Chumappumkal Joseph Bilgimol, Whisenant Thomas C, Cooke Esther J, Zhou Jenny Y, Falah Nicca, De-Pablo Moreno Juan Andres, von Drygalski Annette
Department of Medicine, Division of Hematology/Oncology, University of California San Diego, La Jolla, California, United States of America.
University of California San Diego, Center for Computational Biology and Bioinformatics, La Jolla, California, United States of America.
PLoS One. 2025 May 19;20(5):e0320322. doi: 10.1371/journal.pone.0320322. eCollection 2025.
To investigate if FVIII-Fc Fusion protein (FcFVIII) may modulate inflammation and immune stimulation in hemophilic synovium via the Fc-portion of immunoglobulin used for half-life extension we performed gene expression profiling in FVIII-deficient mice. Hemarthrosis was induced by sub-patellar puncture in FVIII-KO mice, + /- periprocedural recombinant human (rh)FVIII,murine (m)FcFVIII, or mIgG2a. Synovium was harvested at baseline and on days (D) 3 and 14, followed by RNA extraction and sequencing, and histological analysis. RNASeq data were processed using standard protocols followed by differential gene expression (DGE) analysis. Functional enrichment analysis generated molecular pathways (KEGG and Reactome). To distinguish between on-target and off-target (related and unrelated to injury/bleed) effects the following groups were compared: i) Baseline vs. injured-saline, ii) injured-saline vs. injured-rhFVIII, iii) injured-saline vs. injured-mFcFVIII. Knee injury in FVIII-KO mice resulted in hemarthrosis, which was prevented by peri-procedural rhFVIII and mFcFVIII treatments. Only a small proportion of genes was affected by FVIII treatment, exhibiting overlap but also distinct differences between both FVIII-preparations. Acutely (D3), mFcFVIII had unique on-target effects related to immune and inflammatory regulation, whereas rhFVIII mostly affected mRNA and protein processing. On day 14, macrophage profiling indicated a transition from M1 to M2, and only mFcFVIII uniquely influenced pathways and genes associated with tissue remodeling and repair. Some mFcFVIII DGE patterns resembled mIgG2a patterns. Synovial vascular remodeling and cartilage health were better with mFcFVIII than rhFVIII. Interestingly, both FVIII-preparations exerted off-target effects on immune system pathways, albeit with temporal differences. These observations provide proof-of-principle that the type of FVIII preparation can influence synovial processes beyond acute hemostasis control, deserving exploration in the setting of joint bleed control in hemophilia.
为了研究FVIII-Fc融合蛋白（FcFVIII）是否可通过用于延长半衰期的免疫球蛋白的Fc部分来调节血友病性滑膜炎中的炎症和免疫刺激，我们在FVIII缺陷小鼠中进行了基因表达谱分析。通过在FVIII基因敲除（KO）小鼠的髌下穿刺诱导关节积血，手术过程中分别给予或不给予重组人（rh）FVIII、鼠源（m）FcFVIII或mIgG2a。在基线以及第3天和第14天采集滑膜，随后进行RNA提取和测序以及组织学分析。RNA测序（RNASeq）数据使用标准方案进行处理，随后进行差异基因表达（DGE）分析。功能富集分析生成分子途径（京都基因与基因组百科全书（KEGG）和Reactome）。为了区分靶向和非靶向（与损伤/出血相关和不相关）效应，比较了以下几组：i）基线与损伤-生理盐水组，ii）损伤-生理盐水组与损伤-rhFVIII组，iii）损伤-生理盐水组与损伤-mFcFVIII组。FVIII-KO小鼠的膝关节损伤导致关节积血，手术过程中给予rhFVIII和mFcFVIII治疗可预防。只有一小部分基因受FVIII治疗影响，两种FVIII制剂之间表现出重叠但也有明显差异。在急性期（第3天），mFcFVIII具有与免疫和炎症调节相关的独特靶向效应，而rhFVIII主要影响mRNA和蛋白质加工。在第14天，巨噬细胞分析表明从M1型向M2型转变，并且只有mFcFVIII独特地影响与组织重塑和修复相关的途径和基因。一些mFcFVIII的DGE模式类似于mIgG2a模式。mFcFVIII对滑膜血管重塑和软骨健康的改善优于rhFVIII。有趣的是，两种FVIII制剂均对免疫系统途径产生非靶向效应，尽管存在时间差异。这些观察结果提供了原理证明，即FVIII制剂的类型可以影响急性止血控制之外的滑膜过程，在血友病关节出血控制的背景下值得探索。
