量误差估计为 10-11%，对于另一个群体为 62-64%，平均有 19-23 个播种 B 细胞。对来自 GC 的小 B 细胞样本的抗体基因序列进行独立分析，使用适应性的 Yule 多样性估计器，得出了 20-30 个 GC 起始 B 细胞的最小估计值，证实了之前的结果。
我们基于实验的新模型为从嵌合动物的免疫组织化学数据估计 GC 的克隆多样性提供了一种高度改进的方法。基于该模型的计算，并通过独立方法验证，表明 GC 很可能包含广泛不同数量的不同 B 细胞克隆，平均比以前估计的多 5-10 倍。这些发现与最近的结果一致，表明 GC 的大小和寿命也存在高度的可变性，这极大地改变了 GC 动力学的图景。
