Intrahepatic cholangiocarcinoma (ICC) is the second most common liver malignancy. ICC typically features remarkable cellular heterogeneity and a dense stromal reaction. Therefore, a comprehensive ...understanding of cellular diversity and the interplay between malignant cells and niche cells is essential to elucidate the mechanisms driving ICC progression and to develop therapeutic approaches.
Herein, we performed single-cell RNA sequencing (scRNA-seq) analysis on unselected viable cells from 8 human ICCs and adjacent samples to elucidate the comprehensive transcriptomic landscape and intercellular communication network. Additionally, we applied a negative selection strategy to enrich fibroblast populations in 2 other ICC samples to investigate fibroblast diversity. The results of the analyses were validated using multiplex immunofluorescence staining, bulk transcriptomic datasets, and functional in vitro and in vivo experiments.
We sequenced a total of 56,871 single cells derived from human ICC and adjacent tissues and identified diverse tumor, immune, and stromal cells. Malignant cells displayed a high degree of inter-tumor heterogeneity. Moreover, tumor-infiltrating CD4 regulatory T cells exhibited highly immunosuppressive characteristics. We identified 6 distinct fibroblast subsets, of which the majority were CD146-positive vascular cancer-associated fibroblasts (vCAFs), with highly expressed microvasculature signatures and high levels of interleukin (IL)-6. Functional assays indicated that IL-6 secreted by vCAFs induced significant epigenetic alterations in ICC cells, particularly upregulating enhancer of zeste homolog 2 (EZH2) and thereby enhancing malignancy. Furthermore, ICC cell-derived exosomal miR-9-5p elicited high expression of IL-6 in vCAFs to promote tumor progression.
Our single-cell transcriptomic dataset delineates the inter-tumor heterogeneity of human ICCs, underlining the importance of intercellular crosstalk between ICC cells and vCAFs, and revealing potential therapeutic targets.
Intrahepatic cholangiocarcinoma is an aggressive and chemoresistant malignancy. Better understanding the complex transcriptional architecture and intercellular crosstalk of these tumors will help in the development of more effective therapies. Herein, we have identified important interactions between cancer cells and cancer-associated fibroblasts in the tumor stroma, which could have therapeutic implications.
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•Single cell transcriptomic datasets are a valuable resource to dissect cellular diversity and intercellular crosstalk of human ICCs.•Malignant cells displayed remarkable inter-tumor heterogeneity and Tregs revealed highly immunosuppressive characteristics.•Six distinct fibroblast subsets were defined in ICCs and adjacent tissues.•CD146+ vCAFs, comprising most of the fibroblasts, had tight interactions with malignant cells through IL-6/IL-6R axis.•Tumor exosomal miR-9-5p elicited IL-6 expression in vCAFs, contributing to ICC progression via upregulation of EZH2.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Abstract We report here the loading of the antitumor drug doxorubicin (DOX) in preformed multilayer microcapsules and its application in tumor treatment assayed by in vitro cell culture and in vivo ...animal experiments. The microcapsules, consisting completely of polysaccharides, were fabricated by deposition of oppositely charged chitosan and alginate onto carboxylmethyl cellulose (CMC)–doped CaCO3 colloidal particles in a layer-by-layer fashion, followed by cross-linking with glutaraldehyde and decomposition of the cores by disodium ethylenediaminetetraacetic acid. The microcapsules as prepared contain negatively charged CMC—either in a free state or very possibly coupled with the excess chitosan of the first layer. They showed a strong ability to accumulate the positively charged DOX with a factor of tens to hundreds; that is, the drug concentration within the microcapsules was hundreds of times higher than the feeding concentration. Confocal microscopy and transmission electron microscopy revealed homogeneous distribution of the drug. The encapsulated DOX could be released again, following a diffusion-controlled model at the initial stage. In vitro experiments showed that the encapsulated drug can effectively induce the apoptosis of HepG2 tumor cells, as shown by various microscopy techniques after acridine orange, Hoechst 33342, and osmium tetraoxide staining. By seeding the HepG2 hepatoma cells into BALB/c/nu mice, tumors were created for the experimental studies. The results showed that the encapsulated DOX had better efficacy than that of the free drug in terms of tumor inhibition in a 4-week in vivo culture period.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
Following the “green revolution,”
indica
and
japonica
hybrid breeding has been recognized as a new breakthrough in further improving rice yields. However, heterosis-related grain weight QTLs and the ...basis of yield advantage among subspecies has not been well elucidated. We herein
de novo
assembled the chromosome level genomes of an
indica/xian
rice (Luohui 9) and a
japonica/geng
rice (RPY geng) and found that gene number differences and structural variations between these two genomes contribute to the differences in agronomic traits and also provide two different favorable allele pools to produce better derived recombinant inbred lines (RILs). In addition, we generated a high-generation (> F
15
) population of 272 RILs from the cross between Luohui 9 and RPY geng and two testcross hybrid populations derived from the crosses of RILs and two cytoplasmic male sterile lines (YTA,
indica
and Z7A,
japonica
). Based on three derived populations, we totally identified eight 1,000-grain weight (KGW) QTLs and eight KGW heterosis loci. Of QTLs,
qKGW-6.1
and
qKGW-8.1
were accepted as novel KGW QTLs that have not been reported previously. Interestingly, allele genotyping results revealed that heading date related gene (
Ghd8
) in
qKGW-8.1
and
qLH-KGW-8.1
, can affect grain weight in RILs and rice core accessions and may also play an important role in grain weight heterosis. Our results provided two high-quality genomes and novel gene editing targets for grain weight for future rice yield improvement project.
Chlorophyll is the main photosynthetic pigment and is crucial for plant photosynthesis. Leaf color mutants are widely used to identify genes involved in the synthesis or metabolism of chlorophyll. In ...this study, a spontaneous mutant,
(
), was isolated from rice (
). This
mutant showed yellow-green leaves and decreased chlorophyll level and net photosynthetic rate. Brown necrotic spots appeared on the surface of
leaves at the tillering stage. And the agronomic traits of the
mutant, including the plant height, tiller number per plant, and total number of grains per plant, were significantly reduced. Map-based cloning revealed that the candidate
gene was LOC_Os03g21370. Complementation of the
mutant with the wild-type CDS of LOC_Os03g21370 led to the restoration of the mutant to the normal phenotype. Evolutionary analysis revealed that YGL19 protein and its homologues were unique for photoautotrophs, containing a conserved Ycf54 functional domain. A conserved amino acid substitution from proline to serine on the Ycf54 domain led to the ygl19 mutation. Sequence analysis of the
gene in 4726 rice accessions found that the
gene was conserved in natural rice variants with no resulting amino acid variation. The
gene was mainly expressed in green tissues, especially in leaf organs. And the YGL19 protein was localized in the chloroplast for function. Gene expression analysis via qRT-PCR showed that the expression levels of tetrapyrrole synthesis-related genes and photosynthesis-related genes were regulated in the
mutant. Reactive oxygen species (ROS) such as superoxide anions and hydrogen peroxide accumulated in spotted leaves of the
mutant at the tillering stage, accompanied by the regulation of ROS scavenging enzyme-encoding genes and ROS-responsive defense signaling genes. This study demonstrates that a novel yellow-green leaf gene
affects tetrapyrrole biosynthesis, photosynthesis, and ROS metabolism in rice.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Liver disease is an important disease that seriously threatens human health. It accounts for the highest proportion in various malignant tumors, and its incidence rate and mortality are on the rise, ...seriously affecting human health. Modern imaging has developed rapidly, but the application of image segmentation in liver tumor surgery is still rare. The application of image processing technology represented by artificial intelligence (AI) in surgery can greatly improve the efficiency of surgery, reduce surgical complications, and reduce the cost of surgery. Hepatocellular carcinoma is the most common malignant tumor in the world, and its mortality is second only to lung cancer. The resection rate of liver cancer surgery is high, and it is a multidisciplinary surgery, so it is necessary to explore the possibility of effective switching between different disciplines. Resection of hepatobiliary and pancreatic tumors is one of the most challenging and lethal surgical procedures. The operation requires a high level of doctors’ experience and understanding of anatomical structures. The surgical segmentation is slow and there may be obvious complications. Therefore, the surgical system needs to make full use of the relevant functions of AI technology and computer vision analysis software, and combine the processing strategy based on image processing algorithm and computer vision analysis model. Intelligent optimization algorithm, also known as modern heuristic algorithm, is an algorithm with global optimization performance, strong universality, and suitable for parallel processing. This algorithm generally has a strict theoretical basis, rather than relying solely on expert experience. In theory, the optimal solution or approximate optimal solution can be found in a certain time. This work studies the hepatobiliary surgery through intelligent image segmentation technology, and analyzes them through intelligent optimization algorithm. The research results showed that when other conditions were the same, there were three patients who had adverse reactions in hepatobiliary surgery through intelligent image segmentation technology, accounting for 10%. The number of patients with adverse reactions in hepatobiliary surgery by conventional methods was nine, accounting for 30%, which was significantly higher than the former, indicating a positive relationship between intelligent image segmentation technology and hepatobiliary surgery.
Grain quality is an important breeding objective in rice, and the appearance of the grain also affects its commercial value in the market. The aim of this study was to decode the rice grain qualities ...and appearances, such as gelatinization temperature (GT), amylose content (AC), grain protein content (GPC), pericarp color (PC), length/width ratio (LWR), and grain volume (GV) at phenotypic and genetic levels, as well as the relationships among them. A genome-wide association study (GWAS) was used to identify the quantitative trait locus (QTLs) associated with the target traits using mixed linear model (MLM) and Bayesian-information and linkage-disequilibrium iteratively nested keyway (BLINK) methods. In general, AC was negatively correlated with GPC and GV, while it was positively correlated with LWR and PC. GPC was positively correlated with LWR. Using the rice diversity panel 1 (RDP1) population, we identified 11, 6, 2, 7, 11, and 6 QTLs associated with GT, AC, GPC, PC, LWR, and GV, respectively. Five germplasm lines, superior in grain qualities and appearances for basic breeding materials or improvement, were identified. Notably, an F-box gene
OsFbox394
was located in the linkage disequilibrium (LD) block of
qLWR7-2
, which specifically expresses in endosperm and seed tissues, suggesting that this gene may regulate the seed development in rice grain. Besides, different haplotypes of
OsHyPRP45
showed significant differences in AC, indicating that this gene may be related to AC in rice grain.
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused the pandemic of coronavirus disease 2019 (COVID-19). Great international efforts have been put into the development of ...prophylactic vaccines and neutralizing antibodies. However, the knowledge about the B cell immune response induced by the SARS-CoV-2 virus is still limited. Here, we report a comprehensive characterization of the dynamics of immunoglobin heavy chain (IGH) repertoire in COVID-19 patients. By using next-generation sequencing technology, we examined the temporal changes in the landscape of the patient’s immunological status and found dramatic changes in the IGH within the patient’s immune system after the onset of COVID-19 symptoms. Although different patients have distinct immune responses to SARS-CoV-2 infection, by employing clonotype overlap, lineage expansion, and clonotype network analyses, we observed a higher clonotype overlap and substantial lineage expansion of B cell clones 2–3 weeks after the onset of illness, which is of great importance to B-cell immune responses. Meanwhile, for preferences of V gene usage during SARS-CoV-2 infection, IGHV3-74 and IGHV4-34, and IGHV4-39 in COVID-19 patients were more abundant than those of healthy controls. Overall, we present an immunological resource for SARS-CoV-2 that could promote both therapeutic development as well as mechanistic research.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Oncolytic virotherapy is a promising strategy for the treatment of cancer. Influenza A virus has shown potential as an oncolytic agent. In this study, a recombinant PR8 influenza viral vector, called ...delNS1-GM-CSF, was generated with a partial deletion in NS and the granulocyte-macrophage colony-stimulating factor (GM-CSF) coding sequence inserted into the influenza nonstructural protein 1 gene. The morphological characteristics of delNS1-GM-CSF were examined. The delNS1-GM-CSF virus replicated well in various cell lines, including MDCK, A549, SMCC7721, and HepG2 cells. Moreover, selective cytotoxicity of the virus was observed in various hepatocellular carcinoma (HCC) cell lines, while no effect was demonstrated in the normal liver cell line LO2, as indicated by 3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyltetrazolium bromide and crystal violet assays. Importantly, using a model based on the growth of HepG2 cells as a xenograft in nude mice, it was found that a reassortant delNS1-GM-CSF virus inhibited tumor growth significantly following intratumoral injection in a dose-dependent manner. Ex vivo results showed that the tumor inhibition efficacy of delNS1-GM-CSF was observed in HCC clinical samples. Taken together, these results are the first to demonstrate that influenza A viruses may have potential as oncolytic virotherapeutic agents against HCC.
Magnesium protoporphyrin IX methyltransferase (ChlM) catalyzes the formation of magnesium protoporphyrin IX monomethylester (MgPME) from magnesium protoporphyrin IX (MgP) in the chlorophyll synthesis ...pathway. However, no
gene has yet been identified and studied in monocotyledonous plants. In this study, a spontaneous mutant,
(
), was isolated from rice (
). This mutant showed yellow-green leaves, decreased chlorophyll level, and climate-dependent growth differences. Map-based cloning of this mutant identified the
gene LOC_Os06g04150.
is expressed in green tissues, especially in leaf organs, where it functions in chloroplasts. YGL18 showed an amino-acid sequence similarity to that of ChlM from different photosynthetic organisms.
enzymatic assays demonstrated that YGL18 performed ChlM enzymatic activity, but ygl18 had nearly lost all ChlM activity. Correspondingly, the substrate MgP was largely accumulated while the product MgPME was reduced in
leaves.
is required for light-dependent and photoperiod-regulated chlorophyll synthesis. The retarded growth of
mutant plants was caused by the high light intensity. Moreover, the higher light intensity and longer exposure in high light intensity even made the
plants be more susceptible to death. Based on these results, it is suggested that
plays essential roles in light-related chlorophyll synthesis and light intensity-involved plant growth.
UDP glucose pyrophosphorylase (UDPGP) family genes have been reported to play essential roles in cell death or individual survival. However, a systematic analysis on UDPGP gene family has not been ...performed yet. In this study, a total of 454 UDPGP proteins from 76 different species were analyzed. The analyses of the phylogenetic tree and orthogroups divided UDPGPs into three clades, including UDP-
-acetylglucosamine pyrophosphorylase (UAP), UDP-glucose pyrophosphorylase (UGP, containing UGP-A and UGP-B), and UDP-sugar pyrophosphorylase (USP). The evolutionary history of the UDPGPs indicated that the members of UAP, USP, and UGP-B were relatively conserved while varied in UGP-A. Homologous sequences of UGP-B and USP were found only in plants. The expression profile of UDPGP genes in
was mainly motivated under jasmonic acid (JA), abscisic acid (ABA), cadmium, and cold treatments, indicating that UDPGPs may play an important role in plant development and environment endurance. The key amino acids regulating the activity of UDPGPs were analyzed, and almost all of them were located in the NB-loop, SB-loop, or conserved motifs. Analysis of the natural variants of UDPGPs in rice revealed that only a few missense mutants existed in coding sequences (CDSs), and most of the resulting variations were located in the non-motif sites, indicating the conserved structure and function of UDPGPs in the evolution. Furthermore, alternative splicing may play a key role in regulating the activity of UDPGPs. The spatial structure prediction, enzymatic analysis, and transgenic verification of UAP isoforms illustrated that the loss of N- and C-terminal sequences did not affect the overall 3D structures, but the N- and C-terminal sequences are important for UAP genes to maintain their enzymatic activity. These results revealed a conserved UDPGP gene family and provided valuable information for further deep functional investigation of the UDPGP gene family in plants.
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