m6A modulates haematopoietic stem and progenitor cell specification.

N6-methyladenosine (m6A) has been identified as the most abundant modification on eukaryote messenger RNA (mRNA). Although the rapid development of high-throughput sequencing technologies has enabled insight into the biological functions of m6A modification, the function of m6A during vertebrate embryogenesis remains poorly understood. Here we show that m6A determines cell fate during the endothelial-to-haematopoietic transition (EHT) to specify the earliest haematopoietic stem/progenitor cells (HSPCs) during zebrafish embryogenesis. m6A-specific methylated RNA immunoprecipitation combined with high-throughput sequencing (MeRIP-seq) and m6A individual-nucleotide-resolution cross-linking and immunoprecipitation with sequencing (miCLIP-seq) analyses reveal conserved features on zebrafish m6A methylome and preferential distribution of m6A peaks near the stop codon with a consensus RRACH motif. In mettl3-deficient embryos, levels of m6A are significantly decreased and emergence of HSPCs is blocked. Mechanistically, we identify that the delayed YTHDF2-mediated mRNA decay of the arterial endothelial genes notch1a and rhoca contributes to this deleterious effect. The continuous activation of Notch signalling in arterial endothelial cells of mettl3-deficient embryos blocks EHT, thereby repressing the generation of the earliest HSPCs. Furthermore, knockdown of Mettl3 in mice confers a similar phenotype. Collectively, our findings demonstrate the critical function of m6A modification in the fate determination of HSPCs during vertebrate embryogenesis.

Long non-coding RNA SNHG14 aggravates LPS-induced acute kidney injury through regulating miR-495-3p/HIPK1.

Acute kidney injury (AKI) is a complex syndrome with an abrupt decrease of kidney function, which is associated with high morbidity and mortality. Sepsis is the common cause of AKI. Mounting evidence has demonstrated that long non-coding RNAs (lncRNAs) play critical roles in the development and progression of sepsis-induced AKI. In this study, we aimed to illustrate the function and mechanism of lncRNA SNHG14 in lipopolysaccharide (LPS)-induced AKI. We found that SNHG14 was highly expressed in the plasma of sepsis patients with AKI. SNHG14 inhibited cell proliferation and autophagy and promoted cell apoptosis and inflammatory cytokine production in LPS-stimulated HK-2 cells. Functionally, SNHG14 acted as a competing endogenous RNA (ceRNA) to negatively regulate miR-495-3p expression in HK-2 cells. Furthermore, we identified that HIPK1 is a direct target of miR-495-3p in HK-2 cells. We also revealed that the SNHG14/miR-495-3p/HIPK1 interaction network regulated HK-2 cell proliferation, apoptosis, autophagy, and inflammatory cytokine production upon LPS stimulation. In addition, we demonstrated that the SNHG14/miR-495-3p/HIPK1 interaction network regulated the production of inflammatory cytokines (TNF-α, IL-6, and IL-1ß) via modulating NF-κB/p65 signaling in LPS-challenged HK-2 cells. In conclusion, our findings suggested a novel therapeutic axis of SNHG14/miR-495-3p/HIPK1 to treat sepsis-induced AKI.

A Retrospective Population Study to Develop a Predictive Model of Prediabetes and Incident Type 2 Diabetes Mellitus from a Hospital Database in Japan Between 2004 and 2015.

BACKGROUND Type 2 diabetes mellitus is a global public health problem. Prediabetes may be reversed by weight loss, diet, and lifestyle changes. However, without intervention, between 30-50% of individuals with prediabetes develop type 2 diabetes. This retrospective population study was conducted to develop a predictive model of prediabetes and incident type 2 diabetes mellitus using data from 2004 to 2015 from the DRYAD Japanese hospital database. MATERIAL AND METHODS A retrospective longitudinal population study was conducted using the DRYAD database from Murakami Memorial Hospital, Gifu, Japan, to construct a predictive model for prediabetes and incident type 2 diabetes mellitus in the population. Univariate analysis and multivariate analysis were performed to identify the variables that were associated with prediabetes. These variables were used to construct (75% samples) and verify (25% samples) the predictive model. RESULTS From 2004 to 2015, a total of 11,113 cases were identified. Multivariate logistic regression analysis included the six variables of age, waist circumference, smoking history, the presence of fatty liver, fasting blood glucose (FBG), and glycated hemoglobin (HbA1c) level. Data were used to construct (75% samples) and verify (25% samples) in a predictive model. The area under the receiver operating characteristic (ROC) curve (AUC) of the predictive model was 0.87 (0.85-0.89) in the training cohort and 0.87 (0.86-0.90) in the validation cohort. CONCLUSIONS A prognostic model based on six variables was predictive for incident type 2 diabetes mellitus and prediabetes in a healthy population in Japan.

SOFA score is superior to APACHE-II score in predicting the prognosis of critically ill patients with acute kidney injury undergoing continuous renal replacement therapy.

BACKGROUND: Acute kidney injury (AKI) is the most common cause of organ failure in multiple organ dysfunction syndrome (MODS) and is associated with increased mortality. This study aimed at determining the efficacy of sequential organ failure assessment (SOFA), and acute physiology and chronic health evaluation II (APACHE-II) scoring systems in assessing the prognosis of critically ill patients with AKI undergoing CRRT. METHODS: The predictive value of SOFA and APACHE-II scores for 28- and 90-d mortality in patients with AKI undergoing continuous renal replacement therapy (CRRT) were determined by multivariate analysis, sensitivity analysis, and curve-fitting analysis. RESULTS: A total of 836 cases were included in this study. Multivariate Cox logistic regression analysis showed that SOFA scores were associated with 28- and 90-d mortality in patients with AKI undergoing CRRT. The adjusted HR of SOFA for28-d mortality were 1.18 (1.14, 1.21), 1.24 (1.18, 1.31), and 1.19 (1.13, 1.24) in the three models, respectively, and the adjusted HR of SOFA for 90-d mortality was 1.12 (1.09, 1.16), 1.15 (1.10, 1.19), and 1.15 (1.10, 1.19), respectively. The subgroup analysis showed that the SOFA score was associated with 28-d and 90-d mortality in patients with AKI undergoing CRRT. APACHE-II score was not associated with 28- and 90-d mortality patients with AKI undergoing CRRT. Curve fitting analysis showed that SOFA scores increased had a higher prediction accuracy for 28- and 90-d than APACHE-II. CONCLUSIONS: The SOFA score showed a higher accuracy of mortality prediction in critically ill patients with AKI undergoing CRRT than the APACHE-II score.

Higher body mass index is not a protective risk factor for 28-days mortality in critically ill patients with acute kidney injury undergoing continuous renal replacement therapy.

Background: Acute kidney injury (AKI) requiring continuous renal replacement therapy (CRRT) is a fatal and common clinical disorder in critically ill patients. Recent studies have shown that the relationship between BMI and the outcome of patients with AKI undergoing CRRT is conflicting. Methods: A retrospective cohort study based on data reuse. Univariate analysis, multi-factor regression analysis and subgroup analyses were used to explore the association of the BMI with the 28-days mortality risk in patients with AKI undergoing CRRT. Results: From January 2009 to September 2016, a total of 1120 cases met the inclusion criteria and were enrolled in this study. The univariate analysis showed that BMI was associated with 28-days mortality of patients with AKI undergoing CRRT, its HR value was 0.98 (0.96, 0.99). The multi-factor regression analysis showed that BMI was not associated with 28-days mortality of patients with AKI undergoing CRRT in the four models, the adjusted HR value of four models were 1.00 (0.96, 1.04), 1.01 (0.97, 1.04), 1.00 (0.96, 1.04) and 1.00 (0.96, 1.04), respectively. The subgroups analyses showed that the BMI was a risk factor of the 28-days mortality in patients with AKI undergoing CRRT when GFR ≥30 mL/min, its HR value was 1.04 (1.01, 1.09). Conclusion: Higher BMI was not a protective risk of 28-day mortality in patients with AKI undergoing CRRT. Especially, when GFR ≥30 mL/min, higher BMI increased the risk of the 28-day mortality rate in patients with AKI undergoing CRRT.

Inflammatory signaling regulates hematopoietic stem and progenitor cell emergence in vertebrates.

Inflammatory signaling has been shown to be essential for stress hematopoiesis in adult bone marrow, either through increasing proliferation or by directing differentiation of hematopoietic stem and progenitor cells (HSPCs) toward myeloid or lymphoid lineages. However, its role in embryonic normal hematopoiesis has been unknown. Here, we demonstrate that in both zebrafish and mouse embryos, inflammatory signaling is necessary and sufficient for HSPC emergence, in the absence of infection or pathological inflammation. Mechanistically, inflammatory signaling regulates hemogenic endothelium-derived HSPC development through a conserved Toll-like receptor 4 (TLR4)-nuclear factor κ-light-chain enhancer of activated B core (NF-κB) signaling, which then promotes Notch activity, a well-known signal required for HSPC specification in vertebrates. Our findings establish a previously unrecognized link between inflammatory signaling and HSPC emergence, and provide new insights into regenerative medicine and novel therapies to treat innate immune-related diseases.

[Effect of litchi seed aqueous extracts on learning and memory obstacles induced by D-galactose in mice and its mechanism].

OBJECTIVE: To investigate the improving effect of Litchi Seed Aqueous Extracts on learning and memory obstacles model and its mechanism. METHODS: The learning and memory obstacles model was incluced by subcutoneous injection of D-galactose (500 mg/kg) for 8 weeks. The model group and treatment groups were given huperzine A (0.4 mg/kg) and Litchi Seed Aqueous Extracts (15,60 g/kg) respectively for 4 weeks by ig at the 5th week. After huperzine A and Litchi Seed Aqueous Extracts treatment for 4 weeks, water maze test was used to determine the ability of mice's spatial learning and memory. The contents of advanced glycation end products (AGEs) in serum, the content of nitric oxide (NO) and acetylcholine (Ach), the activity of nitric oxide synthase (NOS) and acetylcholinesterase (AchE) in the brain tissue were detected. RESULTS: Litchi Seed Aqueous Extracts significantly ameliorated the learning and memory ability in mice, decreased the level of AGEs in serum, and reduced the content of NO and activity of NOS in brain tissues. No significantly influence was observed for the Ach and Ach-E in brain tissues. CONCLUSION: Litchi Seed Aqueous Extracts possesses improving the learning and memory effects on the model mice induced by D-galactose, which may be related to inhibiting too much AGEs and NO formation and reducing damage in the brain cells.

Pyroptosis regulators exert crucial functions in prognosis, progression and immune microenvironment of pancreatic adenocarcinoma: a bioinformatic and in vitro research.

Pyroptosis is an inflammatory programmed cell death, showing potentials to be a novel anti-cancer approach. However, the roles of pyroptosis-related (PR) genes (PRGs) in pancreatic adenocarcinoma (PAAD) remain elusive. In the present study, we constructed a novel PR risk signature through the lasso regression analysis. The risk signature was greatly conducive to PAAD prognostic assessment. PR risk score was identified as an independent prognostic factor and could distinguish the prognostic differences of most clinical subgroups. Meanwhile, it could improve the traditional prognostic models based on TNM-staging. Next, its prognostic value was also tested in five validation cohorts. Using CIBERSORT, ESTIMATE, and ssGSEA algorithms, the effects of PR risk signature on tumor immune microenvironment (TIM) were explored. High PR risk suppressed antitumor immune through decreasing the infiltrating levels of CD8 T and NK cells. The genomic information and histological expression of risk PRGs were uncovered by USCA and HPA databases. Somatic mutation, methylation alteration, and homozygous CNV of eight PRGs barely occurred in PAAD samples. As for therapeutic correlation, PR risk score may not predict the efficacy of PD-1/L1 inhibitors and was weakly associated with multiple drug susceptibilities. Finally, the biofunctions of toll like receptor 3 (TLR3) in pancreatic cancer (PC) cells were investigated through qPCR, MTT, colony formation, and Transwell assays. Overexpression of TLR3 could promote the proliferation, migration, and invasion of PC cells. In conclusion, PRGs play crucial roles in prognosis, progression, and immune microenvironment of PAAD. TLR3 is expected to be a promising therapeutic target.

The rate of acute kidney injury (AKI) alert detection by the attending physicians was associated with the prognosis of patients with AKI.

Introduction: Early identification of AKI was always considered to improve patients' prognosis. Some studies found that AKI early warning tools didn't affect patients' prognosis. Therefore, additional studies were necessary to explore the reasons. Methods: This study was a secondary analysis of a multicenter randomized controlled trial that found electronic health record warnings for AKI did not influence patients' prognoses. Univariate, multivariate, subgroup, curve fitting, and threshold effect analysis were used to explore the association between AKI warnings detected by attending physicians and the patient's prognosis. Results: A total of 6,030 AKI patients were included in the study. The patients were classified into two groups based on the rate of AKI alerts detected by attending physicians: the partial group (n = 5,377), and the complete group (n = 653). In comparison to the partial group, the complete group significantly decreased 14-day AKI progression, 14-day dialysis, and 14-day mortality, with adjusted ORs of 0.48 (0.33, 0.70), 0.26 (0.09, 0.77), and 0.53 (0.33, 0.84) respectively, and the complete group significantly improve the discharge to home, with an OR value of 1.50 (1.21, 1.87). When the rate of AKI alerts detected by the attending physicians as a continuity variable, we found that the rate of alerts seen by attending physicians was associated with 14-day mortality and the discharge to home, with adjusted ORs of 1.76 (1.11, 2.81) and 1.42 (1.13, 1.80). The sensitivity analysis, curve-fitting analysis, and threshold effect analysis also showed that the rate of alert seen by the attending physician was correlated with the patient's prognosis. Conclusion: The rate of AKI alert detection by attending physician were related to the patient's prognosis. The higher the rate of AKI alert detection by attending physicians, the better the prognosis of patients with AKI.

[Effects and mechanism of berberine on the hypertensive renal injury rats induced by enriched high fat-salt-fructose diet].

OBJECTIVE: To study the effects of berberine on hypertensive renal injury model in rats fed by enriched high fat-salt-fructose diet . METHODS: hypertensive renal injury model was esteblished by feeding enriched high fat-salt-fructose diet for 8 weeks. On the basis of animal blood pressure, hypertensive rats were randomly divided into model group (10 rats, distilled water), captopril group (10 rats,25 mg/kg), berberine high dose group (10 rats, 300 mg/kg) and low dose group (10 rats, 100 mg/kg). These rats were fed by enriched high fat-salt-fructose diet and treated by intragastric administration with drugs for 4 weeks. And normal control group (10 rats) was set Blood pressure was determined at 0, 4, 8, 10, 12 weekend,and after 4 weeks of drugs treatment, getting urine to determine urine protein, taking blood serum to determine blood urea nitrogen, serum creatinine,GHb,MDA and activity of SOD. The content of H2O2 and GSH-Px and activity of CAT in kidney tissues were determined also. RESULTS: Compared with normal control group, blood pressure, urine protein, blood urea nitrogen, serum creatinine and MDA and GHb in serum of model rats obviously increased (P < 0.01), the activity of SOD decreased (P < 0.01), higher content of H2O2 and lower content of GSH-Px and activity of CAT (P < 0.01) in the kidney tissues. Treated with berberine for 4 weeks, elevated blood pressure and heightened levels of urine protein, blood urea nitrogen and serum creatinine in model rats were depressed significantly (P < 0.01), and elevated the activity of SOD, lowed the levels of MDA and GHb in blood serum (P < 0.01). At the same time, berberine increased the activities of GSH-Px and CAT (P < 0.01) and slightly lowed the content of H2O2 in the kidney tissues. CONCLUSION: Berberine has protecting effects on the hypertensive renal impairment model rats fed by enriched high fat-salt-fructose diet, which are concerned with elevated antioxidant capability in body and kidney tissues.

Predictors of Mortality in Hospitalized COVID-19 Patients Complicated With Hypotension and Hypoxemia: A Retrospective Cohort Study.

Introduction: COVID-19 patients with hypotension and hypoxemia had a significantly worse outcome. The purpose of this research was to ascertain the risk factors affecting the prognoses of these patients and to develop appropriate prognostic prediction tools. Methods: From March 1, 2020, to April 16, 2020, a retrospective cohort analysis of COVID-19 patients with hypotension and hypoxemia was performed. The univariate and multivariate analyses were performed to identify the associated risk factors influencing the prognosis of COVID-19 patients with hypotension and hypoxemia, and the selected variables were then utilized to construct and validate the prediction model for these patients. Results: Three hundred and twenty-seven COVID-19 patients with hypotension and hypoxemia who met the inclusion and exclusion criteria were included in this study. Age, temperature, troponin, and blood glucose were related to mortality in COVID-19 patients with hypotension and hypoxemia in both univariate and multivariate analyses. The MFP model (multiple fractional polynomial model), full model, and stepwise model were utilized to build the prediction model, and their AUCs were, respectively, 0.902 (0.868, 0.936), 0.902 (0.868, 0.936), and 0.902 (0.868, 0.936). Because the sample size for this research was limited, we utilized bootstrapping for internal validation. The AUCs of Bootstrap full and Bootstrap stepwise were 0.902 (0.867, 0.936) and 0.902 (0.868, 0.936), respectively. Conclusion: Age, temperature, troponin, and blood glucose levels were associated with mortality in COVID-19 patients with hypotension and hypoxemia. Additionally, the prediction model developed using the variables above showed a high predictive value for predicting the prognosis of these individuals.

Serum Albumin Before CRRT Was Associated With the 28- and 90-Day Mortality of Critically Ill Patients With Acute Kidney Injury and Treated With Continuous Renal Replacement Therapy.

Introduction: Although low serum albumin (ALB) may worsen acute kidney injury (AKI), additional study is needed to establish the connection between ALB and the prognosis of critically ill patients with AKI and treated with continuous renal replacement therapy (CRRT). Methods:A secondary analysis of a bi-center, retrospective, and observational study, such as critically ill patients with AKI and treated with CRRT from January 2009 to September 2016. The univariate analysis, multi-factor regression analysis, sensitivity analysis, and curve-fitting analysis were applied to explore the association of ALB with the 28 and 90 days mortality of critically ill patients with AKI and treated with CRRT, and the removal efficiency of serum phosphorus. Results: From January 2009 to September 2016, 1,132 cases with AKI and treated with CRRT met the inclusion criteria and enrolled in this study. We found that the higher ALB before CRRT, the lower the 28- and 90-day mortality of patients with AKI and treated with CRRT, the higher removal efficiency of serum phosphorus, the adjusted hazard ratio (HR) value for 28-day mortality in the four models were separately 0.92 (0.90, 0.95), 0.91 (0.89, 0.94), 0.92 (0.89, 0.95), and 0.92 (0.89, 0.95); the adjusted HR value for 90 day mortality in the four models were 0.91 (0.89, 0.94), 0.92 (0.89, 0.95), 0.92 (0.89, 0.95), and 0.92 (0.89, 0.96); the adjusted OR value for the removal efficiency of serum phosphorus in the four models were separately -0.04 (-0.07, -0.01), -0.05 (-0.08, -0.01), -0.04 (-0.08, -0.01), and -0.04 (-0.08, -0.01). The sensitivity analysis and curve-fitting analysis also showed that ALB before CRRT was correlated with the 28 and 90 days mortality of critically ill patients with AKI and treated with CRRT and the removal efficiency of serum phosphorus. Conclusion: The higher the serum ALB before CRRT, the lower the mortality of critically ill patients with AKI and treated with CRRT, and the higher the clearance efficiency of serum phosphorus.

Glaucocalyxin A Protects H9c2 Cells Against Hypoxia/Reoxygenation-Induced Injury Through the Activation of Akt/Nrf2/HO-1 Pathway.

Myocardial infarction (MI) is one of the most serious cardiovascular diseases associated with myocardial ischemia/reperfusion (I/R) injury. Glaucocalyxin A (GLA) is a biologically active ent-kauranoid diterpenoid that has been found to ameliorate myocardial I/R injury in mice. However, the mechanism has not been fully investigated. In the present study, we aimed to investigate the effect of GLA on rat cardiomyocytes H9c2 cells exposed to hypoxia/reoxygenation (H/R). The results showed that GLA treatment improved cell viability of H/R-stimulated H9c2 cells. Administration with GLA suppressed the H/R-stimulated reactive oxygen species (ROS) production in H9c2 cells. GLA also elevated the activities of antioxidant enzymes, including superoxide dismutase and glutathione peroxidase in H/R-stimulated H9c2 cells. Moreover, GLA prevented H/R-stimulated cell apoptosis in H9c2 cells, as evidenced by increased bcl-2 expression, decreased bax expression, as well as reduced caspase-3 activity. Furthermore, GLA enhanced the activation of protein kinase B (Akt)/nuclear factor erythroid 2-related factor 2 (Nrf2)/heme oxygenase-1 (HO-1) signaling pathway in H9c2 cells exposed to H/R. Additionally, treatment with LY294002 reserved the protective effects of GLA on H/R-stimulated oxidative injury in H9c2 cells. In conclusion, these findings suggested that GLA protected H9c2 cells from H/R-stimulated oxidative damage, which was mediated by the Akt/Nrf2/HO-1 signaling pathway. Thus, GLA might be a promising therapeutic agent for the prevention and treatment of myocardial I/R.

[Effects of total glucosides of paeony on enhancing insulin sensitivity and antagonizing nonalcoholic fatty liver in rats].

OBJECTIVE: To study the pathological changes of blood glucose, serum lipid, insulin resistance, liver function, liver cell denaturalization of total glucosides of paeony on nonalcoholic fatty liver rats caused by insulin resistance and discuss the acting mechanism. METHOD: Adult SD rats were maintained on high-fat-sugar-salt diet for 56 days. In the 57th day, their fasting blood glucose (FBG) and 2-hours blood glucose after oral glucose tolerance test (OGTT-2 hBG) were mensurated, according to which and the weight the rats were divided randomly into nonalcoholic fatty liver model group, metformin group (0.2 g x kg(-1)) and total glucosides of paeony group (high dosage 0.15 g x kg(-1), low dosage 0.05 g x kg(-1)). All the rats were still administered the same diet and given different drugs by intragastric administration for 28 days. In the 29th day, all of them were killed and the blood was sampled to measure the levels of blood glucose [FBG, OGTT-2 hBG, fasting insulin (Fins)] and serum lipid [free fatty acids (FFA), triglyceride (TG), total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C) and high-density lipoprotein cholesterol (HDL-C)], then the HOMA insulin resistance index (HOMA-IRI, fasting glucosexinsulin) and insulin sensitivity index (ISI) were counted. The activities of aspartate aminotransferase (AST), alanine aminotransferase (ALT), cholinesterase (ChE), superoxide dismutase (SOD) and the contents of malondialdehyde (MDA) were measured also. Livers were weighed and collected to be observed the pathological changes. RESULT: Compared with normal group, in nonalcoholic fatty liver model group the levels of Fins and IRI were increased obviously (P < 0.01), ISI were decreased (P < 0.01), FFA, TG, TC, LDL-C were increased (P < 0.01), HDL-C were decreased (P < 0.05); the content of MDA were increased (P < 0.05), the activities of SOD were decreased (P < 0.01); AST, ALT and ChE were increased (P < 0.05, or P < 0.01), the pathological changes of liver fat were severe (P < 0.01). In glucosides of paeony group and metformin group, hyperinsulinaemia and insulin resistence were resisted (P < 0.05, or P < 0.01); the levels of FFA, TG, TC, LDL-C were decreased and HDL-C were increased (P < 0.05, or P < 0.01); the activities of AST, ALT, ChE were decreased (P < 0.05, or P < 0.01) and SOD were increased (P < 0.01). The contents of MDA were decreased (P < 0.05). The levels of FBG and 2 hBG in metformin group were decreased but in total glucosides of paeony group were not decreased obviously. CONCLUSION: Total glucosides of paeony may protect liver function and modulate serum lipid for the fatty liver rats caused by insulin resistance, and its action mechanism may be concerned with enhancing insulin sensitivity and antioxidative ability, decreasing serum lipid.

[Effect of Semen Litchi containing serum on proliferation and apoptosis of HepG2 cells].

OBJECTIVE: To observe the effect of Semen Litchi containing serum on proliferation and apoptosis of HepG2 cells. METHODS: The Semen litchi or CTX containing serum and control serum were prepared by serologic pharmacology method. MTT assay was used to observe the proliferation inhibition rate of HepG2 cells after incubated with different kinds of drug's containing serum. Nuclear morphological features of HepG2 cells were detected by fluorescencemicroscopy after staining with Hochest33258. The apoptosis rate of HepG2 cells in each group was detected by flow cytometry. RESULTS: The cell viability and the apoptosis rate of HepG2 cells in Semen Litchi containing serum groups were higher than that of control group, and the results of fluorescencemicroscopy observation showed the nuclear morphological change of apoptosis. CONCLUSION: Semen Litchi can inhibit the proliferation of HepG2 cells, the acting mechanism may be concerned with cell apoptosis.

Application of Aorta-gonad-mesonephros Explant Culture System in Developmental Hematopoiesis.

The limitation of using mouse embryos for hematopoiesis studies is the added inconvenience in operations, which is largely due to the intrauterine development of the embryo. Although genetic data from knockout (KO) mice are convincing, it is not realistic to generate KO mice for all genes as needed. In addition, performing in vivo rescue experiments to consolidate the data obtained from KO mice is not convenient. To overcome these limitations, the Aorta-Gonad-Mesonephros (AGM) explant culture was developed as an appropriate system to study hematopoietic stem cell (HSC) development. Especially for rescue experiments, it can be used to recover the impaired hematopoiesis in KO mice. By adding the appropriate chemicals into the medium, the impaired signaling can be reactivated or up-regulated pathways can be inhibited. With the use of this method, many experiments can be performed to identify the critical regulators of HSC development, including HSC related gene expression at mRNA and protein levels, colony formation ability, and reconstitution capacity. This series of experiments would be helpful in defining the underlying mechanisms essential for HSC development in mammals.

The Role of Serotonin beyond the Central Nervous System during Embryogenesis.

Serotonin, or 5-hydroxytryptamine (5-HT), is a well-known neurotransmitter that plays vital roles in neural activities and social behaviors. Clinically, deficiency of serotonin is linked with many psychiatric disorders. Interestingly, a large proportion of serotonin is also produced outside the central nervous system (CNS). There is increasing evidence demonstrating important roles of serotonin in the peripheral tissues. Here, we will describe the multiple biological functions of serotonin in hematopoietic system, such as development of hematopoietic stem and progenitor cells (HSPCs), differentiation of hematopoietic cells, maintenance of vascular system, and relationship with hematological diseases. The roles of serotonin in inflammatory responses mediated by hematopoietic cells as well as in liver regeneration are also discussed. Our recent understandings of the impact of serotonin on hematopoietic system, immune responses, and tissue regeneration support utilization of serotonin as a potential therapeutic target for the treatment of hematological diseases and organ repair in clinic.

The Vascular Niche Regulates Hematopoietic Stem and Progenitor Cell Lodgment and Expansion via klf6a-ccl25b.

In mammals, hematopoietic stem and progenitor cells (HSPCs) rapidly expand in the fetal liver (FL), but the underlying mechanism remains unclear. Here, we characterize zebrafish caudal hematopoietic tissue (CHT) and identify an important cellular and molecular mechanism of HSPC expansion. Time-lapse imaging showed that HSPCs localize adjacent to vascular endothelial cells (ECs), and their migration and expansion display caudal vein-specific orientation in the CHT. RNA sequencing and functional analysis identified that an EC-expressed transcription factor, Krüppel-like factor 6a (Klf6a), is essential for the CHT niche. We further demonstrated that Klf6a directly regulates the expression of the chemokine (C-C motif) ligand 25b to modulate HSPC lodgment and proliferation. Ex vivo culture results support the conserved role of Ccl21/Ccr7 signaling in promoting HSPC expansion in mammals. Together, we identify the Klf6a-Ccl25b/Ccr7 axis in controlling the complex HSPC-CHT niche interaction, which may be applicable to in vitro expansion or engraftment of HSPCs after transplantation.

BLOS2 maintains hematopoietic stem cells in the fetal liver via repressing Notch signaling.

During development, hematopoietic stem cells (HSCs) undergo a rapid expansion in the fetal liver (FL) after their emergence in the aorta-gonad-mesonephros (AGM) region. We recently reported that the endolysosomal trafficking factor BLOS2, encoded by the Bloc1s2 gene, regulates HSC/hematopoietic progenitor cell emergence in the AGM region; however, whether it plays a role in the FL remains unknown. Here, we show that BLOS2 plays an essential role in the regulation of HSC proliferation and differentiation in the FL. Bloc1s2 depletion leads to elevated Notch signaling, with an increased frequency but weakened self-renewal ability of FL HSCs. Functional assays show that Bloc1s2-/- FL HSCs harbor impaired lymphoid and myeloid differentiation abilities. These findings reveal that balanced control of Notch signaling by BLOS2 is required for HSC homeostasis during FL hematopoiesis.

5-hydroxytryptamine synthesized in the aorta-gonad-mesonephros regulates hematopoietic stem and progenitor cell survival.

The in vitro or ex vivo production of transplantable hematopoietic stem cells (HSCs) holds great promise for the treatment of hematological diseases in the clinic. However, HSCs have not been produced from either embryonic or induced pluripotent stem cells. In this study, we report that 5-hydroxytryptamine (5-HT; also called serotonin) can enhance the generation of hematopoietic stem and progenitor cells (HSPCs) in vitro and is essential for the survival of HSPCs in vivo during embryogenesis. In tryptophan hydroxylase 2-deficient embryos, a decrease in 5-HT synthesized in the aorta-gonad-mesonephros leads to apoptosis of nascent HSPCs. Mechanistically, 5-HT inhibits the AKT-Foxo1 signaling cascade to protect the earliest HSPCs in intraaortic hematopoietic clusters from excessive apoptosis. Collectively, our results reveal an unexpected role of 5-HT in HSPC development and suggest that 5-HT signaling may be a potential therapeutic target for promoting HSPC survival.