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Summary: The transcription factor hepatic leukemia factor (HLF) is strongly expressed in hematopoietic stem cells (HSCs) and is thought to influence both HSC self-renewal and leukemogenesis. However, the physiological role of HLF in hematopoiesis and HSC function is unclear. Here, we report that mice lacking Hlf are viable with essentially normal hematopoietic parameters, including an intact HSC pool during steady-state hematopoiesis. In contrast, when challenged through transplantation, Hlf-deficient HSCs showed an impaired ability to reconstitute hematopoiesis and became gradually exhausted upon serial transplantation. Transcriptional profiling of Hlf-deficient HSCs revealed changes associated with enhanced cellular activation, and cell-cycle analysis demonstrated a significant reduction of quiescent HSCs. Accordingly, toxic insults targeting dividing cells completely eradicated the HSC pool in Hlf-deficient mice. In summary, our findings point to HLF as a critical regulator of HSC quiescence and as an essential factor for maintaining the HSC pool during regeneration. : Komorowska et al. report that the transcription factor HLF is required to maintain hematopoietic stem cell (HSC) function during regeneration. Moreover, Hlf-deficient HSCs are less quiescent. In accordance with this, toxic insults targeting dividing cells completely eradicate the HSC pool in Hlf-deficient mice. Keywords: HSC, transcription factor, HLF, quiescence, self-renewal, cell cycle, reconstitution, transplantation, 5FU, stress

Background Cytomegalovirus (CMV) infection is associated with an increased risk of cardiac complications in kidney transplant recipients (KTRs). Some data suggest that CMV may be involved in atherogenesis. The aim of the study was the analysis of CMV medical history in KTRs and its influence on cardiovascular (CV) incidents. Materials and Methods The study observed 254 patients (165 male/89 female) with mean age of 47.2 (range, 15–81) years and duration of dialysis before transplantation 29.2 months who received transplants in 1 university unit (2007–2013). Thirty-six patients were transplanted preemptively. The mean time of observation lasted 7 years. KTRs suffered from diabetes, hypertension, and hyperlipidemia (17.3%, 88.5%, and 61%, respectively). Coronary artery disease was diagnosed in 19.6% patients, 3.5% underwent elective coronary surgery operation, and 9.05% had CV incidents before transplantation. The following CMV donor/recipient (D/R) viral statuses were noticed in the study group: D+/R+ (68.9%), D+/R− (16.9%), D−/R+ (10.2%), and D−/B− (3.9%). D+/R− received universal CMV prophylaxis; the rest were under preemptive CMV prophylaxis. CMV infection affected 87 (34.25%) patients; there were 24 primary infections and 85 secondary infections (some patients had more than 1 CMV). Mean time of diagnosis of the primary and secondary CMV infection was 190.7 and 160.5 days, respectively. Results During observation 22 patients experienced 26 CV incidents: 15 were D+/R+, 6 were D+/R−, and 1 was D−/R+. CMV infections occurred in 40.9% of patients with CV incidents after kidney transplantation. In comparison, 33.6% patients without CV incidents after kidney transplantation suffered from CMV infection. Conclusions CMV infection in KTRs was not a crucial risk factor for CV incidents.

Incidence of malignancy in transplant recipients is higher than in the general population. Malignancy is a major cause of mortality following solid organ transplantation and a major barrier to long-term survival for the kidney. The aim of this study was to estimate the incidence of solid organ malignancy (SOM) and melanoma in renal transplant recipients (RTR) transplanted at 2 representative transplant centers in Poland based on data from the Polish Tumor Registry.We analyzed the medical data of 3069 patients who underwent kidney transplantation (KTx) between 1995 and 2015.In our study 112 SOM (3.6%) were diagnosed. The majority of patients were male (n = 71; 63.4%; P .01). The mean age at KTx was 48.0 ± 13.1 years and the mean age at the time of cancer diagnosis was 55.9 ± 12.7 years. The average time of malignancy occurrence was 5.9 ± 5.0 years after KTx. SOM was the cause of death in 60 patients (53%). The most common were malignancies of gastrointestinal tract (25%), urinary tract tumors (23.2%), lung cancer (n = 18; 16%), and lymphoma (13.4%). We found an increase in the percentage of chronic glomerular nephropathy in the group of SOM (n = 56; 50%) compared with renal insufficiency of other etiologies.RTR in Poland are at a significant risk of malignancy development in a variety of organs, primarily urinary tract tumors and lymphoma. Cancers most frequently occurring in the general population such as lung and colorectal cancer are common in our RTR. On this basis an appropriate tumor screening schedule can be developed in individual countries.

Background Cardiovascular (CV) diseases are the most common cause of death in patients with chronic kidney disease, including patients after kidney transplantation. The aim of the study was to do a retrospective analysis of CV risk in renal transplant recipients (RTRs). Methods The analysis of CV risk was based on the following scales: QRISK2, Framingham (assessment of development of CV disease), PROCAM (assessment of any CV incident), and Pol-SCORE (assessment of CV death) within a 10-year period. Out of 150 RTRs transplanted in 2007–2009, 100 RTRs (65 male/35 female) with an average age of 48.4 years were enrolled in the study. Coronary heart disease and diabetes mellitus were diagnosed in 7% and 15% of participants, respectively. Coronarography was performed in 38% of patients. Hypertension was diagnosed in 98% of participants, myocardial infarction was diagnosed in 6% of participants, and stroke was diagnosed in 2% of participants. Results High and very high risk of CV endpoint according to QRISK2, PROCAM, Framingham, and Pol-SCORE scales was found in 41%, 8%, 10%, and 41% of patients, respectively. After 5 years of follow-up, a total of 13 CV events (myocardial infarction and stroke) were observed in 11 patients. Among these patients, the highest risk of endpoint according to QRISK2, PROCAM, Framingham, and Pol-SCORE scales was found in 36%, 9%, 18%, and 45% of patients, respectively. Conclusions The QRISK2 and Pol-SCORE scales seem to be the most predictive in assessing CV risk in RTRs.

The transcription factor Hepatic leukemia factor (HLF), initially identified as part of a leukemic fusion protein (E2A-HLF) causing B-lymphoid leukemia in children, is expressed in a wide range of tissues. In the mouse hematopoietic system, HLF is highly expressed in hematopoietic stem cells while being hardly detectable in more restricted progenitor populations. We have previously shown that in adult mice, HLF is essential for preserving the hematopoietic stem cell (HSC) pool during regeneration by maintaining their quiescence (Komorowska et al, Cell reports, 2017). Notably, despite this highly specific expression profile and a critical role in hematopoietic regeneration, mice with double knock-out of HLF alleles (KO) appeared to age normally (18-24 months survival > 90% for both WT and KO). In this study, we investigated the role of HLF in HSC during aging and leukemogenesis. We first confirmed that HLF is expressed all through ontogeny by performing quantitative PCR on HSC and progenitor populations from mouse fetal liver (E14.5 FL) and bone marrow (BM) of aged mice (18-month-old mice). We found that not only is HLF expressed in hematopoietic cells from embryonic to aged mice, but also that the expression profile was highly conserved. In fact, we found that HLF expression decreased during hematopoietic differentiation independently of ontogenic age (HLF/HPRT relative expression: 6 in HSC (LSK-CD150+CD48-), 3 in HSPC (LSK), and 5) and secondary recipients (donor contribution: WT: 67% (SEM 2.7), KO: 8.4% (SEM 3.6), n=4). In addition to its function in healthy hematopoiesis, studies have suggested that HLF is a downstream mediator of the MLL/AF9 (MA9) leukemic fusion protein. To address the contribution of HLF to MA9 leukemogenesis, mice were transplanted with either KO- or WT-BM-derived c-Kit+ cells expressing the MA9 fusion protein. Preliminary results showed that HLF did not affect the latency of the leukemia in primary recipients with mice succumbing to a lethal disease at days 40-41 after transplant independently of the leukemic graft genetic background. However, KO leukemic cells had a significantly delayed leukemia development in secondary recipients (time to death; WT: 32-39 days, KO: 39-68 days). Moreover, semi-quantitative PCR showed that not only was HLF expressed in leukemia cells harvested from primary recipients but also that the expression level displayed a tendency to an increase in leukemia cells from secondary recipients. Altogether, our data suggest that while HLF may be dispensable for leukemic transformation, it promotes the development of MLL-AF9 leukemia in mice. We are currently in the process of confirming the role of HLF in MA9 leukemia development. Taken together, we here show that HLF is essential for HSCs from embryonic to aged mice. We further show that HLF is critical for serial propagation of AML cells, suggesting that HLF is needed to maintain leukemia stem cells. Disclosures No relevant conflicts of interest to declare.

In many countries on different continents, at least 10-12% of people are suffering from chronic kidney disease (CKD). These data indicate that CKD is a big problem, much more than previously expected. Therefore, programs should be promoted for early detection of kidney diseases and carry out public education and also the medical community.The aim of this article was the assessment of the nephrology knowledge of participants of the World Kidney Day in Gdansk in 2012. Analysis of the survey conducted for the entire cohort and the separate subgroups of people who were more than 60 years old and less than 60.Among the total sample and in the subgroups, complications of hypertension knowledge was declared by more than 75% of the participants, diabetes more than 70%. However the symptoms of CKD have been known only for about 60%, the concept of creatinine 40%, while the concept of dialysis/transplant for about 80% of respondents. The main source of medical information were non-medical journals for more than 50% of study participants. However the doctor as a source of medical knowledge, identified 47% of respondents.Participants of questionnaires have showed in described analysis mediocre acquaintance of signs of kidney disease, the notion of creatinine in comparison for acquaintance of complication of hypertension and diabetes. However, for all participants of research and for older subgroup, main source of medical knowledge was a physician and unmedical magazines. But internet or television was the main source of knowledge for younger subgroup.

The transcription factor Hepatic Leukemia Factor (HLF) was originally identified in a chromosomal translocation with the gene E2A causing a subset of childhood B-lineage acute lymphoid leukemia. Moreover, HLF has been described as a regulator of circadian rhythm and recent findings have implicated HLF as a candidate “stemness” gene in both normal and malignant stem cells. Accordingly, overexpression of HLF in human hematopoietic stem cells (HSC) results in an enhanced reconstitution capability in NOD-SCID mice. However, little is known about HLF’s physiological role in hematopoiesis and HSC regulation. Using quantitative PCR, we found that HLF is highly expressed in mouse (C57Bl/6) HSC and is downregulated upon differentiation (HSC 3.2 (±0.95) fold (p The conventional HLF knockout (KO) mice (C57bl/6 background) were viable, born at normal Mendelian ratios and showed normal hematopoietic parameters (bone marrow cellularity: WT 2.7x107 (±5.4 x106), KO 3.3x107 (±6.4 x106), p>0.2 n=9). In addition, the HLF KO mice demonstrated normal lineage distribution of both mature cells in the peripheral blood and bone marrow as well as the frequency of immunophenotypic HSC (Lin-Sca1+ckit+CD34-Flt3-: WT 0.0005 (±0.5x10-4)%, KO 0.0005 (±0.1x10-3)%; n>10). However, in a serial competitive transplantation assay using whole bone marrow (200 000 cells 1:1 ratio), HLF KO cells demonstrated a significant reduction in reconstitution capacity in primary recipients (WT 56 (±15)%, KO 40.2 (±16)%, p=0.028, n>10), which was further increased in the secondary recipients (WT 87.2 (±26)%, KO 8.7 (±5.8)%, p10). Almost no engraftment was detected from the HLF KO cells in tertiary recipients. To further evaluate stem cell activity in the absence of HLF, we next enumerated the number of competitive repopulating units (CRU) by limiting dilution assay, which revealed a 2.6 fold reduction, of CRU in the HLF KO mice compared to WT controls (WT 1.6 (±0.4)/105 bone marrow cells, KO 0.6 (±0.2)/105 bone marrow cells). Similarly, transplantation of sorted HSC (Lin-Sca1+ckit+CD34-Flt3-) also showed a 2.4 fold (WT 47.3 (±24)%, KO 19.4 (±25)%, p=0.16, n=9) reduced engraftment of total cells but with enhanced T cell frequency in peripheral blood (WT 19.5 (±6.2)%, KO 40.8 (±7.4)%, p=0.01, n=9). Since we also found that HLF was highly expressed in fetal liver derived HSC, we transplanted fetal liver HLF KO cells from E14.5 in a competitive repopulation setting. In line with the phenotype seen in the adult HLF KO mice, the fetal liver HLF KO cells demonstrated impaired reconstitution ability (WT 52.8 (±16)%, KO 0.9 (±1.4)%, n>10). Intriguingly, the phenotype was stronger than in the adult HLF KO HSC, indicating that HLF is particularly important during the expansion phase of HSC in embryonic development. The underlying mechanism of the reduced HSC activity is still unclear, but preliminary findings show that HLF KO HSC have enhanced ROS levels (WT 337 (±33), KO 510 (±55), p Interestingly, we also found that HLF appears to regulate the identity of HSC by modulating the expression of the SLAM code on the cell surface of the HLF KO HSC. In contrast to the normal frequency of LSK Flt3-CD34- cells, the HLF KO mice displayed a 3.5 fold reduction in the frequency of LSK CD150+CD48- cells (WT 1.94x10-4 (±4.4x10-5)%, KO 0.56x10-4 (±1.5x10-5)%, p10). Strikingly, transplantation of as many as 150 LSK CD150+CD48-HLF KO cells showed a complete lack of repopulating capacity in vivo. This did not correlate to the number of functional HSC seen when transplanting whole bone marrow and indicates that HLF affects the identity of HSC by modulating the expression of the SLAM markers. Taken together, we show here for the first time that HLF has a fundamental role in HSC biology during both fetal and adult hematopoiesis by regulating HSC activity and identity. Disclosures: No relevant conflicts of interest to declare.

Adhesion is a key component of hematopoietic stem cell regulation mediating homing and retention to the niche in the bone marrow. Here, using an RNA interference screen, we identify cytohesin 1 (CYTH1) as a critical mediator of adhesive properties in primary human cord blood–derived hematopoietic stem and progenitor cells (HSPCs). Knockdown of CYTH1 disrupted adhesion of HSPCs to primary human mesenchymal stroma cells. Attachment to fibronectin and ICAM1, 2 integrin ligands, was severely impaired, and CYTH1-deficient cells showed a reduced integrin β1 activation response, suggesting that CYTH1 mediates integrin-dependent functions. Transplantation of CYTH1-knockdown cells to immunodeficient mice resulted in significantly lower long-term engraftment levels, associated with a reduced capacity of the transplanted cells to home to the bone marrow. Intravital microscopy showed that CYTH1 deficiency profoundly affects HSPC mobility and localization within the marrow space and thereby impairs proper lodgment into the niche. Thus, CYTH1 is a novel major regulator of adhesion and engraftment in human HSPCs through mechanisms that, at least in part, involve the activation of integrins.