Highly conserved among species and expressed in a variety of types of cells, numerous roles have already been related to the cellular prion protein (PrPC). to supplementary effects thought as Acute Rays Symptoms commonly.1 Irradiation from the BM problems hematopoietic stem and progenitor cells (HSPC) and perturbs the hematopoietic microenvironment,2,3 leading to radiation-induced severe myelosuppression4,5 and increased susceptibility to infections.6,7 Numerous types of DNA lesions are TNFRSF1A induced by cell contact with ionizing rays. They include foundation modifications, apurinic/apyrimidic sites (AP sites), and solitary- (SSB) and double (DSB)-strand breaks. DSB are the main lesions influencing cell survival. They can arise not only directly by deposition of energy within the DNA, but also as a consequence of the formation of AP sites or AS-605240 supplier SSB.8,9 Indeed, base excision repair (BER) activities, and in particular the processing of abasic sites, have been shown AS-605240 supplier to contribute to radiation-induced DNA damage.10,11 Apurinic/apyrimidic endonuclease-1 (Ape1) is the unique enzyme that converts AP sites into SSB intermediates during BER. Ape1 3-phosphodieterase and -phosphate activities (for a review, observe Laev knockout mice to study the consequences of PrPC deficiency on hematopoiesis of young and aged adult mice, and on the response of hematopoietic stem cells (HSC) and hematopoietic progenitors to gamma-irradiation. Methods Mice Mice experiments were carried out in compliance with the Western Community Council Directive (EC/2010/63) and were authorized by our institutional ethics committee (CetEA-CEA-DRFCn. 17-096). The B6.129S7-Prnptm1Cwe/Orl mice were from your Western Mutant Mouse Archive and bred in our animal facility. We also used ZH3/ZH3 mice provided by A. Aguzzi (Zurich, Switzerland) and C57BL/6 mice were purchased from Charles River. Cell sorting and circulation cytometry analysis of bone marrow cells Murine BM cells were flushed out of femurs, tibiae, hip bone and humeruses using a syringe filled with DPBS and filtered through a 70 m-cell strainer. After red blood cell lysis using NH4Cl answer (STEMCELL Systems), mononuclear cells were phenotyped using different antibody cocktails from Biolegend, e-Bioscience or Beckton Dickinson. Circulation cytometry analysis was performed having AS-605240 supplier a BD FACS AS-605240 supplier LSRIITM circulation cytometer (BD Biosciences) and cell sorting having a FACS Influx cell sorter (Becton Dickinson). Data were analyzed with FlowJo software. Antibodies and gating strategies for hematopoietic subset analysis and sorting are explained in the in AS-605240 supplier different purified hematopoietic subpopulations, i.e. common myeloid progenitors (CMP), granulocyte-monocyte progenitors (GMP), megakaryocyte-erythrocyte progenitors (MEP), MPP, and hematopoietic stem cells (HSC). The highest level of mRNA was found in MEP while they were 2.7-fold and 4.3-fold lower in CMP and GMP, respectively (Number 1A). These variations in mRNA manifestation were also found at the protein level (Number 1B and mRNA level in purified HSC was 2.5-fold higher than in MPP (Number 1C). Open in a separate window Number 1 PrPC contributes to mouse hematopoietic homeostasis. (A) quanta-tive real-time polymerase chain reaction (qRT-PCR) analysis of manifestation, normalized to in the indicated bone marrow (BM) subpopulations: CMP: common myeloid progenitor; GMP: granulocyte-macrophage progenitor; MEP: megakaryocyte-erythrocyte progenitor purified by circulation cytometry from BM of 3-month previous mice (n=7-9). Data are provided as meanstandard mistake of mean (SEM). Means with different words are considerably different (appearance, normalized to in hematopoietic stem cell (HSC) (LSK Compact disc135?) and in multipotent progenitor (MPP) (LSK Compact disc135+) purified by stream cytometry from BM of 3-month previous mice (n=9); Data are provided as meanSEM. ***plating performance of CMP and GMP purified by stream cytometry from BM of WT (dark pubs) and KO (white pubs) mice (n=6-9). Data are provided as meanSEM. **appearance, normalized to Actb in WT and KO HSC and MPP purified by stream cytometry from BM of 3-month and 11-month previous.