scPLAN: a hierarchical computational framework for single transcriptomics data annotation, integration and cell-type label refinement.

MOTIVATION: In the past decade, single-cell RNA sequencing (scRNA-seq) has emerged as a pivotal method for transcriptomic profiling in biomedical research. Precise cell-type identification is crucial for subsequent analysis of single-cell data. And the integration and refinement of annotated data are essential for building comprehensive databases. However, prevailing annotation techniques often overlook the hierarchical organization of cell types, resulting in inconsistent annotations. Meanwhile, most existing integration approaches fail to integrate datasets with different annotation depths and none of them can enhance the labels of outdated data with lower annotation resolutions using more intricately annotated datasets or novel biological findings. RESULTS: Here, we introduce scPLAN, a hierarchical computational framework designed for scRNA-seq data analysis. scPLAN excels in annotating unlabeled scRNA-seq data using a reference dataset structured along a hierarchical cell-type tree. It identifies potential novel cell types in a systematic, layer-by-layer manner. Additionally, scPLAN effectively integrates annotated scRNA-seq datasets with varying levels of annotation depth, ensuring consistent refinement of cell-type labels across datasets with lower resolutions. Through extensive annotation and novel cell detection experiments, scPLAN has demonstrated its efficacy. Two case studies have been conducted to showcase how scPLAN integrates datasets with diverse cell-type label resolutions and refine their cell-type labels. AVAILABILITY: https://github.com/michaelGuo1204/scPLAN.

Prospective Evaluation of the Peripheral Perfusion Index in Assessing the Organ Dysfunction and Prognosis of Adult Patients With Sepsis in the ICU.

Background: The peripheral perfusion index (PI) reflects microcirculatory blood flow perfusion and indicates the severity and prognosis of sepsis. Method: The cohort comprised 208 patients admitted to the intensive care unit (ICU) with infection, among which 117 had sepsis. Demographics, medication history, ICU variables, and laboratory indexes were collected. Primary endpoints were in-hospital mortality and 28-day mortality. Secondary endpoints included organ function variables (coagulation function, liver function, renal function, and myocardial injury), lactate concentration, mechanical ventilation time, and length of ICU stay. Univariate and multivariate analyses were conducted to assess the associations between the PI and clinical outcomes. Sensitivity analyses were performed to explore the associations between the PI and organ functions in the sepsis and nonsepsis groups. Result: The PI was negatively associated with in-hospital mortality (odds ratio [OR] 0.29, 95% confidence interval [CI] 0.15 to 0.55), but was not associated with 28-day mortality. The PI was negatively associated with the coagulation markers prothrombin time (PT) (ß -0.36, 95% CI -0.59 to 0.13) and activated partial thromboplastin time (APTT) (ß -1.08, 95% CI -1.86 to 0.31), and the myocardial injury marker cardiac troponin I (cTnI) (ß -2085.48, 95% CI -3892.35 to 278.61) in univariate analysis, and with the PT (ß -0.36, 95% CI -0.60 to 0.13) in multivariate analysis. The PI was negatively associated with the lactate concentration (ß -0.57, 95% CI -0.95 to 0.19), mechanical ventilation time (ß -23.11, 95% CI -36.54 to 9.69), and length of ICU stay (ß -1.28, 95% CI -2.01 to 0.55). Sensitivity analyses showed that the PI was significantly associated with coagulation markers (PT and APTT) and a myocardial injury marker (cTnI) in patients with sepsis, suggesting that the associations between the PI and organ function were stronger in the sepsis group than the nonsepsis group. Conclusion: The PI provides new insights for assessing the disease severity, short-term prognosis, and organ function damage in ICU patients with sepsis, laying a theoretical foundation for future research.

Remnant cholesterol is associated with unstable carotid plaque in a neurologically healthy population.

BACKGROUND: Remnant cholesterol (RC) is considered to be one of the most significant and important risk factors for atherosclerotic cardiovascular disease (ASCVD). Nonetheless, the association between RC and unstable carotid plaque remains unclear. Our primary objective is to ascertain whether RC exhibits an independent and significant association with unstable carotid plaque in a neurologically healthy population. METHODS: In the cross-sectional study, we enrolled neurologically healthy participants who visited our centre for health checkups between 2021 and 2022. All eligible participants underwent a standardised questionnaire, physical examinations and laboratory testing. The carotid plaque was evaluated with a standard carotid ultrasound and an advanced ultrasound imaging technique called superb microvascular imaging. The correlation between lipids and unstable carotid plaque was primarily assessed utilising univariate and multivariate logistic regression. RESULTS: The study totally enrolled 1100 participants who had an average age of 57.00 years (IQR: 49.00-63.00), with 67.55% being men. Among the participants, 321 (29.18%) had unstable carotid plaque. In the multivariate logistic regression analysis, higher RC had an independent association with an elevated incidence of unstable carotid plaque compared with the lowest concentrations of RC (OR=1.673, 95% CI 1.113 to 2.515, p=0.0134), but not other lipids. In addition, apolipoprotein A1 was negatively related to unstable carotid plaque (OR=0.549, 95% CI 0.364 to 0.830, p=0.0045). CONCLUSIONS: Elevated concentrations of RC are independently and excellently correlated with unstable carotid plaque within a neurologically healthy population.

SPANN: annotating single-cell resolution spatial transcriptome data with scRNA-seq data.

MOTIVATION: The rapid development of spatial transcriptome technologies has enabled researchers to acquire single-cell-level spatial data at an affordable price. However, computational analysis tools, such as annotation tools, tailored for these data are still lacking. Recently, many computational frameworks have emerged to integrate single-cell RNA sequencing (scRNA-seq) and spatial transcriptomics datasets. While some frameworks can utilize well-annotated scRNA-seq data to annotate spatial expression patterns, they overlook critical aspects. First, existing tools do not explicitly consider cell type mapping when aligning the two modalities. Second, current frameworks lack the capability to detect novel cells, which remains a key interest for biologists. RESULTS: To address these problems, we propose an annotation method for spatial transcriptome data called SPANN. The main tasks of SPANN are to transfer cell-type labels from well-annotated scRNA-seq data to newly generated single-cell resolution spatial transcriptome data and discover novel cells from spatial data. The major innovations of SPANN come from two aspects: SPANN automatically detects novel cells from unseen cell types while maintaining high annotation accuracy over known cell types. SPANN finds a mapping between spatial transcriptome samples and RNA data prototypes and thus conducts cell-type-level alignment. Comprehensive experiments using datasets from various spatial platforms demonstrate SPANN's capabilities in annotating known cell types and discovering novel cell states within complex tissue contexts. AVAILABILITY: The source code of SPANN can be accessed at https://github.com/ddb-qiwang/SPANN-torch. CONTACT: dengmh@math.pku.edu.cn.

The collateral activity of RfxCas13d can induce lethality in a RfxCas13d knock-in mouse model.

BACKGROUND: The CRISPR-Cas13 system is an RNA-guided RNA-targeting system and has been widely used in transcriptome engineering with potentially important clinical applications. However, it is still controversial whether Cas13 exhibits collateral activity in mammalian cells. RESULTS: Here, we find that knocking down gene expression using RfxCas13d in the adult brain neurons caused death of mice, which may result from the collateral activity of RfxCas13d rather than the loss of target gene function or off-target effects. Mechanistically, we show that RfxCas13d exhibits collateral activity in mammalian cells, which is positively correlated with the abundance of target RNA. The collateral activity of RfxCas13d could cleave 28s rRNA into two fragments, leading to translation attenuation and activation of the ZAKα-JNK/p38-immediate early gene pathway. CONCLUSIONS: These findings provide new mechanistic insights into the collateral activity of RfxCas13d in mammalian cells and warn that the biosafety of the CRISPR-Cas13 system needs further evaluation before application to clinical treatments.

A pH-neutral bioactive glass coated 3D-printed porous Ti6Al4V scaffold with enhanced osseointegration.

Osseointegration is vital for the success of non-degradable implants like those made of titanium alloys. In order to promote osseointegration, implants are made porous, providing space for bone ingrowth. Despite extensive optimization of the pore geometry and porosity, bone ingrowth into implants is still marginal; further modification to promote bone ingrowth as well as osseointegration becomes paramount. In this study, a pH neutral bioactive glass with the composition of 10.8% P2O5-54.2% SiO2-35% CaO (mol%; hereinafter referred to as PSC) was successfully coated on 3D-printed porous Ti6Al4V scaffolds using an in situ sol-gel method. This PSC coating is strongly bonded to the substrate and quickly induces the formation of hydroxyapatite on the scaffold surface upon contact with body fluid. In vitro, the PSC-coated Ti6Al4V scaffolds showed superior biocompatibility, cell proliferation promotion, cell adhesion, osteogenic differentiation and mineralization compared to their bare counterparts, implying better osseointegration. In vivo experiments confirmed this expectation; after being implanted, the coated scaffolds had more bone ingrowth and osseointegration, and consequently, higher push-out strength was achieved, proving the validity of the proposed concept in this study. In conclusion, PSC coating on 3D-printed porous Ti6Al4V scaffolds can improve osteogenesis, bone ingrowth, and osseointegration. Together with the versatility of this in situ sol-gel coating method, titanium alloy implants with better biological performances may be developed for immediate clinical applications.

Performance Enhancement of All-Inorganic Carbon-Based CsPbIBr2 Perovskite Solar Cells Using a Moth-Eye Anti-Reflector.

All-inorganic carbon-based CsPbIBr2 perovskite solar cells (PSCs) have attracted increasing interest due to the low cost and the balance between bandgap and stability. However, the relatively narrow light absorption range (300 to 600 nm) limited the further improvement of short-circuit current density (JSC) and power conversion efficiency (PCE) of PSCs. Considering the inevitable reflectance loss (~10%) at air/glass interface, we prepared the moth-eye anti-reflector by ultraviolet nanoimprint technology and achieved an average reflectance as low as 5.15%. By attaching the anti-reflector on the glass side of PSCs, the JSC was promoted by 9.4% from 10.89 mA/cm2 to 11.91 mA/cm2, which is the highest among PSCs with a structure of glass/FTO/c-TiO2/CsPbIBr2/Carbon, and the PCE was enhanced by 9.9% from 9.17% to 10.08%. The results demonstrated that the larger JSC induced by the optical reflectance modulation of moth-eye anti-reflector was responsible for the improved PCE. Simultaneously, this moth-eye anti-reflector can withstand a high temperature up to 200 °C, and perform efficiently at a wide range of incident angles from 40° to 90° and under various light intensities. This work is helpful to further improve the performance of CsPbIBr2 PSCs by optical modulation and boost the possible application of wide-range-wavelength anti-reflector in single and multi-junction solar cells.

Constructing Hierarchically Porous N-Doped Carbons Derived from Poly(ionic liquids) with the Multifunctional Fe-Based Template for CO2 Adsorption.

Nitrogen-doped hierarchical porous carbons with a rich pore structure were prepared via direct carbonization of the poly(ionic liquid) (PIL)/potassium ferricyanide compound. Thereinto, the bisvinylimidazolium-based PIL was a desirable carbon source, and potassium ferricyanide as a multifunctional Fe-based template, could not only serve as the pore-forming agent, including metallic components (Fe and Fe3C), potassium ions (etching carbon framework during carbonization), and gas generated during the pyrolysis process, but also introduce the N atoms to porous carbons, which were in favor of CO2 capture. Moreover, the hierarchically porous carbon NDPC-1-800 (NDPC, nitrogen-doped porous carbon) had taken advantage of the highest specific surface area, exhibiting an excellent CO2 adsorption capacity and selectivity compared with NDC-800 (NDC, nitrogen-doped carbon) directly carbonized from the pure PIL. Furthermore, its hierarchical porous architectures played an important part in the process of CO2 capture, which was described briefly as follows: the synergistic effect of mesopores and micropores could accelerate the CO2 molecules' transportation and storage. Meanwhile, the appropriate microporous size distribution of NDPC-1-800 was conducive to enhancing CO2/N2 selectivity. This study was intended to open up a new pathway for designing N-doped porous carbons combining both PILs and the multifunctional Fe-based template potassium ferricyanide with wonderful gas adsorption and separation performance.

Induction of alarmin S100A8/A9 mediates activation of aberrant neutrophils in the pathogenesis of COVID-19.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic poses an unprecedented public health crisis. Evidence suggests that SARS-CoV-2 infection causes dysregulation of the immune system. However, the unique signature of early immune responses remains elusive. We characterized the transcriptome of rhesus macaques and mice infected with SARS-CoV-2. Alarmin S100A8 was robustly induced in SARS-CoV-2-infected animal models as well as in COVID-19 patients. Paquinimod, a specific inhibitor of S100A8/A9, could rescue the pneumonia with substantial reduction of viral loads in SARS-CoV-2-infected mice. Remarkably, Paquinimod treatment resulted in almost 100% survival in a lethal model of mouse coronavirus infection using the mouse hepatitis virus (MHV). A group of neutrophils that contributes to the uncontrolled pathological damage and onset of COVID-19 was dramatically induced by coronavirus infection. Paquinimod treatment could reduce these neutrophils and regain anti-viral responses, unveiling key roles of S100A8/A9 and aberrant neutrophils in the pathogenesis of COVID-19, highlighting new opportunities for therapeutic intervention.

A Solvent-Exchange Strategy to Regulate Noncovalent Interactions for Strong and Antiswelling Hydrogels.

Physical hydrogels from existing polymers consisting of noncovalent interacting networks are highly desired due to their well-controlled compositions and environmental friendliness; and therefore, applied as adhesives, artificial tissues, and soft machines. Nevertheless, these gels have suffered from weak mechanical strength and low water resistance. Current methodologies used to fabricate these hydrogels mainly involve the freezing-thawing process (cryogels), which are complicated in preparation and short in adjustment of polymer conformation. Here, taking the merits of noncovalent bonds in adjustability and reversibility, a solvent-exchange strategy is developed to construct a class of exogels. Based on the exchange from a good solvent subsequently to a poor one, the intra- and interpolymer interactions are initially suppressed and then recovered, resulting in dissolving and cross-linking to polymers, respectively. Key to this approach is the good solvent, which favors of a stretched polymer conformation to homogenize the network, forming cross-linked hydrogel networks with remarkable stiffness, toughness, antiswelling properties, and thus underwater adhesive performance. The exogels highlight a facile but highly effective strategy of turning the solvent and consequently the noncovalent interactions to achieve the rational design of enhanced hydrogels and hydrogel-based soft materials.

Wrinkled double network hydrogel via simple stretch-recovery.

The wrinkled structures in biological tissues play a key role in nutrition transportation and organ protection and cannot be easily achieved in synthetic hydrogels using universal and convenient strategies. Wrinkles are highly desirable for the nascent applications of hydrogels in biomaterials and artificial organs. Here, we propose a strategy for inducing different viscoelastic behaviours inside a double network hydrogel to achieve regular wrinkles that are formed by the mass redistribution. The wrinkles can be designed on multiple dimensions and can be well reserved in repeated tensile loadings. These wrinkled hydrogels exhibit unusual characteristics, such as the anisotropy of mechanics and J type tensile curves. This strategy is particularly valuable for biomaterials and artificial organs and may become a universal method for designing the surface morphology of soft materials in large-scale preparation.

LSD1 contributes to programmed oocyte death by regulating the transcription of autophagy adaptor SQSTM1/p62.

In female mammals, the size of the initially established primordial follicle (PF) pool within the ovaries determines the reproductive lifespan of females. Interestingly, the establishment of the PF pool is accompanied by a remarkable programmed oocyte loss for unclear reasons. Although apoptosis and autophagy are involved in the process of oocyte loss, the underlying mechanisms require substantial study. Here, we identify a new role of lysine-specific demethylase 1 (LSD1) in controlling the fate of oocytes in perinatal mice through regulating the level of autophagy. Our results show that the relatively higher level of LSD1 in fetal ovaries sharply reduces from 18.5 postcoitus (dpc). Meanwhile, the level of autophagy increases while oocytes are initiating programmed death. Specific disruption of LSD1 resulted in significantly increased autophagy and obviously decreased oocyte number compared with the control. Conversely, the oocyte number is remarkably increased by the overexpression of Lsd1 in ovaries. We further demonstrated that LSD1 exerts its role by regulating the transcription of p62 and affecting autophagy level through its H3K4me2 demethylase activity. Finally, in physiological conditions, a decrease in LSD1 level leads to an increased level of autophagy in the oocyte when a large number of oocytes are being lost. Collectively, LSD1 may be one of indispensible epigenetic molecules who protects oocytes against preterm death through repressing the autophagy level in a time-specific manner. And epigenetic modulation contributes to programmed oocyte death by regulating autophagy in mice.

SP1 governs primordial folliculogenesis by regulating pregranulosa cell development in mice.

Establishment of the primordial follicle (PF) pool is pivotal for the female reproductive lifespan; however, the mechanism of primordial folliculogenesis is poorly understood. Here, the transcription factor SP1 was shown to be essential for PF formation in mice. Our results showed that SP1 is present in both oocytes and somatic cells during PF formation in the ovary. Knockdown of Sp1 expression, especially in pregranulosa cells, significantly suppressed nest breakdown, oocyte apoptosis, and PF formation, suggesting that SP1 expressed by somatic cells functions in the process of primordial folliculogenesis. We further demonstrated that SP1 governs the recruitment and maintenance of Forkhead box L2-positive (FOXL2+) pregranulosa cells using an Lgr5-EGFP-IRES-CreERT2 (Lgr5-KI) reporter mouse model and a FOXL2+ cell-specific knockdown model. At the molecular level, SP1 functioned mainly through manipulation of NOTCH2 expression by binding directly to the promoter of the Notch2 gene. Finally, consistent with the critical role of granulosa cells in follicle survival in vitro, massive loss of oocytes in Sp1 knockdown ovaries was evidenced before puberty after the ovaries were transplanted under the renal capsules. Conclusively, our results reveal that SP1 controls the establishment of the ovarian reserve by regulating pregranulosa cell development in the mammalian ovary.

HDAC3 maintains oocyte meiosis arrest by repressing amphiregulin expression before the LH surge.

It is known that granulosa cells (GCs) mediate gonadotropin-induced oocyte meiosis resumption by releasing EGF-like factors in mammals, however, the detailed molecular mechanisms remain unclear. Here, we demonstrate that luteinizing hormone (LH) surge-induced histone deacetylase 3 (HDAC3) downregulation in GCs is essential for oocyte maturation. Before the LH surge, HDAC3 is highly expressed in GCs. Transcription factors, such as FOXO1, mediate recruitment of HDAC3 to the amphiregulin (Areg) promoter, which suppresses AREG expression. With the LH surge, decreased HDAC3 in GCs enables histone H3K14 acetylation and binding of the SP1 transcription factor to the Areg promoter to initiate AREG transcription and oocyte maturation. Conditional knockout of Hdac3 in granulosa cells in vivo or inhibition of HDAC3 activity in vitro promotes the maturation of oocytes independent of LH. Taking together, HDAC3 in GCs within ovarian follicles acts as a negative regulator of EGF-like growth factor expression before the LH surge.

Prostaglandin-E2 deficiency during late pregnancy and the associated increase in interleukin-1ß derived from periaortic lymph nodes lead to abortion.

Prostaglandin E2 (PGE2) is a hormone with many physiological functions. During pregnancy, it is generally believed that there is a high level of PGE2 at the final stage of pregnancy, which induces the contraction of uterine smooth muscle and promotes the occurrence of childbirth. However, we find that high PGE2 levels are present throughout late pregnancy in mice, not just during childbirth, and that PGE2 deficiency induced by indomethacin during late pregnancy causes damage to the placental labyrinth and eventually leads to abortion. Interestingly, the damage is closely related to inflammation, which involves the role of inflammatory factors produced by the periaortic lymph nodes (PLNs) near the uterus. Further, through RNA sequencing, we reveal that PLNs produce a large amount of interleukin-1ß (IL-1ß) when exposed to PGE2 deficiency, which causes damage to the placental labyrinth, probably via destroying the extracellular matrix. Finally, events leading to abortion following indomethacin administration are effectively prevented by supplementing PGE2 or by PLN removal. These results suggest that high levels of PGE2 during late pregnancy protect fetuses from inflammatory damage related to IL-1ß. This work suggests a new role of PGE2 during late pregnancy and may provide potential therapeutic strategies for pathological pregnancy.

Oocyte-derived E-cadherin acts as a multiple functional factor maintaining the primordial follicle pool in mice.

In mammals, female fecundity is determined by the size of the primordial follicle (PF) pool, which is established during the perinatal period. As a non-renewable resource, the preservation of dormant PFs is crucial for sustaining female reproduction throughout life. Although studies have revealed that several oocyte-derived functional genes and pathways, such as newborn ovary homeobox (NOBOX) and 3-phosphoinositide-dependent protein kinase-1, participate in maintaining the PF pool, our understanding of the underlying molecular mechanisms is still incomplete. Here, we demonstrate that E-cadherin (E-cad) plays a crucial role in the maintenance of PFs in mice. E-cad is specifically localized to the cytomembrane of oocytes in PFs. Knockdown of E-cad in neonatal ovaries resulted in significant PF loss owing to oocyte apoptosis. In addition, the expression pattern of NOBOX is similar to that of E-cad. Knockdown of E-cad resulted in a decreased NOBOX level, whereas overexpression of Nobox partially rescued the follicle loss induced by silencing E-cad. Furthermore, E-cad governed NOBOX expression by regulating the shuttle protein, ß-catenin, which acts as a transcriptional co-activator. Notably, E-cad, which is a transmembrane protein expressed in the oocytes, was also responsible for maintaining the PF structure by facilitating cell-cell adhesive contacts with surrounding pregranulosa cells. In conclusion, E-cad in oocytes of PFs plays an indispensable role in the maintenance of the PF pool by facilitating follicular structural stability and regulating NOBOX expression. These findings shed light on the physiology of sustaining female reproduction.