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Although various genome projects have provided us enormous static sequence information, understanding of the sophisticated biology continues to require integrating the computational modeling, system analysis, technology development for experiments, and quantitative experiments all together to analyze the biology architecture on various levels, which is just the origin of systems biology subject. This review discusses the object, its characteristics, and research attentions in systems biology, and summarizes the analysis methods, experimental technologies, research developments, and so on in the four key fields of systems biology--systemic structures, dynamics, control methods, and design principles.

Objective To identify potential target genes for isocitrate dehydrogenase (IDH) mutations in cholangiocarcinoma. Methods DNA methylation 450K data and mRNA expression data of the TCGA CHOL project were downloaded. Probe-wise differential methylation analysis and differential expression analysis were performed through limma and DESeq2 in the IDH-mutant vs IDH-WT comparison. Then, Spearman correlation analysis was performed between the normalized expression levels of differentially expressed genes and the methylation levels of corresponding differentially methylated sites. Genes with high methylation, low expression, and strong negative correlation were screened for enrichment analysis to explore their functions. A protein interaction network was constructed and the core modules were found by MCODE. Results The analysis revealed 11 605 differentially methylated sites, of which 10 427 were hypermethylated; and 651 of the 735 differentially expressed genes were down-regulated. Among them, 143 down-regulated genes and 328 hypermethylation sites formed 330 strongly negatively correlated combinations. The above 143 genes are enriched in the regulation of epidermal growth factor receptor signaling pathway, cell extravasation and cell division. By constructing a protein-protein interaction (PPI) network, 10 genes of its core module were identified, which were involved in biological processes such as epithelial cell differentiation and development, and cellular protein localization. Conclusions By integrating the TCGA cholangiocarcinoma transcriptome and DNA methylation data, the potential target genes for IDH mutations are obtained and the core modules of their PPI network are identified, providing new clues for the role of IDH mutations in the development of cholangiocarcinoma.

Disuse osteoporosis is characterized by decreased bone mass caused by abnormal mechanical stimulation of bone. Piezo1 is a major mechanosensitive ion channel in bone homeostasis. However, whether intervening in the action of Piezo1 can rescue disuse osteoporosis remains unresolved. In this study, a commonly‐used hindlimb‐unloading model is employed to simulate microgravity. By single‐cell RNA sequencing, bone marrow‐derived mesenchymal stem cells (BMSCs) are the most downregulated cell cluster, and coincidentally, Piezo1 expression is mostly enriched in those cells, and is substantially downregulated by unloading. Importantly, activation of Piezo1 by systemically‐introducing yoda1 mimics the effects of mechanical stimulation and thus ameliorates bone loss under simulated microgravity. Mechanistically, Piezo1 activation promotes the proliferation and osteogenic differentiation of Gli1+ BMSCs by activating the β‐catenin and its target gene activating transcription factor 4 (ATF4). Inhibiting β‐catenin expression substantially attenuates the effect of yoda1 on bone loss, possibly due to inhibited proliferation and osteogenic differentiation capability of Gli1+ BMSCs mediated by ATF4. Lastly, Piezo1 activation also slightly alleviates the osteoporosis of OVX and aged mice. In conclusion, impaired function of Piezo1 in BMSCs leads to insufficient bone formation especially caused by abnormal mechanical stimuli, and is thus a potential therapeutic target for osteoporosis. [ABSTRACT FROM AUTHOR]

Expansin proteins, a crucial class of intracellular proteins, are known to play a vital role in facilitating processes like cell wall relaxation and cell growth. Recent discoveries have revealed that expansin proteins also have significant functions in plant growth, development, and response to resistance. However, the expansin gene family, particularly in emmer wheat, has not been thoroughly studied, particularly in terms of evolution. In this study, we identified 63 TdEXPs and 49 TtEXPs from the latest genome versions of wild emmer wheat (WEW) and durum wheat (DW), respectively. The physicochemical properties of the encoded expansin proteins exhibited minimal differences, and the gene structures remained relatively conserved. Phylogenetic analysis categorized the proteins into three subfamilies, namely EXPA, EXPB, and EXLA, in addition to the EXLB subfamily. Furthermore, codon preference analysis revealed an increased usage frequency of the nucleotide "T" in expansin proteins throughout the evolution of WEW and DW. Collinearity analysis demonstrated higher orthology between the expansin proteins of WEW and DW, with a Ka/Ks ratio ranging from 0.4173 to 0.9494, indicating purifying selection during the evolution from WEW to DW. Haplotype analysis of the expansin gene family identified five genes in which certain haplotypes gradually became dominant over the course of evolution, enabling adaptation for survival and improvement. Expression pattern analysis indicated tissue-specific expression of expansin genes in emmer wheat, and some of these genes were quantified through qRT-PCR to assess their response to salt stress. These comprehensive findings present the first systematic analysis of the expansin protein gene family during the evolution from WEW to DW, providing a foundation for further understanding the functions and biological roles of expansin protein genes in emmer wheat. [ABSTRACT FROM AUTHOR]

Purpose: Pyroptosis plays an important role in the occurrence and progression of many tumors; however, the specific mechanisms involved remain unknown. Here, we construct a pyroptosis-related gene signature that can be used to predict survival prognosis of skin cutaneous melanoma (SKCM) and provide guidance for clinical treatment. Methods: By integrating data from the two databases from the GTEx and TCGA, differentially expressed genes (DEGs) from normal tissues and skin cutaneous tumor tissues were identified. The main signaling pathways and function enrichment of these differential genes were determined. Univariate and multivariate COX regression analysis, and risk score analysis were used to construct a signature to assess its predictive value for overall survival. The mRNA expression of these five genes in melanoma cells was determined by quantitative polymerase chain reaction (qPCR). The pRRophetic algorithm was used to estimate the half-maximal inhibitory concentration (IC50) of chemotherapy drugs in SKCM patients. The expression of multiple immune checkpoint genes also was evaluated. Results: Sixteen DEGs associated with pyroptosis in SKCM and normal skin tissues were identified. Of these, 12 pyroptosis-related DEGs were associated with the prognosis of SKCM. A five-gene signature (GSDMA, GSDMC, IL-18, NLRP6, and AIM2) model was constructed. Patients were divided into high-risk and low-risk groups using the risk scores. Of these, the high-risk group had a worse survival prognosis. There are significant differences in the predicted sensitivity of the high-risk and low-risk groups to chemotherapeutic drugs. In addition, compared with the high-risk group, the low-risk group showed higher expression of PD-1, PDL-1, CTLA-4, LAG-3, and VSIR. Conclusion: In this study, we constructed a novel prognostic pyroptosis-related gene-signature for SKCM. These genes showed good predictive value for patient prognosis and could provide guidance for better treatment of SKCM patients. [ABSTRACT FROM AUTHOR]

Background: Myocardial ischemia reperfusion injury (MIRI) is an important mechanism of post-myocardial infarction injury and a main cause of death in patients with ischemic heart disease. Electroacupuncture (EA) pretreatment is effective for the prevention and treatment of MIRI, but mechanisms mediating the effects of cardiovascular disease EA treatments remain unclear. Objectives: To determine whether the lateral hypothalamus (LHA) and the cerebellar fastigial nucleus (FN) are involved in the protective effects of EA stimulation on MIRI. Methods: EA pretreatment was performed for 7 days before the establishment of the MIRI model. ST-segment changes on electrocardiograms were recorded and the Curtis-Walker arrhythmia score was used to evaluate changes in reperfusion injury. Hematoxylin-eosin staining was applied to evaluate the pathological and morphological changes in myocardial tissue. c-fos expression in the LHA and FN was determined by immunofluorescence staining. Glutamic (Glu) and γ-Aminobutyric acid (GABA) levels were measured using a high-performance liquid chromatography-electrochemical method. Results: EA pretreatment reduced ST-segment elevation, arrhythmia scores, and morphological changes in MIRI myocardial cells in rats, and decreased the c-fos protein expression in LHA/FN nuclei. MIRI was associated with an imbalance between GABA and Glu levels, whereas EA pretreatment increased GABA levels and decreased Glu levels in the LHA/FN. Conclusion: FN and LHA are involved in the EA-mediated attenuation of MIRI. Pretreatment with EA plays a protective role in the myocardium by regulating Glu and GABA release in the LHA and FN. [ABSTRACT FROM AUTHOR]

Summary: The tea plant (Camellia sinensis) is a thermophilic cash crop and contains a highly duplicated and repeat‐rich genome. It is still unclear how DNA methylation regulates the evolution of duplicated genes and chilling stress in tea plants. We therefore generated a single‐base‐resolution DNA methylation map of tea plants under chilling stress. We found that, compared with other plants, the tea plant genome is highly methylated in all three sequence contexts, including CG, CHG and CHH (where H = A, T, or C), which is further proven to be correlated with its repeat content and genome size. We show that DNA methylation in the gene body negatively regulates the gene expression of tea plants, whereas non‐CG methylation in the flanking region enables a positive regulation of gene expression. We demonstrate that transposable element‐mediated methylation dynamics significantly drives the expression divergence of duplicated genes in tea plants. The DNA methylation and expression divergence of duplicated genes in the tea plant increases with evolutionary age and selective pressure. Moreover, we detect thousands of differentially methylated genes, some of which are functionally associated with chilling stress. We also experimentally reveal that DNA methyltransferase genes of tea plants are significantly downregulated, whereas demethylase genes are upregulated at the initial stage of chilling stress, which is in line with the significant loss of DNA methylation of three well‐known cold‐responsive genes at their promoter and gene body regions. Overall, our findings underscore the importance of DNA methylation regulation and offer new insights into duplicated gene evolution and chilling tolerance in tea plants. Significance Statement: Our results show solid evidence that DNA methylation not only drives transposable element (TE)‐mediated functional diversification of duplicated genes in the tea plant, but also facilitates the enhancement of chilling tolerance in the tea plant through suppressing the activities of DNA methyltransferase under chilling stress. The overall findings underscore the importance of DNA methylation regulation on gene expression through TE insertions and broaden our understanding of duplicated gene evolution and chilling tolerance in plants. [ABSTRACT FROM AUTHOR]

Galangin, a natural flavonoid product derived from the root of galangal, is emerging as a promising anticancer agent against multiple cancers. Yet, whether it also has antitumor effects on cholangiocarcinoma (CCA) and the underlying mechanism is still unknown. Herein, we demonstrate that galangin exhibits multiple antitumor effects on CCA cells including decreases cell viability; inhibits proliferation, migration, and invasion; and induces apoptosis. Moreover, those phenotypic changes are associated with downregulated microRNA-21 (miR-21) expression. To support, overexpression of miR-21 blocks galangin-mediated antisurvival and metastasis effects on CCA cells. Mechanically, galangin increases the expression of phosphatase and tensin homolog (PTEN), a direct target of miR-21, resulting in decreased phosphorylation of AKT, a protein kinase which plays a critical role in controlling survival and apoptosis. In contrast, overexpression of miR-21 abrogates galangin-regulated PTEN expression and AKT phosphorylation. Taken together, these findings indicate that galangin inhibits CCA cell proliferation and metastasis and induces cell apoptosis through a miR-21-dependent manner, and galangin may provide a novel potential therapeutic adjuvant to treat CCA. [ABSTRACT FROM AUTHOR]

Enterovirus A71 (EV-A71) is transmitted through the respiratory tract, gastrointestinal system, and fecal-oral routes. The main symptoms caused by EV-A71 are hand, foot, and mouth disease (HFMD) or vesicular sore throat. Upf1 (Up-frameshift protein 1) was reported to degrade mRNA containing early stop codons, known as nonsense-mediated decay (NMD). Upf1 is also involved in the NMD mechanism as a host factor detrimental to viral replication. In this study, we dissected the potential roles of Upf1 in the EV-A71-infected cells. Upf1 was virulently down-regulated in three different EV-A71-infected cells, RD, Hela, and 293T, implying that Upf1 is a host protein unfavorable for EV-A71 replication. Knockdown of Upf1 protein resulted in increased viral RNA expression and production of progeny virus, and conversely, overexpression of Upf1 protein resulted in decreased viral RNA expression and production of progeny virus. Importantly, we observed increased RNA levels of asparagine synthetase (ASNS), one of the indicator substrates for the NMD mechanism, which indirectly suggests that EV-A71 infection of cells suppresses NMD activity in the host. The results shown in this study are useful for subsequent analysis of the relationship between the NMD/Upf1 mechanism and other picornaviruses, which may lead to the development of anti-picornavirus drugs. [ABSTRACT FROM AUTHOR]

Main conclusion: Circular RNA (circRNA) discovery, expression patterns and experimental validation in developing tea leaves indicates its correlation with circRNA-parental genes and potential roles in ceRNA interaction network.Circular RNAs (circRNAs) have recently emerged as a novel class of abundant endogenous stable RNAs produced by circularization with regulatory potential. However, identification of circRNAs in plants, especially in non-model plants with large genomes, is challenging. In this study, we undertook a systematic identification of circRNAs from different stage tissues of tea plant (Camellia sinensis) leaf development using rRNA-depleted circular RNA-seq. By combining two state-of-the-art detecting tools, we characterized 3174 circRNAs, of which 342 were shared by each approach, and thus considered high-confidence circRNAs. A few predicted circRNAs were randomly chosen, and 20 out of 24 were experimental confirmed by PCR and Sanger sequencing. Similar in other plants, tissue-specific expression was also observed for many C. sinensis circRNAs. In addition, we found that circRNA abundances were positively correlated with the mRNA transcript abundances of their parental genes. qRT-PCR validated the differential expression patterns of circRNAs between leaf bud and young leaf, which also indicated the low expression abundance of circRNAs compared to the standard mRNAs from the parental genes. We predicted the circRNA-microRNA interaction networks, and 54 of the differentially expressed circRNAs were found to have potential tea plant miRNA binding sites. The gene sets encoding circRNAs were significantly enriched in chloroplasts related GO terms and photosynthesis/metabolites biosynthesis related KEGG pathways, suggesting the candidate roles of circRNAs in photosynthetic machinery and metabolites biosynthesis during leaf development. [ABSTRACT FROM AUTHOR]

Background: Non-small-cell lung cancer (NSCLC) is the major leading cause of cancer-related death around the world. The resistance to chemotherapy limits the effects of clinical treatment. The aim of this study was to identify novel mechanisms involved in NSCLC chemoresistance. Materials and methods: We explored the public database and commercial tissue microarray to evaluate the expression of G protein-coupled receptor 35 (GPR35). We established the chemoresistant A549 cell line to further investigate the biological function of GPR35 in vitro and in vivo. Then, we measured the altered signalings that GPR35 knocking down by Western blot assay. Results: We demonstrated that GPR35 expression was significantly elevated in NSCLC tissues and correlated with poor prognosis. GPR35 was upregulated in our in vitro chemoresistance cell model. GPR35 depletion reduced the half maximal inhibitory concentration of chemodrugs and restored the sensitivity both in vitro and in vivo. Mechanically, we found that GPR35-mediated chemoresistance occurred partially via β-arrestin-2/Akt signaling. Furthermore, inhibition of β-arrestin-2 or Akt activation could suppress the GPR35 expression and overcome chemoresistance. Conclusion: Our results suggested that GPR35 might serve as a novel therapeutic target to enhance the chemotherapy efficacy in NSCLC. [ABSTRACT FROM AUTHOR]

N6-methyladenosine (m6A) RNA methylation is the most abundant modification on mRNAs and plays important roles in various biological processes. The formation of m6A is catalyzed by a methyltransferase complex including methyltransferase-like 3 (METTL3) as a key factor. However, the in vivo functions of METTL3 and m6A modification in mammalian development remain unclear. Here, we show that specific inactivation of Mettl3 in mouse nervous system causes severe developmental defects in the brain. Mettl3 conditional knockout (cKO) mice manifest cerebellar hypoplasia caused by drastically enhanced apoptosis of newborn cerebellar granule cells (CGCs) in the external granular layer (EGL). METTL3 depletion–induced loss of m6A modification causes extended RNA half-lives and aberrant splicing events, consequently leading to dysregulation of transcriptome-wide gene expression and premature CGC death. Our findings reveal a critical role of METTL3-mediated m6A in regulating the development of mammalian cerebellum. [ABSTRACT FROM AUTHOR]

Objective: To establish an unbiased, 3‐dimensional (3‐D) approach that quantifies subchondral bone plate (SBP) changes in mouse joints, and to investigate the mechanism that mediates SBP sclerosis at a late stage of osteoarthritis (OA). Methods: A new micro‐computed tomography (micro‐CT) protocol was developed to characterize the entire thickness of the SBP in the distal femur of a normal mouse knee. Four mouse models of severe joint OA were generated: cartilage‐specific Egfr–knockout (Egfr‐CKO) mice at 2 months after surgical destabilization of the medial meniscus (DMM), Egfr‐CKO mice with aging‐related spontaneous OA, wild‐type (WT) mice at 10 months after DMM, and WT mice at 14 weeks after DMM plus hemisectomy of the meniscus (DMMH) surgery. As an additional model, mice with knockout of the sclerostin gene (Sost‐KO) were subjected to DMMH surgery. Knee joints were examined by micro‐CT, histology, and immunohistochemical analyses. Results: Examination of the mouse distal femur by 3‐D micro‐CT revealed a positive correlation between SBP thickness and the loading status in normal knees. In all 4 mouse models of late‐stage OA, SBP sclerosis was restricted to the areas under severely eroded articular cartilage. This was accompanied by elevated bone formation at the bone marrow side of the SBP and a drastic reduction in the levels of sclerostin in osteocytes within the SBP. Unlike in WT mice, no further increase in the thickness of the SBP was observed in response to DMMH in Sost‐KO mice. Conclusion: Since focal stress on the SBP underlying sites of cartilage damage increases during late stages of OA, these findings establish mechanical loading–induced attenuation of sclerostin expression and elevation of bone formation along the SBP surface as the major mechanisms characterizing subchondral bone phenotypes associated with severe late‐stage OA in mice. [ABSTRACT FROM AUTHOR]

Diabetic nephropathy (DN) is one of the most important causes that leads to end-stage renal disease and the efficacy of strategies currently available for the prevention of DN remains unsatisfactory. Sitagliptin (SIT), which is a dipeptidyl peptidase-4 inhibitor, exhibited a modest beneficial effect on glycated hemoglobin levels and is capable of ameliorating renal ischemia reperfusion injury. By determining the expression of transforming growth factor-ß1 (TGF-ß1), connective tissue growth factor (CTGF), collagen type IV (ColIV) and fibronectin (FN) levels in high glucose-cultured glomerular mesangial cells (MCs), the present study aimed to assess the anti-proliferative and anti-fibrotic effects of SIT on the therapeutic potential for the prevention of DN and its mechanism. Specifically, cell proliferation was determined via cell counting kit-8 assay, and the expression levels of TGF-ß1 and CTGF mRNA were detected by reverse transcription polymerase chain reaction analysis. Furthermore, the secretion of TGF-ß1, CTGF, ColIV and FN proteins was measured via enzyme-linked immunosorbent assays. The results demonstrated that high glucose induced the proliferation of MCs and enhanced the expression of TGF-ß1, CTGF, ColIV and FN. Furthermore, treatment with SIT inhibited cell proliferation and the expression of TGF-ß1, CTGF, ColIV and FN induced by high glucose. In conclusion, SIT inhibits cell proliferation and the expression of the major extracellular matrix proteins induced by high glucose, indicating its value for treating or relieving DN. [ABSTRACT FROM AUTHOR]

Ankylosing spondylitis (AS) is a severe chronic inflammatory disease that may ultimately result in the development of a 'bamboo-like' spine. Although the pathological changes that occur in AS have been extensively investigated, the mechanism underlying spinal fusion during AS remains elusive. Differentially expressed genes (DEGs) in paraspinal tissues from patients with AS compared with those from healthy controls were therefore investigated. Polymerase chain reaction (PCR)-based suppression subtractive hybridization was performed using total mRNA from the supraspinal ligaments of three patients with AS and three patients with spinal fractures as controls. From this, 27 genes were identified in all of the three independent forward libraries, which were defined as DEGs associated with AS. Reverse transcription-quantitative PCR demonstrated that six DEGs were overexpressed in the tissues from patients with AS compared with those from individuals in the control group, including those encoding transforming growth factor β types I and III receptor, vascular endothelial growth factor, matrix metalloproteinase-3, core-binding factor α1 and bone morphogenetic protein 2. Western blot analysis showed increased expression in all six of these proteins in the samples from patients with AS compared with those in the control groups. These findings suggested that changes in the expression of these genes and proteins are associated with the development of spinal fusion during the pathogenesis of AS. Furthermore, these genes may be novel markers of the risk of developing AS, in addition to being targets for the treatment of this disease. [ABSTRACT FROM AUTHOR]

Perilla frutescens (L.) Britt. is a self-pollinating annual species and is widely cultivated in China, Korea and Japan as an economic crop and a source of medicine and spices. In this study, we sequenced one cultivar variety (PF98095) of P. frutescens (L.) var. frutescens Britt., which was assembled as reference and other three varieties (PF11109, weedy of var. frutescens , PF06336 and PF06353, cultivars of varieties crispa ) in order to carry out comparative expression profiling within cultivar and weedy in varieties frutescens and between varieties frutescens and varieties crispa of cultivar type in P. frutescens . Assembly of PF98095, annotation mapping, DEG (differentially expressed gene) profiling, and comparative analysis were performed. We found that more than 65% of the reads were mapped to the reference of P. frutescens gene set. Moreover, we detected 22,962 DEGs in the weedy variety compared to the cultivar, and also, 22,138 and 23,845 DEGs were identified in two cultivars according to the reference, respectively. The DEGs and functional classification were developed to analyze the differences between weedy and cultivar and between varieties frutescens and varieties crispa of Perilla . Furthermore, candidate genes for the different color and seed size of Perilla were identified that could be further investigated in future study. The herein results may play a significant role, and contribute in functional transcriptome studies of Perilla . [ABSTRACT FROM AUTHOR]

Purpose. To explore the association between differential Tim-3 and PD-1 expression patterns and HIV disease progression, and to investigate the impact of common γ-chain cytokines on Tim-3 and PD-1 expression patterns on T cells. Methods. Tim-3/PD-1 expression on the T cells of patients with early and chronic HIV infections was detected. The expression levels and functional profiles of T cells with differential Tim-3 and PD-1 expression patterns induced by γ-chain cytokines were studied. Results. The elevation of differential Tim-3 and PD-1 expression patterns on T cells appeared early in HIV infection. Co-expression of Tim-3 and PD-1 (Tim-3+PD-1+) correlates with more severe exhaustion of T cells during HIV infection. In vitro stimulation of common γ-chain cytokines can induce differential expression patterns of Tim-3 and PD-1 on T cells. The enhancement of Tim-3 and PD-1 expression by common γ-chain IL-2 can inhibit the function of T cells re-stimulated by HIV gag and TCR, not by the re-stimulation of IL-2. Conclusions. The elevation of differential Tim-3 and PD-1 expression patterns on T cells represents a state of T cell exhaustion and can be induced by common γ-chain cytokines. These findings provide insights into HIV pathogenesis and help inform immune intervention strategies. [ABSTRACT FROM AUTHOR]

Methionine sulfoxide reductase plays a regulatory role in plant growth and development, especially in scavenging reactive oxygen species by restoration of the oxidation of methionine in protein. A fulllength cDNA sequence encoding methionine sulfoxide reductase (MSR) from mulberry, which we designated MMSR, was cloned based on mulberry expressed sequence tags (ESTs). Sequence analysis showed that the MMSR is 810 bp long, encoding 194 amino acids with a predicted molecular weight of 21.6 kDa and an isoelectric point of 6.78. The expression level of the MMSR gene under conditions of drought and salt stresses was quantified by qRT-PCR. The results show that the expression level changed significantly under the stress conditions compared to the normal growth environment. It helps us to get a better understanding of the molecular basis for signal transduction mechanisms underlying the stress response in mulberry. [ABSTRACT FROM AUTHOR]

Upon aging, the number of hematopoietic stem cells (HSCs) in the bone marrow increases while their repopulation potential declines. Moreover, aged HSCs exhibit lineage bias in reconstitution experiments with an inclination toward myeloid at the expense of lymphoid potential. The adaptor protein Lnk is an important negative regulator of HSC homeostasis, as Lnk deficiency is associated with a 10-fold increase in HSC numbers in young mice. However, the age-related increase in functional HSC numbers found in wild-type HSCs was not observed in Lnk-deficient animals. Importantly, HSCs from aged Lnk null mice possess greatly enhanced self-renewal capacity and diminished exhaustion, as evidenced by serial transplant experiments. In addition, Lnk deficiency ameliorates the aging-associated lineage bias. Transcriptome analysis revealed that WT and Lnk-deficient HSCs share many aging-related changes in gene expression patterns. Nonetheless, Lnk null HSCs displayed altered expression of components in select signaling pathways with potential involvement in HSC self-renewal and aging. Taken together, these results suggest that loss of Lnk partially mitigates age-related HSC alterations. [ABSTRACT FROM AUTHOR]

Hematopoietic stem cell (HSC) function is tightly regulated by cytokine signaling. Although phospho-flow cytometry allows us to study signaling in defined populations of cells, there has been tremendous hurdle to carry out this study in rare HSCs due to unrecoverable critical HSC markers, low HSC number, and poor cell recovery rate. Here, we overcame these difficulties and developed a 'HSC phospho-flow' method to analyze cytokine signaling in murine HSCs at the single-cell level and compare HSC signaling profile to that of multipotent progenitors (MPPs), a cell type immediately downstream of HSCs, and commonly used Lin− cKit+ cells (LK cells, enriched for myeloid progenitors). We chose to study signaling evoked from three representative cytokines, stem cell factor (SCF) and thrombopoietin (TPO) that are essential for HSC function and granulocyte macrophage-colony-stimulating factor (GM-CSF) that is dispensable for HSCs. HSCs display a distinct TPO and GM-CSF signaling signature from MPPs and LK cells, which highly correlates with receptor surface expression. In contrast, although majority of LK cells express lower levels of cKit than HSCs and MPPs, SCF-evoked ERK1/2 activation in LK cells shows a significantly increased magnitude for a prolonged period. These results suggest that specific cellular context plays a more important role than receptor surface expression in SCF signaling. Our study of HSC signaling at the homeostasis stage paves the way to investigate signaling changes in HSCs under conditions of stress, aging, and hematopoietic diseases. S tem C ells 2012;30:1447-1514 [ABSTRACT FROM AUTHOR]

The SSM/SLSM (spliceosomal Smith (SM)/SM-like (LSM)) genes are the central components of the spliceosome in eukaryotes, which play an important role in regulating RNA splicing, participating in diverse biological processes. Although it has been detected in Arabidopsis and rice etc. plants, the members and significance of the SSM/SLSM gene family in wheat are still not reported. In this study, we identified the SSM/SLSM genes in wheat and its progenitors at genome-scale, where 57 SSM/SLSM genes were identified in wheat, together with 41, 17and 19 found in Triticum dicoccoides, Triticum urartu, and Aegilops tauschii. Furthermore, their phylogenetic relationship, gene structures, conserved motifs, and cis-regulatory elements were systematically analyzed. By synteny analysis, good collinearity of SSM/SLSM genes was found among bread wheat and its progenitors' genomes, and the distribution of SMD2 genes in wheat chromosome 5A, 4B and 4D located in the 4AL-5AL-7BS chromosome model, due to the translocation. Then, the positively selected genes were further investigated based on the non-synonymous to synonymous (dN/dS) analysis of the orthologous pairs. Finally, the expression profiles of the SSM/SLSM genes were detected using RNA-seq datasets, and eight stress-responsive candidate genes were selected to validate their expression through qPCR (real-time quantitative polymerase chain reaction). According to the co-expression network analysis, the correlation between the LSM7-7A gene and related genes was illustrated through Gene Ontology (GO) enrichment analysis. Furthermore, the LSM7-7A gene was related to the Arabidopsis homologous salt tolerance gene RCY1. This investigation systematically identified the complete candidates of SSM/SLSM genes and their characters in wheat and its progenitors, and provided clues to a better understanding of their contribution during the wheat polyploidy process. [ABSTRACT FROM AUTHOR]

Pancreatic stellate cells (PSCs) are crucial for metabolism and disease progression in pancreatic ductal adenocarcinoma (PDAC). However, detailed mechanisms of PSCs in glutamine (Gln) metabolism and tumor-stromal metabolic interactions have not been well clarified. Here we showed that tumor tissues displayed Gln deficiency in orthotopic PDAC models. Single-cell RNA sequencing analysis revealed metabolic heterogeneity in PDAC, with significantly higher expression of Gln catabolism pathway in stromal cells. Significantly higher glutamine synthetase (GS) protein expression was further validated in human tissues and cells. Elevated GS levels in tumor and stroma were independently prognostic of poorer prognosis in PDAC patients. Gln secreted by PSCs increased basal oxygen consumption rate in PCCs. Depletion of GS in PSCs significantly decreased PCCs proliferation in vitro and in vivo. Mechanistically, activation of Wnt signaling induced directly binding of β-catenin/TCF7 complex to GS promoter region and upregulated GS expression. Rescue experiments testified that GS overexpression recovered β-catenin knockdown-mediated function on Gln synthesis and tumor-promoting ability of PSCs. Overall, these findings identify the Wnt/β-catenin/TCF7/GS-mediated growth-promoting effect of PSCs and provide new insights into stromal Gln metabolism, which may offer novel therapeutic strategies for PDAC. • Metabolic heterogeneity generates Gln crosstalk between tumor and stroma in PDAC. • Gln synthesis is higher in PSCs compared to PCCs. • High GS expression in either tumor or stroma associates with poor prognosis in PDAC patients. • Targeting stromal GS induces tumor regression in vitro and in vivo. • The Wnt-β-catenin-TCF7-GS-Gln axis plays a key role in the growth-promoting effect of PSCs in PDAC. [ABSTRACT FROM AUTHOR]

Chronic myelogenous leukemia (CML) results from transformation of a long-term hematopoietic stem cell (LTHSC) by expression of the BCR-ABL fusion gene. However, BCR-ABL-expressing LTHSCs are heterogeneous in their capacity as leukemic stem cells (LSCs). Although discrepancies in proliferative, self-renewal, and differentiation properties of normal LTHSCs are being increasingly recognized, the mechanisms underlying heterogeneity of leukemic LTHSCs are poorly understood. Using a CML mouse model, we identified gene expression differences between leukemic and nonleukemic LTHSCs. Expression of the thrombopoietin (THPO) receptor MPL was elevated in leukemic LTHSC populations. Compared with LTHSCs with low MPL expression, LTHSCs with high MPL expression showed enhanced JAK/STAT signaling and proliferation in response to THPO in vitro and increased leukemogenic capacity in vivo. Although both G0 and S phase subpopulations were increased in LTHSCs with high MPL expression, LSC capacity was restricted to quiescent cells. Inhibition of MPL expression in CML LTHSCs reduced THPO-induced JAK/STAT signaling and leukemogenic potential. These same phenotypes were also present in LTHSCs from patients with CML, and patient LTHSCs with high MPL expression had reduced sensitivity to BCR-ABL tyrosine kinase inhibitor treatment but increased sensitivity to JAK inhibitors. Together, our studies identify MPL expression levels as a key determinant of heterogeneous leukemia-initiating capacity and drug sensitivity of CML LTHSCs and suggest that high MPL-expressing CML stem cells are potential targets for therapy. [ABSTRACT FROM AUTHOR]

Highlights: [•] Lentiviral vector construction and virus packaging. [•] Lentiviral ERK2 siRNA detection both in vitro and vivo. [•] Lentiviral ERK2 siRNA is able to prevent collagen expression and scar adhesion. [•] Lentiviral ERK2 siRNA is able to suppress the expression of TGF-β1 and IL-6. [ABSTRACT FROM AUTHOR]

Abstract: Tuberculosis remains a major infectious disease worldwide due to the low efficacy of available vaccine of the Mycobacterium bovis Bacillus Calmette–Guérin (BCG). DNA vaccines are especially promising candidates; however, the efficacy of DNA vaccine expressing single antigen of Mycobacterium tuberculosis (MTb) is limited. In this study, a plasmid DNA vaccine, pAEH, was constructed and designed to express a fusion protein of the Ag85B, Esat6, and HspX of MTb. Its immunogenicity and protective efficacy as well as therapeutic effect were assessed in a mouse model of tuberculosis. Vaccination with the pAEH significantly increased the frequency of peripheral blood CD4+ and CD8+ T cells, but not γδT cells, similar to that of vaccination with the BCG, and induced significantly higher levels of HspX-specific T cell proliferation, as compared with vaccination with BCG or the pHspX. Furthermore, vaccination with the pAEH increased the frequency of Ag85B, Esat6 and HspX-specific IFNγ-secreting T cells, accompanied by significantly higher levels of IFN-γ and IL-2 production ex vivo, as compared with that of the BCG or pHspX-vaccinated mice. Apparently, vaccination with the pAEH induced potent Th1 responses in mice. More importantly, vaccination with the pAEH inhibited the replication of virulent MTb in the lungs and spleens, even after MTb infection, and related lung inflammation in mice. Potentially, the newly developed pAEH vaccine may be used for the prevention and therapeutic intervention of MTb infection. [Copyright &y& Elsevier]