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Publikationssystem UB Tuebingen advisor Triebskorn, Rita (Prof. Dr.) author Doll, Larissa 2025-10-02T13:09:12Z 2025-10-02 http://hdl.handle.net/10900/170703 Severe congenital neutropenia (CN) is a genetically heterogeneous condition characterized by an inborn neutrophil deficiency. It is caused by germline mutations in various genes, including autosomal recessive mutations in HAX1, JAGN1, or COPZ1. CN patients suffer from severe infections from birth, and without treatment, these can be life-threatening. Current understanding of CN pathophysiology is mainly based on in vitro assays and clinical observations. To date, no faithful mouse models have been described. Mice deficient in HAX1 or JAGN1 are prematurely lethal and don't develop neutropenia, and a COPZ1 model has not yet been established. The evolutionary conservation of most processes underlying hematopoiesis between humans and zebrafish makes it an ideal alternative vertebrate model system to study normal and malignant granulopoiesis. The aim of this study was to establish HAX1-, JAGN1-, and COPZ1-associated CN zebrafish models to study the underlying pathomechanisms and the discovery of new therapeutic approaches. We have successfully established three CN models by mimicking the human mutations in the zebrafish orthologs hax1, jagn1b, and copz1. Using these models, we studied different processes, such as the induction of unfolded protein response (UPR), apoptosis of myeloid progenitors, and the impairment of the G-CSFR pathway, which were previously described as potential causes of neutropenia development. Despite active UPR upon different Jagn1b alterations and an increase in apoptotic cells after downregulating hax1 or jagn1b, we dismissed these processes as solely neutropenia-causing mechanisms since the chemical induction of UPR or apoptosis did not lead to a reduced number of neutrophils. In the HAX1- and JAGN1- associated CN models, the analyses of the G-CSFR pathway showed an altered signaling, resulting in a decreased expression of the central regulator of steady-state granulopoiesis, cebpa. Similarly, a reduced expression of CEBPA was seen in HSPCs carrying truncated COPZ1. Moreover, treating our zebrafish CN models with the HIF1a activator, IOX2, or the pan-CDK inhibitor, flavopiridol, rescued their neutropenia phenotype. The activation of HIF1a is associated with CEBPA activation and the inhibition of CDK2/4 mimics C/EBPa function. These mechanisms of action could compensate for the reduced cebpa expression and explain the induction of granulopoiesis after IOX2 or flavopiridol treatment. However, the underlying cause of neutropenia development is a complex process involving several interrelated mechanisms. Our CN zebrafish models, established during this PhD, have allowed us to get closer to understanding these mechanisms and are a potential tool for further studies. In addition, these models enable rapid testing of the in vivo effects of candidate therapeutics on granulopoiesis, an essential step toward the clinical translation of our experimental findings. en ubt-podno Establishment of Experimental Severe Congenital Neutropenia (CN) Models in Zebrafish PhDThesis URL https://publikationen.uni-tuebingen.de/xmlui/bitstream/10900/170703/1/Dissertation_Doll%2cLarissa.pdf File MD5 56150b980963e4fab6cf0a3b24ac493e 14337536 application/pdf Dissertation_Doll,Larissa.pdf URL https://publikationen.uni-tuebingen.de/xmlui/bitstream/10900/170703/3/Dissertation_Doll%2cLarissa.pdf.txt File MD5 76b76d48f905b93973bf3305eaa439b6 550883 text/plain Dissertation_Doll,Larissa.pdf.txt