CD93 regulates breast cancer growth and vasculogenic mimicry through the PI3K/AKT/SP2 signaling pathway activated by integrin ß1.

In women, breast cancer (BC) accounts for 7%-10% of all cancer cases and is one of the most common cancers. To identify a new method for treating BC, the role of CD93 and its underlying mechanism were explored. MDA-MB-231 cells were used in this study and transfected with si-CD93, si-MMRN2, oe-CD93, si-integrin ß1, or oe-SP2 lentivirus. After MDA-MB-231 cells were transfected with si-NC or si-CD93, they were injected into nude mice by subcutaneous injection at a dose of 5 × 106/mouse to construct a BC animal model. The expression of genes and proteins and cell migration, invasion and vasculogenic mimicry were detected by RT‒qPCR, western blot, immunohistochemistry, immunofluorescence, Transwell, and angiogenesis assays. In pathological samples and BC cell lines, CD93 was highly expressed. Functionally, CD93 promoted the proliferation, migration, and vasculogenic mimicry of MDA-MB-231 cells. Moreover, CD93 interacts with MMRN2 and integrin ß1. Knockdown of CD93 and MMRN2 can inhibit the activation of integrin ß1, thereby inhibiting the PI3K/AKT/SP2 signaling pathway and inhibiting BC growth and vasculogenic mimicry. In conclusion, the binding of CD93 to MMRN2 can activate integrin ß1, thereby activating the PI3K/AKT/SP2 signaling pathway and subsequently promoting BC growth and vasculogenic mimicry.

Nitrogen starvation modulates the sensitivity of rhizobacterial community to drought stress in Stevia rebaudiana.

Alterations in water regimes or nitrogen (N) availability lead to shifts in the assemblage of rhizosphere microbial community; however, how the rhizosphere microbiome response to concurrent changes in water and N availability remains largely unclear. Herein, we investigated the taxonomic and functional characteristics of rhizobacteria associated with stevia (Stevia rebaudiana Bertoni) under varying combinations of water and N levels. Community diversity and predicted functions of rhizobacteria were predominantly altered by drought stress, with N-starvation modulating these effects. Moreover, N fertilization simplified the ecological interactions within rhizobacterial communities and heightened the relative role of stochastic processes on community assembly. In terms of rhizobacterial composition, we observed both common and distinctive changes in drought-responsive bacterial taxa under different N conditions. Generally, the relative abundance of Proteobacteria and Bacteroidetes phyla were depleted by drought stress but the Actinobacteria phylum showed increases. The rhizobacterial responses to drought stress were influenced by N availability, where the positive response of δ-proteobacteria and the negative response of α- and γ-proteobacteria, along with Bacteroidetes, were further heightened under N starvation. By contrast, under N fertilization conditions, an amplified negative or positive response to drought were demonstrated in Firmicutes and Actinobacteria phyla, respectively. Further, the drought-responsive rhizobacteria were mostly phylogenetically similar, but this pattern was modulated under N-rich conditions. Overall, our findings indicate an N-dependent specific restructuring of rhizosphere bacteria under drought stress. These changes in the rhizosphere microbiome could contribute to enhancing plant stress tolerance.

Selective activation of cholinergic neurotransmission from the medial septal nucleus to hippocampal pyramidal neurones improves sepsis-induced cognitive deficits in mice.

BACKGROUND: Sepsis-associated encephalopathy is characterised by cognitive dysfunction, and might be mediated by deficits in neurotransmission. Reduced cholinergic neurotransmission in the hippocampus impairs memory function. We assessed real-time alterations of acetylcholine neurotransmission from the medial septal nucleus to the hippocampus, and explored whether sepsis-induced cognitive deficits can be relieved by activating upstream cholinergic projections. METHOD: Lipopolysaccharide (LPS) injection or caecal ligation and puncture (CLP) was used to induce sepsis and associated neuroinflammation in wild-type and mutant mice. Adeno-associated viruses for calcium and acetylcholine imaging, and for optogenetic and chemogenetic modulation of cholinergic neurones were injected into the hippocampus or medial septum, and a 200-µm-diameter optical fibre was implanted to collect acetylcholine and calcium signals. Cholinergic activity of the medial septum was manipulated and combined with cognitive assessment after LPS injection or CLP. RESULTS: Intracerebroventricular LPS injection reduced postsynaptic acetylcholine (from 0.146 [0.001] to 0.0047 [0.0005]; p=0.004) and calcium (from 0.0236 [0.0075] to 0.0054 [0.0026]; p=0.0388) signals in hippocampal Vglut2-positive glutamatergic neurones, whereas optogenetic activation of cholinergic neurones in the medial septum reversed LPS-induced reductions in these two signals. Intraperitoneal LPS injection decreased acetylcholine concentration in the hippocampus (476 [20] pg ml-1 to 382 [14] pg ml-1; p=0.0001). Reduction in long-term potentiation (238 [23] % to 150 [12] %; p=0.0082) and enhancement of hippocampal pyramidal neurone action potential frequency (5.8 [1.5] Hz to 8.2 [1.8] Hz; p=0.0343) were relieved, and neurocognitive performance was improved by chemogenetic activation of cholinergic innervation of the hippocampus 3 days after LPS injection in septic mice. CONCLUSIONS: Systemic or local LPS reduced cholinergic neurotransmission from the medial septum to hippocampal pyramidal neurones, and their selective activation alleviated defects in hippocampal neuronal function and synaptic plasticity and ameliorated memory deficits in sepsis model mice through enhanced cholinergic neurotransmission. This provides a basis for targeting cholinergic signalling to the hippocampus in sepsis-induced encephalopathy.

Comparative transcriptome analysis provides insights into steviol glycoside synthesis in stevia (Stevia rebaudiana Bertoni) leaves under nitrogen deficiency.

KEY MESSAGE: Transcriptome analysis revealed the potential mechanism of nitrogen regulating steviol glycosides synthesis via shifting of leaf carbon metabolic flux or inducing certain transcription factors. Nitrogen (N) plays key regulatory roles in both stevia (Stevia rebaudiana) growth and the synthesis of its functional metabolite steviol glycosides (SGs), but the mechanism by which this nutrient regulates SGs synthesis remains to be elucidated. To address this question, a pot experiment was performed in a greenhouse where stevia plants fertilized with N (the control as CK plants) and compared with plants without the supply of N. Physiological and biochemical analyses were conducted to test the growth and metabolic responses of plants to N regimes. Our results showed that N deficiency significantly inhibited plant growth and leaf photosynthesis, while increased leaf SGs contents in stevia (49.97, 46.64 and 84.80% respectively for rebaudioside A, stevioside, and rebaudioside C), which may be partly due to "concentration effect". Then, transcriptome analysis was conducted to understand the underlying mechanisms. A total of 535 differentially expressed genes were identified, and carbon metabolism-related events were highlighted by Gene Ontology and Kyoto Encyclopedia of Genes and Genomes. Many of these genes were significantly upregulated by N-deficiency, including those involved in "phenylpropanoid biosynthesis", "flavonoid biosynthesis" and "starch and sucrose metabolism". Our study also analyzed the expression patterns of SGs synthesis-related genes under two N regimes and the potential transcription factors linking N nutrition and SG metabolism. N-deficiency may promote SGs synthesis by changing the carbon metabolism flux or inducing certain transcription factors. Our results provide deeper insight into the relationship between N nutrition and SGs synthesis in stevia plants.

Application of E-nose technology combined with artificial neural network to predict total bacterial count in milk.

Total bacterial count (TBC) is a widely accepted index for assessing microbial quality of milk, and cultivation-based methods are commonly used as standard methods for its measurement. However, these methods are laborious and time-consuming. This study proposes a method combining E-nose technology and artificial neural network for rapid prediction of TBC in milk. The qualitative model generated an accuracy rate of 100% when identifying milk samples with high, medium, or low levels of TBC, on both the testing and validating subsets. Predicted TBC values generated by the quantitative model demonstrated strong coefficient of multiple determination (R2 > 0.99) with reference values. Mean relative difference between predicted and reference values (mean ± standard deviation) of TBC were 1.1 ± 1.7% and 0.4 ± 0.8% on the testing and validating subsets involving 24 and 28 tested samples, respectively. Paired t-test implied that the difference between predicted and reference values of TBC was insignificant for both the testing and validating subsets. As low as ~1 log cfu/mL of TBC present in tested samples were precisely predicted. Results of this study indicated that combination of E-nose technology and artificial neural network generated reliable predictions of TBC in milk. The method proposed in this study was reliable, rapid, and cost efficient for assessing microbial quality milk, and thus would potentially have realistic application in dairy section.

Transcriptomic Characterization of Nitrate-Enhanced Stevioside Glycoside Synthesis in Stevia (Stevia rebaudiana) Bertoni.

Nitrogen forms (nitrate (NO3-) or ammonium (NH4+)) are vital to plant growth and metabolism. In stevia (Stevia rebaudiana), it is important to assess whether nitrogen forms can influence the synthesis of the high-value terpene metabolites-steviol glycosides (SGs), together with the underlying mechanisms. Field and pot experiments were performed where stevia plants were fertilized with either NO3- or NH4+ nutrition to the same level of nitrogen. Physiological measurements suggested that nitrogen forms had no significant impact on biomass and the total nitrogen content of stevia leaves, but NO3--enhanced leaf SGs contents. Transcriptomic analysis identified 397 genes that were differentially expressed (DEGs) between NO3- and NH4+ treatments. Assessment of the DEGs highlighted the responses in secondary metabolism, particularly in terpenoid metabolism, to nitrogen forms. Further examinations of the expression patterns of SGs synthesis-related genes and potential transcription factors suggested that GGPPS and CPS genes, as well as the WRKY and MYB transcription factors, could be driving N form-regulated SG synthesis. We concluded that NO3-, rather than NH4+, can promote leaf SG synthesis via the NO3--MYB/WRKY-GGPPS/CPS module. Our study suggests that insights into the molecular mechanism of how SG synthesis can be affected by nitrogen forms.

A beta-glucosidase gene from Stevia rebaudiana may be involved in the steviol glycosides catabolic pathway.

We herein report the preparation of a full-length raucaffricine-O-beta-D-glucosidase gene of stevia rebaudiana Bertoni (named SrRG1, GenBank accession number MK920450). Sequence analysis indicated SrRG1 consists of a 1650 bp open reading frame encoding a protein of 549 amino acids. Its deduced amino acid sequence showed a high identity of 82% with a raucaffricine-O-beta-D-glucosidase from H. annuus of glycoside hydrolase family 1. The expression pattern analyzed by real-time quantitative PCR showed no significant difference among different tissues, developmental stages, and cultivars under normal growth conditions. Furthermore, the gene function of SrRG1 was preliminarily studied by agrobacterium-mediated transformation on instantaneous expression. In the test of agrobacterium-mediated transformation on instantaneous expression, it was observed that overexpression of SrRG1 increased the accumulation of steviol content and decreased the major components and total SGs contents. Such results demonstrated that SrRG1 may participate in the steviol glycosides catabolic pathway. However, the effect of silencing construct infiltration on steviol and SGs content was not significant and its expression pattern was constitutive, which most probably, attributed the hydrolysis of SGs to the secondary activity of SrRG1. This study firstly identified the bate-glucosidase in stevia and advances our understanding of steviol glycosides hydrolyzation.

Identification of GH1 gene family fgt members in Stevia rebaudiana and their expression when grown in darkness.

Stevia rebaudiana Bertoni is an important economic crop that is well known for its secondary metabolites, steviol glycosides (SGs), found in leaves. Because the enzymes of deglycosylation (glycoside hydrolases) play important roles in SGs biosynthetic processes, our study is focused on the functions of ß-glucosidases in SGs catabolism in stevia. We cloned and characterized 19 stevia GH1 genes based on transcriptomic sequences. The 19 genes were divided into five putative subfamilies in Arabidopsis. Conserved motifs in the SrGH1 proteins were analysed using the online motif-based sequence analysis tool, MEME. Most of the identified proteins contain the conserved 'TFNEP' motif (contains the catalytic acid/base) and 'ITENG' motif (contains the catalytic nucleophile). Furthermore, the steviol glycoside content and expression of these 19 genes were characterized under constant darkness. The dark treatment lowered the steviol glycoside content significantly, while SrBGLU16 responded to darkness and was markedly upregulated. This study is the first transcriptome-wide analysis of the GH1 family in Stevia rebaudiana. The sequences of 19 SrGH1 members and their expression when grown in darkness were characterized. Among the 19 genes, SrBGLU16 was markedly upregulated by darkness. Thus, we identified SrBGLU16 for further investigation as a possible steviol glycoside beta-glucosidase.

The evolutionary history of the sucrose synthase gene family in higher plants.

BACKGROUND: Sucrose synthase (SUS) is widely considered a key enzyme participating in sucrose metabolism in higher plants and regarded as a biochemical marker for sink strength in crops. However, despite significant progress in characterizing the physiological functions of the SUS gene family, knowledge of the trajectory of evolutionary processes and significance of the family in higher plants remains incomplete. RESULTS: In this study, we identified over 100 SUS genes in 19 plant species and reconstructed their phylogenies, presenting a potential framework of SUS gene family evolution in higher plants. Three anciently diverged SUS gene subfamilies (SUS I, II and III) were distinguished based on their phylogenetic relationships and unique intron/exon structures in angiosperms, and they were found to have evolved independently in monocots and dicots. Each subfamily of SUS genes exhibited distinct expression patterns in a wide range of plants, implying that their functional differentiation occurred before the divergence of monocots and dicots. Furthermore, SUS III genes evolved under relaxed purifying selection in dicots and displayed narrowed expression profiles. In addition, for all three subfamilies of SUS genes, the GT-B domain was more conserved than the "regulatory" domain. CONCLUSIONS: The present study reveals the evolution of the SUS gene family in higher plants and provides new insights into the evolutionary conservation and functional divergence of angiosperm SUS genes.

Exogenous glutathione increased lead uptake and accumulation in Iris lactea var. chinensis exposed to excess lead.

Long- and short-term hydroponic experiments were conducted to study the effect of different concentrations of exogenous glutathione (GSH) on Pb uptake, translocation, and gene expresses in Iris lactea var. chinensis exposed to excess lead (Pb). Exogenous GSH remarkedly promoted Pb uptake and translocation in long-term (14 d) experiment, and the GSH-dose-dependent increases in shoot and root Pb contents existed obviously when GSH concentrations were lower than 800 mg·L-1. The fresh weight in gradual rise in plants was observed with the increase of exogenous GSH concentration. In short-term (24 h) experiment, Pb contents in roots under Pb with L-buthionine sulfoximine (BSO, a known inhibitor of GSH biosynthesis) treatments were significantly lower than that under Pb exposure alone. The transcript levels of three genes (Ilγ-ECS, IlGS, and IlPCS) involved in GSH synthesis and metabolism, showed no significant change in expression pattern except that upregulation after 24 h of treatment with Pb and GSH in comparison with that of the single Pb treatment. Further, the level of IlGS transcript after exposure for 4 h was much higher than that of Ilγ-ECS and IlPCS transcripts. All these results obtained here suggest that exogenous GSH can increase Pb accumulation, detoxification, and translocation to the shoot.

Enzyme treatment reverse transcription-PCR to differentiate infectious and inactivated F-specific RNA phages.

F-specific (F+) RNA phages are recommended as indicators of fecal contamination and the presence of enteric viruses and as viral surrogates to elucidate the resistance of viruses to adverse conditions or to assess the effectiveness of inactivating processes. Reverse transcription (RT)-PCR methods have been used to detect, quantify, or identify subgroups of F+ RNA phages. However, these methods may overestimate the infectivity of F+ RNA phages in test samples, since the presence of both infectious and inactivated phages (or naked RNA) can lead to positive RT-PCR signals. In this study, we evaluated the ability of an enzyme treatment (ET) with proteinase K and RNase A prior to RNA extraction, followed by RT-PCR, to differentiate infectious and inactivated F+ RNA phages. The results indicated that ET RT-PCR reduced, but did not completely eliminate, false-positive signals encountered with RT-PCR alone. The two-step ET RT-PCR, in which the enzymes were added sequentially, was more effective at reducing false-positive signals than the one-step ET RT-PCR, which involved addition of both enzymes together. Despite its inability to completely eliminate false-positive signals, ET RT-PCR gave more reliable information on the infectivity of F+ RNA phages. Thus, the method is better than RT-PCR alone for detecting F+ RNA phages as indicators to assess the risk of fecal contamination by enteric pathogens or to evaluate the effectiveness of virus-inactivating processes.

Relationship between the baking quality of wheat (Triticum aestivum L.) and the protein composition and structure after shading.

Although wheat (Triticum aestivum L.) grain protein content is increased by shade stress, the relationship between the baking quality of wheat flour and protein composition and structure remains unclear. Here, we investigated the effects of shade stress on wheat flour protein composition and structure. The contents of the flour protein, α/ß-gliadins and disulfide and hydrogen bonds were significantly increased by shade stress. Glutenins, UPP%, and ß-sheet contents also increased, whereas that of α-helices decreased. Spearman correlations revealed that the flour protein content, Glu:Gli ratio, and disulfide, hydrogen, and ionic bonds can predict the specific volume and number of crumb cells in bread, whereas α/ß-gliadins content can predict the crumb cell wall thickness and diameter of bread. Under shade stress, variations in protein composition and structure help increase the specific volume and crumb cells number and decrease crumb cell wall thickness and diameter of bread, ultimately leading to improved baking quality.

Changes in free amino acid and protein polymerization in wheat caryopsis and endosperm during filling after shading.

Over the past several decades, a decreasing trend in solar radiation has been observed during the wheat growing season. The effects of shade stress on grain yield formation have been extensively studied. However, little information on shade stress's effects on protein formation warrants further investigation. Two wheat cultivars were grown under three treatments, no shade as the control group (CK), shading from the joint to the anthesis stage (S1), and shading from the joint to the mature stage (S2), to investigate the effects of shade stress on the free amino acids of the caryopsis and endosperm and protein accumulation during grain filling. The dry mass of caryopsis and endosperm was significantly decreased under shade stress, whereas Glu, Ser, Ala, and Asp and protein relative content increased during grain filling. The observed increases in total protein in S1 and S2 were attributed to the increases in the SDS-isoluble and SDS-soluble protein extracts, respectively. S1 improved polymer protein formation, but S2 delayed the conversion of albumins and globulins into monomeric and polymeric proteins. Moreover, shade stress increased the proportion of SDS-unextractable polymeric protein, which represented an increase in the degree of protein polymerization. The polymerization of protein interrelations between protein components and accumulation in caryopsis and endosperm provided novel insights into wheat quality formation under shade stress.

Dysfunction of astrocytic glycophagy exacerbates reperfusion injury in ischemic stroke.

Glycophagy has evolved from an alternative glycogen degradation pathway into a multifaceted pivot to regulate cellular metabolic hemostasis in peripheral tissues. However, the pattern of glycophagy in the brain and its potential therapeutic impact on ischemic stroke remain unknown. Here, we observed that the dysfunction of astrocytic glycophagy was caused by the downregulation of the GABA type A receptor-associated protein like 1 (GABARAPL1) during reperfusion in ischemic stroke patients and mice. PI3K-Akt pathway activation is involved in driving GABARAPL1 downregulation during cerebral reperfusion. Moreover, glycophagy dysfunction-induced glucosamine deficiency suppresses the nuclear translocation of specificity protein 1 and TATA binding protein, the transcription factors for GABARAPL1, by decreasing their O-GlcNAcylation levels, and accordingly feedback inhibits GABARAPL1 in astrocytes during reperfusion. Restoring astrocytic glycophagy by overexpressing GABARAPL1 decreases DNA damage and oxidative injury in astrocytes and improves the survival of surrounding neurons during reperfusion. In addition, a hypocaloric diet in the acute phase after cerebral reperfusion can enhance astrocytic glycophagic flux and accelerate neurological recovery. In summary, glycophagy in the brain links autophagy, metabolism, and epigenetics together, and glycophagy dysfunction exacerbates reperfusion injury after ischemic stroke.

Spatial resolved transcriptomics reveals distinct cross-talk between cancer cells and tumor-associated macrophages in intrahepatic cholangiocarcinoma.

BACKGROUND: Multiple studies have shown that tumor-associated macrophages (TAMs) promote cancer initiation and progression. However, the reprogramming of macrophages in the tumor microenvironment (TME) and the cross-talk between TAMs and malignant subclones in intrahepatic cholangiocarcinoma (iCCA) has not been fully characterized, especially in a spatially resolved manner. Deciphering the spatial architecture of variable tissue cellular components in iCCA could contribute to the positional context of gene expression containing information pathological changes and cellular variability. METHODS: Here, we applied spatial transcriptomics (ST) and digital spatial profiler (DSP) technologies with tumor sections from patients with iCCA. RESULTS: The results reveal that spatial inter- and intra-tumor heterogeneities feature iCCA malignancy, and tumor subclones are mainly driven by physical proximity. Tumor cells with TME components shaped the intra-sectional heterogenetic spatial architecture. Macrophages are the most infiltrated TME component in iCCA. The protein trefoil factor 3 (TFF3) secreted by the malignant subclone can induce macrophages to reprogram to a tumor-promoting state, which in turn contributes to an immune-suppressive environment and boosts tumor progression. CONCLUSIONS: In conclusion, our description of the iCCA ecosystem in a spatially resolved manner provides novel insights into the spatial features and the immune suppressive landscapes of TME for iCCA.

Flexible nano-liposomes-encapsulated recombinant UL8-siRNA (r/si-UL8) based on bioengineering strategy inhibits herpes simplex virus-1 infection.

Herpes simplex virus-1 (HSV-1) infection can cause various diseases and the current therapeutics have limited efficacy. Small interfering RNA (siRNA) therapeutics are a promising approach against infectious diseases by targeting the viral mRNAs directly. Recently, we employed a novel tRNA scaffold to produce recombinant siRNA agents with few natural posttranscriptional modifications. In this study, we aimed to develop a specific prodrug against HSV-1 infection based on siRNA therapeutics by bioengineering technology. We screened and found that UL8 of the HSV-1 genome was an ideal antiviral target based on RNAi. Next, we used a novel bio-engineering approach to manufacture recombinant UL8-siRNA (r/si-UL8) in Escherichia coli with high purity and activity. The r/si-UL8 was selectively processed to mature si-UL8 and significantly reduced the number of infectious virions in human cells. r/si-UL8 delivered by flexible nano-liposomes significantly decreased the viral load in the skin and improved the survival rate in the preventive mouse zosteriform model. Furthermore, r/si-UL8 also effectively inhibited HSV-1 infection in a 3D human epidermal skin model. Taken together, our results highlight that the novel siRNA bioengineering technology is a unique addition to the conventional approach for siRNA therapeutics and r/si-UL8 may be a promising prodrug for curing HSV-1 infection.

Disparate macrophage responses are linked to infection outcome of Hantan virus in humans or rodents.

Hantaan virus (HTNV) is asymptomatically carried by rodents, yet causes lethal hemorrhagic fever with renal syndrome in humans, the underlying mechanisms of which remain to be elucidated. Here, we show that differential macrophage responses may determine disparate infection outcomes. In mice, late-phase inactivation of inflammatory macrophage prevents cytokine storm syndrome that usually occurs in HTNV-infected patients. This is attained by elaborate crosstalk between Notch and NF-κB pathways. Mechanistically, Notch receptors activated by HTNV enhance NF-κB signaling by recruiting IKKß and p65, promoting inflammatory macrophage polarization in both species. However, in mice rather than humans, Notch-mediated inflammation is timely restrained by a series of murine-specific long noncoding RNAs transcribed by the Notch pathway in a negative feedback manner. Among them, the lnc-ip65 detaches p65 from the Notch receptor and inhibits p65 phosphorylation, rewiring macrophages from the pro-inflammation to the pro-resolution phenotype. Genetic ablation of lnc-ip65 leads to destructive HTNV infection in mice. Thus, our findings reveal an immune-braking function of murine noncoding RNAs, offering a special therapeutic strategy for HTNV infection.

Comparative persistence of subgroups of F-specific RNA phages in river water.

F-specific (F+) RNA phages are widely used as indicators for the presence of fecal contamination and/or enteric viruses in water, and identifying subgroups of F+ RNA phages provides an approach for microbial source tracking. Different survival characteristics of the F+ RNA phage subgroups result in a misinterpretation of their original proportion in water, thus giving misleading information when they are used for microbial source tracking. This study investigated the comparative persistence of subgroups of F+ RNA phages in river water under different conditions. Results suggested that temperature and pH are the major factors affecting the persistence of F+ RNA phages in river water, and organic substances promote phage survival. The comparative persistence patterns of subgroups of F+ RNA phages varied and may bias extrapolation of their initial proportions in surface water. Thus, the characteristics of water should be taken into consideration and the results should be carefully interpreted when F+ RNA phages are used for microbial source tracking.

An overview of extrahepatic cholangiocarcinoma: from here to where?

Extrahepatic cholangiocarcinoma (eCCA) contains perihilar cholangiocarcinoma and distal cholangiocarcinoma both of which can arise at any point of the biliary tree and originate from disparate anatomical sites. Generally, the incidence of eCCA is increasing globally. Though surgical resection is the principal treatment of choice for the early stages of eCCA, optimal survival remains restricted by the high risk of recurrence when most patients are present with unresectable disease or distant metastasis. Furthermore, both intra- and intertumoral heterogeneity make it laborious to determine molecularly targeted therapies. In this review, we mainly focused on current findings in the field of eCCA, mostly including epidemiology, genomic abnormalities, molecular pathogenesis, tumor microenvironment, and other details while a summary of the biological mechanisms driving eCCA may shed light on intricate tumorigenesis and feasible treatment strategies.

Screening genes related to embryo implantation in Dazu black goats (Capra Hircus) by morphological and transcriptome analyses.

Embryo implantation is a critical step in the establishment of pregnancy. However, the mechanisms of embryo implantation during early pregnancy in goats remain unclear due to the lack of published studies examining the genes involved in embryo implantation. As a popular goat breed in southwest China, Dazu black goats (DBGs) are highly adaptable and exhibit high fertility, making this breed a good model in which to study reproductive performance of goats. Here, morphological analysis showed that compared with the non-pregnant (NP) groups, the endometrial thickness of the goats in the P15 and P19 groups (15 and 19-day pregnant groups, respectively) were increased (P < 0.01). Proliferating Cell Nuclear Antigen (PCNA) staining showed that PCNA was expressed in the NP, P15, and P19 groups. Transcriptome analysis was then conducted to identify gene expression patterns in uterine tissue during DBG embryo implantation. By comparing uterine tissue at different stages of embryonic implantation, 48 in NP_vs._P15, 318 in NP_vs._P19, and 1439 in P15_vs._P19, differentially expressed mRNAs were identified. Gene Ontology (GO) enrichments of the differentially expressed genes were enriched in the extracellular region, extracellular space, transporter activity, extracellular region, immune system process, immune response, and defense response etc. Through Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis, the biological metabolic pathways with which the differentially expressed genes are associated were explored. Through KEGG analysis, the DBGs were associated with oxidative phosphorylation, complement and coagulation cascades, arginine and proline metabolism, metabolic pathways, arachidonic acid metabolism, and ECM-receptor interaction. These candidate genes (CSF1, C1S, CST6, SLC24A4, HOXA10, HOXA11, MMP9, and ITGA11) and enriched signaling pathways could be valuable references for exploring the molecular mechanisms underlying goat embryo implantation.

Mammalian embryo implantation refers to the process that the embryo normally develops to the blastocyst stage, contacts the maternal endometrium, and establishes one kind structural connection. This intimate connection allows for the process of maternal­fetal material exchange, which is one of the key steps in the successful pregnancy. The success of embryo implantation depends on two aspects of the endometrium and the embryo, 1) the maternal endometrium is in a receptive state, and 2) the embryo develops normally, both of which are indispensable. In this stage, the mechanism of embryo implantation early in goat pregnancy is not clear, as only few limited studies have been conducted into gene expression in the uterus during embryo implantation. In this study, goat uterine tissue was systematically collected during the periods of non-pregnancy, pregnancy recognition, and embryo adhesion. And the morphological changes of the uterus in the different gestational stages were also observed, and gene expression associated with embryo implantation was further analyzed by RNA-seq method. This study provides a preliminary dataset for analyzing the molecular mechanisms regulating goat embryo implantation.