An improved DIIS method using a versatile residual matrix to accelerate SCF starting from a crude guess.

The minimization of the commutator of the Fock and density matrices as the error matrix in the direct inversion of the iterative subspace (CDIIS) developed by Pulay is a powerful self-consistent field (SCF) acceleration technique for the construction of optimum Fock matrix, if initiated with a fair initial guess. In this work, we present an alternative minimized error matrix to the commutator in the CDIIS, namely the residual or the gradient of the energy-functional for a Slater determinant subject to the orthonormality constraints among orbitals, representing the search for a newly improved Fock matrix in the direction of the residual in the direct inversion of the iterative subspace (RDIIS). Implemented in the computational chemistry package GAMESS, the RDIIS is compared with the standard CDIIS and the second order SCF orbital optimization (SOSCF) for tested molecules started with a crude guess. As a result, the RDIIS stably and efficiently performs the SCF convergence acceleration. Furthermore, the RDIIS is considerably independent on the subspace size with the concentrated linear coefficients accounting proportionally for the Fock matrices close to the current iteration.

CEA cell adhesion molecule 5 enriches functional human hematopoietic stem cells capable of long-term multi-lineage engraftment.

Hematopoietic stem cell (HSC) surface markers improve the understanding of cell identity and function. Here, we report that human HSCs can be distinguished by their expression of the CEA Cell Adhesion Molecule 5 (CEACAM5, CD66e), which serves as a marker and a regulator of HSC function. CD66e+ cells exhibited a 5.5-fold enrichment for functional long term HSCs compared to CD66e- cells. CD66e+CD34+CD90+CD45RA- cells displayed robust multi-lineage repopulation and serial reconstitution ability in immunodeficient mice compared to CD66e-CD34+CD90+CD45RA-cells. CD66e expression also identified almost all repopulating HSCs within the CD34+CD90+CD45RA- population. Together, these results indicated that CEACAM5 is a marker that enriches functional human hematopoietic stem cells capable of long-term multi-lineage engraftment.

Single-cell dissection of human hematopoietic reconstitution after allogeneic hematopoietic stem cell transplantation.

Hematopoietic stem cell transplantation is an effective regenerative therapy for many malignant, inherited, or autoimmune diseases. However, our understanding of reconstituted hematopoiesis in transplant patients remains limited. Here, we uncover the reconstitution dynamics of human allogeneic hematopoietic stem and progenitor cells (HSPCs) at single-cell resolution after transplantation. Transplanted HSPCs underwent rapid and measurable changes during the first 30 days after transplantation, characterized by a strong proliferative response on the first day. Transcriptomic analysis of HSPCs enabled us to observe that immunoregulatory neutrophil progenitors expressing high levels of the S100A gene family were enriched in granulocyte colony-stimulating factor-mobilized peripheral blood stem cells. Transplant recipients who developed acute graft-versus-host disease (aGVHD) infused fewer S100Ahigh immunoregulatory neutrophil progenitors, immunophenotyped as Lin-CD34+CD66b+CD177+, than those who did not develop aGVHD. Therefore, our study provides insights into the regenerative process of transplanted HSPCs in human patients and identifies a potential criterion for identifying patients at high risk for developing aGVHD early after transplant.

Temporal molecular program of human hematopoietic stem and progenitor cells after birth.

Hematopoietic stem and progenitor cells (HSPCs) give rise to the blood system and maintain hematopoiesis throughout the human lifespan. Here, we report a transcriptional census of human bone-marrow-derived HSPCs from the neonate, infant, child, adult, and aging stages, showing two subpopulations of multipotent progenitors separated by CD52 expression. From birth to the adult stage, stem and multipotent progenitors shared similar transcriptional alterations, and erythroid potential was enhanced after the infant stage. By integrating transcriptome, chromatin accessibility, and functional data, we further showed that aging hematopoietic stem cells (HSCs) exhibited a bias toward megakaryocytic differentiation. Finally, in comparison with the HSCs from the cord blood, neonate bone-marrow-derived HSCs were more quiescent and had higher long-term regeneration capability and durable self-renewal. Taken together, this work provides an integral transcriptome landscape of HSPCs and identifies their dynamics in post-natal steady-state hemopoiesis, thereby helping explore hematopoiesis in development and diseases.

Red light-transmittance bagging promotes carotenoid accumulation of grapefruit during ripening.

Light, a crucial environmental signal, is involved in the regulation of secondary metabolites. To understand the mechanism by which light influences carotenoid metabolism, grapefruits were bagged with four types of light-transmitting bags that altered the transmission of solar light. We show that light-transmitting bagging induced changes in carotenoid metabolism during fruit ripening. Compared with natural light, red light (RL)-transmittance treatment significantly increases the total carotenoid content by 62%. Based on weighted gene co-expression network analysis (WGCNA), 'blue' and 'turquoise' modules are remarkably associated with carotenoid metabolism under different light treatment (p < 0.05). Transcriptome analysis identifies transcription factors (TFs) bHLH128, NAC2-like/21/72, MYB-like, AGL11/AGL61, ERF023/062, WRKY20, SBPlike-7/13 as being involved in the regulation of carotenoid metabolism in response to RL. Under RL treatment, these TFs regulate the accumulation of carotenoids by directly modulating the expression of carotenogenic genes, including GGPPS2, PDS, Z-ISO, ZDS2/7, CRTISO3, CYP97A, CHYB, ZEP2, CCD1-2. Based on these results, a network of the regulation of carotenoid metabolism by light in citrus fruits is preliminarily proposed. These results show that RL treatments have great potential to improve coloration and nutritional quality of citrus fruits.

New insights into Human Hematopoietic Stem and Progenitor Cells via Single-Cell Omics.

Residing at the apex of the hematopoietic hierarchy, hematopoietic stem and progenitor cells (HSPCs) give rise to all mature blood cells. In the last decade, significant progress has been made in single-cell RNA sequencing as well as multi-omics technologies that have facilitated elucidation of the heterogeneity of previously defined human HSPCs. From the embryonic stage through the adult stage to aging, single-cell studies have enabled us to trace the origins of hematopoietic stem cells (HSCs), demonstrating different hematopoietic differentiation during development, as well as identifying novel cell populations. In both hematological benign diseases and malignancies, single-cell omics technologies have begun to reveal tissue heterogeneity and have permitted mapping of microenvironmental ecosystems and tracking of cell subclones, thereby greatly broadening our understanding of disease development. Furthermore, advances have also been made in elucidating the molecular mechanisms for relapse and identifying therapeutic targets of hematological disorders and other non-hematological diseases. Extensive exploration of hematopoiesis at the single-cell level may thus have great potential for broad clinical applications of HSPCs, as well as disease prognosis.

Clonal phylogeny and evolution of critical cytogenetic aberrations in multiple myeloma at single-cell level by QM-FISH.

Single-cell analysis is of significant importance in delineating the exact phylogeny of the subclonal population and in discovering subtle diversification. So far, studies of intratumor heterogeneity and clonal evolution in multiple myeloma (MM) were largely focused on the bulk tumor population level. We performed quantitative multigene fluorescence in situ hybridization (QM-FISH) in 129 longitudinal samples of 57 MM patients. All the patients had newly diagnosed and relapsed paired samples. An expanded cohort of 188 MM patients underwent conventional FISH (cFISH) to validate the cytogenetic evolution in bulk tumor level. Forty-three of 57 patients (75.4%) harbored 3 or 4 cytogenetic clones at diagnosis. We delineated the phylogeny of the subclonal tumor population and derived the evolutionary architecture in each patient. Patients with clonal stabilization had a significantly improved overall survival (OS) than those with other evolutionary patterns (median OS, 71.2 months vs 39.7 months vs 35.2 months vs 25.5 months, for stable, differential, branching, and linear patterns, respectively; P = .001). A high degree of consistency and complementarity across QM-FISH and cFISH was observed in the evaluation of cytogenetic evolution patterns in MM. Survival after relapse was greater influenced by the presence of high-risk aberrations at relapse (hazard ratio = 2.07) rather than present at diagnosis (hazard ratio = 1.55). This study shows that QM-FISH is a valuable tool to elucidate the clonal architecture at the single-cell level. Clonal evolution pattern is of prognostic significance, highlighting the need for repeated cytogenetic evaluation in relapsed MM.

Single-cell transcriptomic landscape of human blood cells.

High throughput single-cell RNA-seq has been successfully implemented to dissect the cellular and molecular features underlying hematopoiesis. However, an elaborate and comprehensive transcriptome reference of the whole blood system is lacking. Here, we profiled the transcriptomes of 7551 human blood cells representing 32 immunophenotypic cell types, including hematopoietic stem cells, progenitors and mature blood cells derived from 21 healthy donors. With high sequencing depth and coverage, we constructed a single-cell transcriptional atlas of blood cells (ABC) on the basis of both protein-coding genes and long noncoding RNAs (lncRNAs), and showed a high consistence between them. Notably, putative lncRNAs and transcription factors regulating hematopoietic cell differentiation were identified. While common transcription factor regulatory networks were activated in neutrophils and monocytes, lymphoid cells dramatically changed their regulatory networks during differentiation. Furthermore, we showed a subset of nucleated erythrocytes actively expressing immune signals, suggesting the existence of erythroid precursors with immune functions. Finally, a web portal offering transcriptome browsing and blood cell type prediction has been established. Thus, our work provides a transcriptional map of human blood cells at single-cell resolution, thereby offering a comprehensive reference for the exploration of physiological and pathological hematopoiesis.

Radiation-induced bystander effects impair transplanted human hematopoietic stem cells via oxidative DNA damage.

Total body irradiation (TBI) is commonly used in host conditioning regimens for human hematopoietic stem cell (HSC) transplantation to treat various hematological disorders. Exposure to TBI not only induces acute myelosuppression and immunosuppression, but also injures the various components of the HSC niche in recipients. Our previous study demonstrated that radiation-induced bystander effects (RIBE) of irradiated recipients decreased the long-term repopulating ability of transplanted mouse HSCs. However, RIBE on transplanted human HSCs have not been studied. Here, we report that RIBE impaired the long-term hematopoietic reconstitution of human HSCs as well as the colony-forming ability of human hematopoietic progenitor cells (HPCs). Our further analyses revealed that the RIBE-affected human hematopoietic cells showed enhanced DNA damage responses, cell-cycle arrest, and p53-dependent apoptosis, mainly because of oxidative stress. Moreover, multiple antioxidants could mitigate these bystander effects, though at different efficacies in vitro and in vivo. Taken together, these findings suggest that RIBE impair human HSCs and HPCs by oxidative DNA damage. This study provides definitive evidence for RIBE on transplanted human HSCs and further justifies the necessity of conducting clinical trials to evaluate different antioxidants to improve the efficacy of HSC transplantation for the patients with hematological or nonhematological disorders.

Correction: PDGFB-expressing mesenchymal stem cells improve human hematopoietic stem cell engraftment in immunodeficient mice.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Cytoplasmic p27 is a novel prognostic biomarker and oncogenic protein for nasopharyngeal carcinoma.

Cytoplasmic p27 plays an important role in regulating the cell cycle. Recent studies have revealed p27 protein translocation from the nucleus to the cytoplasm in many tumour cells. The aim of this study was to investigate the role and molecular mechanisms of cytoplasmic p27 in the progression of nasopharyngeal carcinoma (NPC) and to explore its prognostic value. We found increased cytoplasmic p27 expression by immunohistochemistry in NPC tissues, and its expression level was significantly correlated with the T classification and TNM clinical stage of NPC. The survival rate was significantly lower for NPC patients with cytoplasmic p27 immunopositivity than for NPC patients with cytoplasmic p27 immunonegativity, and cytoplasmic p27 was an independent risk factor that affected the prognosis of patients with NPC. Cytoplasmic p27 promoted the proliferation, cell cycle progression, migration, and invasion of NPC cells, increased Bim-1 and Twist1 protein levels, and decreased RhoA-GTP level. Collectively, these findings suggest that cytoplasmic relocalization of p27 is involved in the pathogenesis of NPC and is closely related to the unfavourable prognosis of patients with NPC. Therefore, cytoplasmic p27 might be a useful prognostic factor and potential therapeutic target for patients with NPC.

PDGFB-expressing mesenchymal stem cells improve human hematopoietic stem cell engraftment in immunodeficient mice.

The bone marrow (BM) niche regulates multiple hematopoietic stem cell (HSC) processes. Clinical treatment for hematological malignancies by HSC transplantation often requires preconditioning via total body irradiation, which severely and irreversibly impairs the BM niche and HSC regeneration. Novel strategies are needed to enhance HSC regeneration in irradiated BM. We compared the effects of EGF, FGF2, and PDGFB on HSC regeneration using human mesenchymal stem cells (MSCs) that were transduced with these factors via lentiviral vectors. Among the above niche factors tested, MSCs transduced with PDGFB (PDGFB-MSCs) most significantly improved human HSC engraftment in immunodeficient mice. PDGFB-MSC-treated BM enhanced transplanted human HSC self-renewal in secondary transplantations more efficiently than GFP-transduced MSCs (GFP-MSCs). Gene set enrichment analysis showed increased antiapoptotic signaling in PDGFB-MSCs compared with GFP-MSCs. PDGFB-MSCs exhibited enhanced survival and expansion after transplantation, resulting in an enlarged humanized niche cell pool that provide a better humanized microenvironment to facilitate superior engraftment and proliferation of human hematopoietic cells. Our studies demonstrate the efficacy of PDGFB-MSCs in supporting human HSC engraftment.

Reactive Oxygen Species and Nrf2: Functional and Transcriptional Regulators of Hematopoiesis.

Hematopoietic stem cells (HSCs) are characterized by self-renewal and multilineage differentiation potentials. Although they play a central role in hematopoietic homeostasis and bone marrow (BM) transplantation, they are affected by multiple environmental factors in the BM. Here, we review the effects of reactive oxygen species (ROS) and Nrf2 on HSC function and BM transplantation. HSCs reside in the hypoxic microenvironment of BM, and ROS play an important role in HSPC regulation. Recently, an extraphysiologic oxygen shock/stress phenomenon was identified in human cord blood HSCs collected under ambient air conditions. Moreover, Nrf2 has been recently recognized as a master transcriptional factor that regulates multiple antioxidant enzymes. Since several years, the role of Nrf2 in hematopoiesis has been extensively studied, which has functional similarities of cellular oxygen sensor hypoxia-inducible factor-1 as transcriptional factors. Increasing evidence has revealed that abnormally elevated ROS production due to factors such as genetic defects, aging, and ionizing radiation unexceptionally resulted in lethal impairment of HSC function and hematopoiesis. Both experimental and clinical studies have identified elevated ROS levels as a major culprit of ineffective BM transplantation. Lastly, we discuss the possibility of using small molecule antioxidants, such as N-acetyl cysteine, resveratrol, and curcumin, to augment HSC function and improve the therapeutic efficacy of BM transplantation. Further research on the function of ROS levels and improving the efficacy of BM transplantation may have a great potential for broad clinical applications of HSCs.

Effect of Light-Emitting Diodes and Ultraviolet Irradiation on the Soluble Sugar, Organic Acid, and Carotenoid Content of Postharvest Sweet Oranges (Citrus sinensis (L.) Osbeck).

Mature 'Hamlin' sweet oranges (Citrus sinensis (L.) Osbeck) were irradiated using light-emitting diodes (LEDs) and ultraviolet (UV) light for six days after harvest. Based on evaluation of the basic ripening parameters of fruits, the contents of soluble sugars, organic acids, and carotenoids were analyzed (in pulps) on the sixth day by high-performance liquid chromatography (HPLC). The results showed that LED and UV irradiation not only accelerated orange ripening but also caused significant changes in the soluble sugar, organic acid, and carotenoid content. Compared with fruit subjected to dark shade (DS) treatment, the total soluble sugar, fructose, and glucose contents increased significantly in UV-treated (UVA, UVB, and UVC) fruits, while the sucrose content increased remarkably in white light, UVB, and UVC-treated fruits (p < 0.05). UV treatment was associated with inducing the largest effect on the total soluble sugar content. Except for UVB, other types of light notably induced an accumulation of the total organic acid content, none but blue light and red light markedly induced citric acid accumulation (p < 0.05). Interestingly, only the red light and dark shade treatments had markedly positive effects in terms of inducing carotenoid accumulation, including the total carotenoid, isolutein, zeaxanthin, lutein, neoxanthin, all-trans-violaxanthin, phytofluene, cis-ζ-carotene, and ß-carotene concentrations. Other light treatments had significantly negative effects on carotenoid accumulation (p < 0.05). Therefore, soluble sugar, organic acid, and carotenoid accumulation in sweet oranges vary depending on the levels of UV and LED irradiation. Appropriate light irradiation is a potentially effective way to maintain or improve postharvest fruit quality.

Effect of Post-Harvest LED and UV Light Irradiation on the Accumulation of Flavonoids and Limonoids in the Segments of Newhall Navel Oranges (Citrus sinensis Osbeck).

To investigate the effect of post-harvest light irradiation on the accumulation of flavonoids and limonoids, harvested Newhall navel oranges were continuously exposed to light-emitting diode (LED) and ultraviolet (UV) light irradiation for 6 days, and the composition and content of flavonoids and limonoids in the segments were determined using UPLC-qTOF-MS at 0, 6, and 15 days after harvest. In total, six polymethoxylated flavonoids (PMFs), five flavone-O/C-glycosides, seven flavanone-O-glycosides, and three limonoids were identified in the segments. The accumulation of these components was altered by light irradiation. Red and blue light resulted in higher levels of PMFs during exposure periods. The accumulation of PMFs was also significantly induced after white light, UVB and UVC irradiation were removed. Red and UVC irradiation induced the accumulation of flavone and flavanone glycosides throughout the entire experimental period. Single light induced limonoid accumulation during exposure periods, but limonoid levels decreased significantly when irradiation was removed. Principal component analysis showed a clear correlation between PMFs and white light, between flavonoid glycosides and red light and UVC, and between limonoids and UVC. These results suggest that the accumulation of flavonoids and limonoids in citrus is regulated by light irradiation. White light, red light and UVC irradiation might be a good potential method for improving the nutrition and flavor quality of post-harvest citrus.

Theoretical study on the reaction mechanism of OH radical with Z(E)-CF3CH[double bond, length as m-dash]CHF.

Understanding the atmospheric fate of hydrofluoroolefins (HFOs) is of great significance to assess their potential risk to the environment. As an important type of HFO, the comprehensive transformation mechanism and kinetics of Z(E)-CF3CH[double bond, length as m-dash]CHF initiated by OH radicals were investigated by performing quantum chemical calculations at the CCSD(T)/aug-cc-pVTZ//MP2/cc-pVDZ level. The results show that the OH-addition pathways are the most favorable for the title reaction. The rate constants are obtained by transition state theory with Wigner tunneling correction (TST/W). The calculated total rate constants are in good agreement with the experimental data. At 298 K, the computed rate constant and lifetime of Z(E)-CF3CH[double bond, length as m-dash]CHF are 9.66 × 10-13 (4.02 × 10-13) cm3 molecule-1 s-1 and 12.3 (29.7) days, respectively, which demonstrates that Z(E)-CF3CH[double bond, length as m-dash]CHF is atmospherically persistent.

Megakaryocyte-derived excessive transforming growth factor ß1 inhibits proliferation of normal hematopoietic stem cells in acute myeloid leukemia.

Impaired production of healthy hematopoietic cells from residual hematopoietic stem cells (HSCs) leads to high mortality in acute myeloid leukemia (AML). Previous studies have identified p21 and Egr3 as intrinsic factors responsible for the growth arrest and differentiation blockade of normal HSCs in leukemia; however, the related extrinsic factors remain unknown. In this study, we found that transforming growth factor ß (TGFß) signaling was upregulated in HSCs from bone marrow of mice with MLL-AF9-induced acute myeloid leukemia (AML) because of excessive production of TGFß1, especially from megakaryocytes, and overactivation of latent TGFß1 protein. We also found that SMAD3, a signal transducer of TGFß1, directly bound to Egr3 and upregulated its expression to arrest proliferation of HSCs. Our study provides evidence for targeting TGFß1 in AML to rectify normal hematopoiesis defects in clinical practice.

Clinical application of MRI-respiratory gating technology in the evaluation of children with obstructive sleep apnea hypopnea syndrome.

The objective of the present study was to investigate the clinical application of magnetic resonance imaging (MRI)-respiratory gating technology for assessing illness severity in children with obstructive sleep apnea hypopnea syndrome (OSAHS).MRI-respiratory gating technology was used to scan the nasopharyngeal cavities of 51 children diagnosed with OSAHS during 6 respiratory phases. Correlations between the ratio of the area of the adenoid to the area of the nasopalatine pharyngeal cavity (Sa/Snp), with the main indexes of polysomnography (PSG), were analyzed. Receiver operator characteristic (ROC) curve and Kappa analysis were used to determine the diagnostic accuracy of Sa/Snp in pediatric OSAHS.The Sa/Snp was positively correlated with the apnea hypopnea index (AHI) (P < .001) and negatively correlated with the lowest oxygen saturation of blood during sleep (LaSO2) (P < .001). ROC analysis in the 6 respiratory phases showed that the area under the curve (AUC) of the Sa/Snp in the end-expiratory phase was the largest (0.992, P < .001), providing a threshold of 69.5% for the diagnosis of severe versus slight-moderate OSAHS in children. Consistency analysis with the AHI showed a diagnosis accordance rate of 96.0% in severe pediatric OSAHS and 96.2% in slight-moderate pediatric OSAHS (Kappa = 0.922, P < .001).Stenosis of the nasopalatine pharyngeal cavity in children with adenoidal hypertrophy was greatest at the end-expiration phase during sleep. The end-expiratory Sa/Snp obtained by a combination of MRI and respiratory gating technology has potential as an important imaging index for diagnosing and evaluating severity in pediatric OSAHS.

A molecular pathology method for sequential fluorescence in situ hybridization for multi-gene analysis at the single-cell level.

Multi-gene detection at the single-cell level is desirable to enable more precise genotyping of heterogeneous hematology and oncology samples. This study aimed to establish a single-cell multi-gene fluorescence in situ hybridization (FISH) method for use in molecular pathology analyses. Five fluorochromes were used to label different FISH gene probes, and 5 genes were detected using a five-color FISH protocol. After the first hybridization, the previous FISH probe set was stripped, and a second set of five-color FISH probes was used for rehybridization. After each hybridization, the fluorescence signals were recorded in 6 fluorescence filter channels that included DAPI, Spectrum Green™, Cy3™ v1, Texas Red, Cy5, and PF-415. A digital automatic relocation procedure was used to ensure that exactly the same microscopic field was studied in each stripping and hybridization cycle. By using this sequential stripping and rehybridization strategy, up to 20 genes can be detected within a single nucleus. In conclusion, a practical molecular pathology method was developed for analyzing multiple genes at the single-cell level.