ARID1A loss promotes RNA editing of CDK13 in an ADAR1-dependent manner.

BACKGROUND: ARID1A, a subunit of the SWI/SNF chromatin remodeling complex, is thought to play a significant role both in tumor suppression and tumor initiation, which is highly dependent upon context. Previous studies have suggested that ARID1A deficiency may contribute to cancer development. The specific mechanisms of whether ARID1A loss affects tumorigenesis by RNA editing remain unclear. RESULTS: Our findings indicate that the deficiency of ARID1A leads to an increase in RNA editing levels and alterations in RNA editing categories mediated by adenosine deaminases acting on RNA 1 (ADAR1). ADAR1 edits the CDK13 gene at two previously unidentified sites, namely Q113R and K117R. Given the crucial role of CDK13 as a cyclin-dependent kinase, we further observed that ADAR1 deficiency results in changes in the cell cycle. Importantly, the sensitivity of ARID1A-deficient tumor cells to SR-4835, a CDK12/CDK13 inhibitor, suggests a promising therapeutic approach for individuals with ARID1A-mutant tumors. Knockdown of ADAR1 restored the sensitivity of ARID1A deficient cells to SR-4835 treatment. CONCLUSIONS: ARID1A deficiency promotes RNA editing of CDK13 by regulating ADAR1.

Cellular heterogeneity and immune microenvironment revealed by single-cell transcriptome in venous malformation and cavernous venous malformation.

Venous malformation (VM) and cavernous venous malformation (CVM) are two types of vascular malformations. Even if the two diseases are similar in appearance and imaging, the distinct cellular components and signaling pathways between them might help distinguish the two from a molecular perspective. Here, we performed single-cell profiling of 35,245 cells from two VM samples and three CVM samples, with a focus on endothelial cells (ECs), smooth muscle cells (SMCs) and immune microenvironment (IME). Clustering analysis based on differential gene expression unveiled 11 specific cell types, and determined CVM had more SMCs. Re-clustering of ECs and SMCs indicated CVM was dominated by arterial components, while VM is dominated by venous components. Gene set variation analysis suggested the activation of inflammation-related pathways in VM ECs, and upregulation of myogenesis pathway in CVM SMCs. In IME analysis, immune cells were identified to accounted for nearly 30% of the total cell number, including macrophages, monocytes, NK cells, T cells and B cells. Notably, more macrophages and monocytes were discovered in VM, indicating innate immune responses might be more closely related to VM pathogenesis. In addition, angiogenesis pathway was highlighted among the significant pathways of macrophages & monocytes between CVM and VM. In VM, VEGFA was highly expressed in macrophages & monocytes, while its receptors were all abundantly present in ECs. The close interaction of VEGFA on macrophages with its receptors on ECs was also predicted by CellPhoneDB analysis. Our results document cellular composition, significant pathways, and critical IME in CVM and VM development.

Proteomic analysis reveals the heterogeneity of metabolic reprogramming in lacrimal gland tumors.

Lacrimal gland adenoid cystic carcinoma (ACC) is associated with high recurrence and mortality rates. Many recent studies have focused on the clinical features of the disease, and a better understanding of its underlying molecular mechanisms may help guide future treatment strategies. For proteomics quantitation, we analyzed normal tissues, benign tumor tissues and ACC tissues by LC-MS/MS with Tandem mass tags (TMTs) labeling. Bioinformatics analysis of the KEGG pathway found that, compared with normal tissues, the expression levels of major proteins related to cell metabolism were lower in benign tumors and cancer tissues of the lacrimal gland. In addition, we also performed IHC staining to verify the expression of representative proteins in tissue samples. All of these results indicated that compared with normal tissues, lacrimal gland tumors had unique metabolic reprogramming characteristics. Further Short Time-series Expression Miner (STEM) analysis revealed that glycine, serine and threonine metabolism in ACC tissues was significantly enhanced compared with that in normal tissues and benign tumor tissues. This finding suggested that glycine, serine and threonine metabolism might be the key to the malignant transformation of ACC; thus, assessing the metabolism in these tissues could be an effective approach enabling the early diagnosis of ACC, and the proteins involved in these metabolic pathways could represent therapeutic targets.

Clinical characteristics and germline mutation spectrum of RB1 in Chinese patients with retinoblastoma: A dual-center study of 145 patients.

Retinoblastoma (Rb) is the most common primary intraocular childhood malignancy and one of the main causes of blindness in children. In China, most tumors are diagnosed at an advanced stage and have relatively poor outcomes compared to developed countries. Here, we aimed to update the clinical manifestations and RB transcriptional corepressor 1 (RB1) mutation spectrum in Chinese Rb patients. Medical charts of 184 eyes in 145 Chinese Rb patients belonging to unrelated families were reviewed. Genomic DNA was isolated from peripheral blood of the patients and their parents. Mutation analysis of whole coding regions, promoter regions and flanking splice sites in the RB1 gene was performed. In addition, multiplex ligation-dependent probe amplification (MLPA) was done to detect gross aberrations. Germline RB1 mutations were observed in 37.2% (54/145) of Rb patients. RB1-mutated patients presented with earlier age of diagnosis (p = 0.019), with a significantly larger proportion of bilateral cases (p = <0.001) and secondary malignancies (p = 0.027) relative to those without RB1 mutations. For ocular clinical presentations, RB1-mutated retinoblastomas presented with a larger proportion of ectropion uveae (p = 0.017) and iris neovascularization (p = 0.001). These RB1 mutations comprised of 13 (24.1%) nonsense mutation, 13 (24.1%) splicing mutations, 11 (20.4%) frameshift deletions, 11 (20.4%) gross mutations, 3 (5.6%) missense mutations, 2 (3.7%) promoter mutations and 1 (1.9%) non-frameshift deletion. In addition, 8 novel RB1 mutations were identified. These germline RB1 mutations were not related to age at diagnosis or laterality. Here, we provide a comprehensive spectrum of RB1 germline mutations in Chinese Rb patients and describe correlations between RB1 mutations and clinical presentations. Our study also provides new evidence that will inform management and genetic counselling of Rb patients and families.

BMP9 attenuates occurrence of venous malformation by maintaining endothelial quiescence and strengthening vessel walls via SMAD1/5/ID1/α-SMA pathway.

Venous malformation (VM) is a type of vascular morphogenic defect in humans with an incidence of 1%. Although gene mutation is considered as the most common cause of VM, the pathogenesis of those without gene mutation remains to be elucidated. Here, we aimed to explore the relation of bone morphogenetic protein 9 (BMP9) and development of VM. At first, we found serum and tissue BMP9 expression in VM patients was significantly lower than that in healthy subjects, detected via enzyme-linked immunosorbent assay. Next, with wound healing assay, transwell assay and tube formation assay, we discovered BMP9 could inhibit migration and enhance tube formation activity of human umbilical vein endothelial cells (HUVECs) via receptor activin receptor-like kinase 1 (ALK1). Besides, BMP9 improved the expression of structural proteins alpha-smooth muscle actin (α-SMA) and Desmin in human umbilical vein smooth muscle cells (HUVSMCs) via activation of the SMAD1/5-ID1 pathway, determined by RNA-based next-generation sequencing, qPCR, immunofluorescence and western blotting. Intriguingly, this effect could be blocked by receptor ALK1 inhibitor, SMAD1/5 inhibitor and siRNAs targeting ID1, verifying the BMP9/ALK1/SMAD1/5/ID1/α-SMA pathway. Meanwhile, knocking out BMP9 in C57BL/6 mice embryo led to α-SMA scarcity in walls of lung and mesenteric vessels, as well as walls of small trachea. BMP9-/- zebrafish also exhibited abnormal vascular maturity, indicating a critical role of BMP9 in vascular maturity and remodeling. Finally, a VM mice model revealed that BMP9 might have therapeutic effect in VM progression. Our study discovered that BMP9 might inhibit the occurrence of VM by strengthening the vessel wall and maintaining endothelium quiescence. These findings provide promising evidences of new therapeutic targets that might be used for the management of VM.

A novel LncRNA transcript, RBAT1, accelerates tumorigenesis through interacting with HNRNPL and cis-activating E2F3.

BACKGROUND: Long non-coding RNAs (lncRNAs) have been identified as important epigenetic regulators that play critical roles in human cancers. However, the regulatory functions of lncRNAs in tumorigenesis remains to be elucidated. Here, we aimed to investigate the molecular mechanisms and potential clinical application of a novel lncRNA, retinoblastoma associated transcript-1 (RBAT1), in tumorigenesis. METHODS: RBAT1 expression was determined by real-time PCR in both retinoblastoma (Rb) and bladder cancer (BCa) cell lines and clinical tissues. Chromatin isolation using RNA purification (ChIRP) assays were performed to identify RBAT1-interacting proteins. Patient-derived xenograft (PDX) retinoblastoma models were established to test the therapeutic potential of RBAT1-targeting GapmeRs. RESULTS: Here, we found that RBAT1 expression was significantly higher in Rb and BCa tissues than that in adjacent tissues. Functional assays revealed that RBAT1 accelerated tumorigenesis both in vitro and in vivo. Mechanistically, RBAT1 recruited HNRNPL protein to E2F3 promoter, thereby activating E2F3 transcription. Therapeutically, GapmeR-mediated RBAT1 silencing significantly inhibited tumorigenesis in orthotopic xenograft retinoblastoma models derived from Rb cell lines and Rb primary cells. CONCLUSIONS: RBAT1 overexpression upregulates a known oncogene, E2F3, via directly recruiting HNPNPL to its promoter and cis-activating its expression. Our finding provides a novel mechanism of lncRNA biology and provides potential targets for diagnosis and treatment of Rb and BCa.

The fidelity of cancer cells in PDX models: Characteristics, mechanism and clinical significance.

Patient-derived xenograft (PDX) models are widely used as preclinical cancer models and are considered better than cell culture models in recapitulating the histological features, molecular characteristics and intratumoral heterogeneity (ITH) of human tumors. While the PDX model is commonly accepted for use in drug discovery and other translational studies, a growing body of evidence has suggested its limitations. Recently, the fidelity of cancer cells within a PDX has been questioned, which may impede the future application of these models. In this review, we will focus the variable phenotypes of xenograft tumors and the genomic instability and molecular inconsistency of PDX tumors after serial transplantation. Next, we will discuss the underlying mechanism of ITH and its clinical relevance. Stochastic selection bias in the sampling process and/or deterministic clonal dynamics due to murine selective pressure may have detrimental effects on the results of personalized medicine and drug screening studies. In addition, we aim to identify a possible solution for the issue of fidelity in current PDX models and to discuss emerging next-generation preclinical models.

A novel variant in GPAA1, encoding a GPI transamidase complex protein, causes inherited vascular anomalies with various phenotypes.

Vascular anomalies (VAs), comprising wide subtypes of tumors and malformations, are often caused by variants in multiple tyrosine kinase (TK) receptor signaling pathways including TIE2, PIK3CA and GNAQ/11. Yet, a portion of individuals with clinical features of VA do not have variants in these genes, suggesting that there are undiscovered pathogenic factors underlying these patients and possibly with overlapping phenotypes. Here, we identified one rare non-synonymous variant (c.968A > G) in the seventh exon of GPAA1 (Glycosylphosphatidylinositol Anchor Attachment Protein 1), shared by the four affected members of a large pedigree with multiple types of VA using whole-exome sequencing. GPAA1 encodes a glycosylphosphatidylinositol (GPI) transamidase complex protein. This complex orchestrates the attachment of the GPI anchor to the C terminus of precursor proteins in the endoplasmic reticulum (ER). We showed such variant led to scarce expression of GPAA1 protein in vascular endothelium and induced a localization change from ER membrane to cytoplasm and nucleus. In addition, expressing wild-type GPAA1 in endothelial cells had an effect to inhibit cell proliferation and migration, while expressing variant GPAA1 led to overgrowth and overmigration, indicating a loss of the quiescent status. Finally, a gpaa1-deficient zebrafish model displayed several types of developmental defects as well as vascular dysplasia, demonstrating that GPAA1 is involved in angiogenesis and vascular remodeling. Altogether, our results indicate that the rare coding variant in GPAA1 (c.968A > G) is causally related to familial forms of VAs.

High-throughput transcriptional profiling combined with angiogenesis antibody array analysis in an orbital venous malformation cohort.

Orbital venous malformations (OVMs) are the most common benign orbital vascular disorders in adults and are characterized as enlarging encapsulated vascular neoplasms. These painless lesions grow slowly and become symptomatic with proptosis or visual disturbance. However, the pathogenic mechanism and diagnostic markers of OVMs remain poorly understood. To identify potential pathways involved in OVM formation, a cDNA microarray analysis was conducted with OVM samples and normal vascular tissues. These data were deposited in the National Omics Data Encyclopedia (NODE) database (accession number: OER033009). These pathway expression data were further confirmed by reverse transcription qPCR (RT-qPCR) in an OVM cohort. To explore the diagnostic markers in OVM, an angiogenesis antibody array was analyzed. The altered factors were further validated by enzyme-linked immunosorbent assay (ELISA) in the OVM cohort. Transcriptome screening revealed upregulated autophagy and VEGF pathways and downregulated Hippo, Wnt, hedgehog and vascular smooth muscle contraction signaling pathways in OVM samples. Furthermore, plasma EGF (p < 0.001) and Leptin (p < 0.01) levels were significantly elevated in OVM patients. Here, for the first time, we revealed the transcriptional background and plasma diagnostic markers in OVM, providing a novel understanding of OVM pathogenesis and facilitating the early diagnosis of OVM.

The oncolytic virus H101 combined with GNAQ siRNA-mediated knockdown reduces uveal melanoma cell viability.

BACKGROUND: Uveal melanoma (UM) is a severe human malignancy with a high mortality rate, as well as high metastasis and recurrence potential. The active mutation of G protein subunit alpha q (GNAQ) or G protein subunit alpha 11 (GNA11) is a major trigger for UM. Oncolytic adenovirus H101 (H101) is the first oncolytic virus approved for clinical applications in cancer therapy by the China Food and Drug Administration. We investigated whether combining H101 with the downregulation of GNAQ expression would act synergistically in UM therapy. METHODS: Three UM cell lines OMM2.3 and 92.1, harboring GNAQ mutation, and OCM1, harboring B-Raf proto-oncogene mutation, were chosen for our research. The cellular toxicity of adenoviral infection and the cell growth rate were measured with a Cell Counting Kit-8. Western blot analysis was used to detect GNAQ, p-MEK1/2, YAP, and p-YAP expression. The apoptosis and cell-cycle distribution of cells were evaluated with annexin-V and propidium iodide staining. RESULTS: Our results revealed that OMM2.3 and 92.1 cells were more sensitive to H101 infection than OCM1 cells. GNAQ expression was markedly reduced by small interfering RNA, siGNAQ. Combined treatment of siGNAQ and H101 inhibited the proliferation and activated the apoptosis of OMM2.3 and 92.1 cells by blocking the phosphorylation of MEK1/2 and increasing the phosphorylation of YAP. CONCLUSIONS: In summary, a therapy combining H101 and siGNAQ is feasible, with potential utility as a novel targeted molecular therapy for UM, especially those carrying a GNAQ mutation.

Targeting secreted cytokine BMP9 gates the attenuation of hepatic fibrosis.

Liver fibrosis is overly exuberant wound healing that leads to portal hypertension or liver cirrhosis. Recent studies have demonstrated the functions of bone morphogenetic protein 9 (BMP9) in liver fibrosis, and thus, targeting liver-specific BMP9 abnormalities will become an attractive approach for developing therapeutics to treat liver fibrosis. Here, we reveal that BMP9 serves as a valuable serum diagnostic indicator and efficient therapeutic target to attenuate liver fibrogenesis. Our analysis of biopsies from liver fibrotic patients revealed that higher BMP9 levels accompanied advanced stages of liver fibrosis. In mouse models, recombinant Bmp9 overexpression accelerated liver fibrosis, and adenovirus-mediated Bmp9 knockdown attenuated liver fibrogenesis. Intriguingly, BMP9 directly stimulated hepatic stellate cell activation via the SMAD signaling pathway to enhance hepatic fibrosis. Moreover, an inhibitory monoclonal antibody targeting Bmp9 was efficacious in treatment of mice with liver fibrosis. These observations delineate a novel model in which BMP9 directly drives SMAD/ID1 signaling in hepatic stellate cells, which modulates liver fibrogenesis development. Moreover, the findings unveil a promising surrogate biomarker for the diagnosis of hepatic fibrosis, thereby representing an efficient "BMP9 neutralization" approach in alleviating hepatic fibrosis.

Characterization of a conjunctival melanoma cell line CM-AS16, newly-established from a metastatic Han Chinese patient.

Conjunctival melanoma (CM) is associated with metastases formation, can be fatal, and occurs in all different races. While cell lines are essential for experimental research, all available CM cell lines are derived from Caucasian patients. Furthermore, they are not derived from metastases. We aimed to establish a new CM cell line from a parotid metastasis in a Han Chinese patient and to depict its characteristics. The novel cell line, CM-AS16, was obtained from a surgical parotid sample and determined as a unique one with short tandem repeat (STR) analysis. It has been successively sub-cultured in vitro for more than 100 passages and exhibits rapid proliferation and migration. Chromosome analysis shows abundant chromosome aberrations, while whole exome sequencing (WES) reveals a typical NRAS mutation (Q61R). In vivo tumor growth was successfully established in a NOD/SCID mice model, and the immunophenotypes, such as HMB45, Melan A, S100, SOX10 and Ki67, manifested similar between the original tumor and the xenograft by immunohistochemistry. A MEK inhibitor binimetinib prominently suppressed in vitro cell growth by inhibiting ERK1/2 phosphorylation. In addition, monoclonal cells were used to demonstrate the drug sensitivity of different cells. In conclusion, the first cell line, CM-AS16, that is derived from a CM in a Han Chinese patient has highly malignant characteristics and a typical NRAS mutation. It may be used as a tool for further exploration of the molecular mechanisms of CM.

Enhanced Cd activation by Coprinus comatus endophyte Bacillus thuringiensis and the molecular mechanism.

Fungal endophytes not only tolerate and activate Cd in soil but also promote host growth, yet its Cd activation capacity and mechanism remain unrevealed. Our previous study isolated a robust endophyte Bacillus thuringiensis L1 from Coprinus comatus fruiting body with splendid Cd resistance and activation abilities under laboratory conditions. In this study, those peculiarities were investigated in the actual soil environment. L1 could significantly increase the soil bioavailable Cd content and effectively compensate for alkali-hydro nitrogen losses and microbial inhibition caused by Cd. Furthermore, L1 inoculation improved the soil's bacterial community structure and increased the relative abundance of Cd-resistant bacteria, such as Actinobacteria, Chloroflexi, Acidobacter, and Firmicutes, closely associated with the soil enzyme activity shift. The genome sequencing analysis revealed the presence of genes related to growth promotion, resistance to Cd stress, and Cd activation, which were significantly up-regulated under Cd stress. Notably, L1 mainly activates Cd in soil by secreting citric acid, succinic acid, siderophore, and soluble phosphorus substances to chelate with Cd or dissolve bounded Cd. Meanwhile, the metal-responsive transcription repressor (CadC) and the Cd-translocating protein P-type ATPase (CadA) can help the L1 to suppress the toxicity of Cd. Those results help to unveil the possible mechanism of L1 in Cd-contaminated soil remediation, providing a clear strategy for Cd bio-extraction from soil.

Different survival strategies of the phosphate-mineralizing bacterium Enterobacter sp. PMB-5 in response to cadmium stress: Biomineralization, biosorption, and bioaccumulation.

The phosphate-mineralizing bacteria (PMBs) has shown great potential as a sustainable solution to support pollution remediation through its induced mineralization capacity. However, few studies have been conducted on the mechanism of cadmium (Cd) tolerance in PMBs. In this study, a PMB strain, Enterobacter sp. PMB-5, screened from Cd-contaminated rhizosphere soil, has high resistance to Cd (540 - 1220 mg/L) and solubilized phosphate (232.08 mg/L). The removal experiments showed that the strain PMB-5 removed 71.69-98.24% and 34.83-76.36% of Cd with and without biomineralization, respectively. The characterization result of SEM, EDS, TEM, XPS and XRD revealed that PMB-5 induced Cd to form amorphous phosphate precipitation through biomineralization and adopted different survival strategies, including biomineralization, bioaccumulation, and biosorption to resistance Cd in the microbial induced phosphate precipitation (MIPP) system and the non-MIPP system, respectively. Moreover, the results of whole genome sequencing and qRT-PCR indicated that phosphorus metabolism genes such as pst, pit, phn, ugp, ppk, etc. and heavy metal tolerance genes (including ion transport, ion efflux, redox, antioxidant stress), such as czcD, zntA, mgtA, mgtC, katE, SOD2, dsbA, cysM, etc. were molecular for the PMB-5 mineralization and Cd tolerance of PMB-5. Together, our findings suggested Enterobacter sp. PMB-5 is a potential target for developing more effective bioinoculants for Cd contamination remediation.

EHMT2 promotes tumorigenesis in GNAQ/11-mutant uveal melanoma via ARHGAP29-mediated RhoA pathway.

Constitutive activation of GNAQ/11 is the initiative oncogenic event in uveal melanoma (UM). Direct targeting GNAQ/11 has yet to be proven feasible as they are vital for a plethora of cellular functions. In search of genetic vulnerability for UM, we found that inhibition of euchromatic histone lysine methyltransferase 2 (EHMT2) expression or activity significantly reduced the proliferation and migration capacity of cancer cells. Notably, elevated expression of EHMT2 had been validated in UM samples. Furthermore, Kaplan-Meier survival analysis indicated high EHMT2 protein level was related to poor recurrence-free survival and a more advanced T stage. Chromatin immunoprecipitation sequencing analysis and the following mechanistic investigation showed that ARHGAP29 was a downstream target of EHMT2. Its transcription was suppressed by EHMT2 in a methyltransferase-dependent pattern in GNAQ/11-mutant UM cells, leading to elevated RhoA activity. Rescuing constitutively active RhoA in UM cells lacking EHMT2 restored oncogenic phenotypes. Simultaneously blocking EHMT2 and GNAQ/11 signaling in vitro and in vivo showed a synergistic effect on UM growth, suggesting the driver role of these two key molecules. In summary, our study shows evidence for an epigenetic program of EHMT2 regulation that influences UM progression and indicates inhibiting EHMT2 and MEK/ERK simultaneously as a therapeutic strategy in GNAQ/11-mutant UM.

High-risk histopathologic features in local advanced conjunctival melanoma.

PURPOSE: To identify high-risk histopathologic and molecular features of local recurrence, nodal metastasis, distant metastasis (DM) and disease-specific death (DSD) in conjunctival melanoma (CoM). METHODS: Ninety patients with pathologically diagnosed CoM between 2008 and 2023 were enrolled. Immunohistochemistry staining of BRAFV600E, NRASQ61R, CD117, PD-1 and PD-L1 was performed in 65 and 45 patients, respectively. Cox regression and Kaplan-Meier survival analysis were conducted to identify risk factors for local recurrence, nodal metastasis, DM and DSD. RESULTS: Pathologically, ulceration (hazard ratio [HR]: 3.170; 95% CI: 1.312-7.659; p = 0.01) and regression (HR: 3.196; 95% CI: 1.094-9.335; p = 0.034) were risk factors for DM. Tumour thickness ≥ 4 mm (HR: 4.889; 95% CI: 1.846-12.946; p = 0.001) and regression (HR: 4.011; 95% CI: 1.464-10.991; p = 0.007) were risk factors for DSD. For patients with tumour thickness < 4 mm, the presence of ulceration indicated a higher risk of nodal metastasis (log-rank p = 0.0011), DM (log-rank p = 0.00051) and DSD (log-rank p = 0.02). Patients with regression (+)/tumour-infiltrating lymphocytes (TILs) (+) had a higher risk for DM (log-rank p = 0.011) and DSD (log-rank p = 0.0032). Molecularly, the positive rate of BRAFV600E, NRASQ61R, CD117, PD-1 and PD-L1 was 40.00% (26/65), 43.08% (28/65), 70.77% (46/65), 46.67% (21/45) and 28.89% (13/45), respectively. Positive BRAFV600E was identified as an independent risk factor for DM (HR: 2.533; 95% CI: 1.046-6.136, p = 0.039). The expression level of BRAFV600E was positively correlated with vascular invasion (p = 0.01), as well as the expression levels of PD-1 (p = 0.038) and PD-L1 (p = 0.049). CONCLUSIONS: Tumour thickness ≥ 4 mm, ulceration, the coexistence of regression and TILs, and positive BRAFV600E were risk factors for poor prognosis of CoM patients. Besides, expression level of BRAFV600E was positively correlated with the expression levels of PD-1 and PD-L1.

Single-cell sequencing depicts tumor architecture and empowers clinical decision in metastatic conjunctival melanoma.

Conjunctival melanoma (CoM) is a potentially devastating tumor that can lead to distant metastasis. Despite various therapeutic strategies for distant metastatic CoM, the clinical outcomes remain unfavorable. Herein, we performed single-cell RNA sequencing (scRNA-seq) of 47,017 cells obtained from normal conjunctival samples (n = 3) and conjunctival melanomas (n = 7). Notably, we noticed a higher abundance of cancer-associated fibroblasts (CAFs) in tumor microenvironment (TME), correlated with enhanced angiogenic capacity and increased VEGFR expression in distal metastatic CoM. Additionally, we observed a significant decrease in the proportion of total CD8+ T cells and an increase in the proportion of naive CD8+ T cells, contributing to a relatively quiescent immunological environment in distal metastatic CoM. These findings were confirmed through the analyses of 70,303 single-cell transcriptomes of 7 individual CoM samples, as well as spatially resolved proteomes of an additional 10 samples of CoMs. Due to the increase of VEGFR-mediated angiogenesis and a less active T cell environment in distal metastatic CoMs, a clinical trial (ChiCTR2100045061) has been initiated to evaluate the efficacy of VEGFR blockade in combination with anti-PD1 therapy for patients with distant metastatic CoM, showing promising tumor-inhibitory effects. In conclusion, our study uncovered the landscape and heterogeneity of the TME during CoM tumorigenesis and progression, empowering clinical decisions in the management of distal metastatic CoM. To our knowledge, this is the initial exploration to translate scRNA-seq analysis to a clinical trial dealing with cancer, providing a novel concept by accommodating scRNA-seq data in cancer therapy.

Microbial response to antimony-arsenic distribution and geochemical factors at arable soil around an antimony mining site.

Antimony (Sb) mining often causes severe Sb pollution and associate arsenic (As) compound contamination. To further understand the interaction mechanism among soil microorganisms, heavy metal distribution, and geochemical factors, the effects of environmental factors on soil microbial communities under different levels of Sb-As co-contamination were studied in situ of Chashan antimony mine, Guangxi Province. The results showed that the range of Sb and As contents in soil were 1339.63-7762.28 mg/kg and 2170.3-10,371.36 mg/kg, respectively, and the residual fraction accounted for more than 98.0% with less than 2.0% of bioavailable fraction. Besides, the concentration of the two metals is both related to the distance to surface runoff. Different microbial communities in arable soils of each sample site were analyzed, which was significantly affected by soil environmental factors such as pH, ALN, AP, OM, Tot-Sb, Tot-As, Bio-As, and Bio-Sb. The phylum of Actinobacteria in sites 1, 4, and 5 was the most dominant and the phylum of Proteobacteria were the most dominant in sites 2 and 3. Moreover, the results of redundancy analysis (RDA), variation partition analysis (VPA), and Spearman correlation analyses demonstrated that microorganisms, heavy metal distribution, and geochemical factors interacted with each other and together shaped the microbial community. Our findings are beneficial for understanding the response of soil microorganisms to As-Sb distribution and geochemical factors in arable soils under Sb mining areas.

Enhanced cadmium phytoextraction efficiency of ryegrass (Lolium perenne L.) by porous media immobilized Enterobacter sp. TY-1.

Although studies on immobilized microorganisms have been conducted, their performance remains unclear for enhancing plants to remediate cadmium (Cd)-contaminated soil. In this study, a Cd-resistant strain TY-1 with good plant growth promotion traits was immobilized by biochar (BC) or oyster shell (OS) power to strengthen ryegrass to remediate Cd-contaminated soil. SEM-EDS combined with FTIR showed that TY-1 could tolerate Cd toxicity by surface precipitation, and functional groups such as hydroxyl and carbonyl groups might be involved. In the biocomposite treatments, soil pH increased, and the activity of fertility-related enzymes such as dehydrogenase increased by 109.01%-128.01%. The relative abundance of genus Saccharimonadales decreased from 7.97% to 3.35% in BS-TY and 2.61% in OS-TY, respectively. Thus, a suitable environment for ryegrass growth was created. The fresh weight, dry weight, plant height and Cd accumulation of ryegrass in TY treatment increased by 122.92%, 114.81%, 42.08% and 8.05%, respectively, compared to the control. Cd concentration in ryegrass was further increased in BC-TY and OS-TY by 24.14% and 40.23%, respectively. The improvement in soil microcosm and plant biomass forms an ongoing virtuous cycle, demonstrating that using carrier materials to improve the efficiency of microbial-assisted phytoremediation is realistic and feasible.

Application potential of Vaccinium ashei R. for cadmium migration retention in the mining area soil.

Despite numerous reports on phytoremediation of heavy metals contaminated soil, there are few reports on plant retention of heavy metals in the mining area slope. This study was the first of its kind to explore the cadmium (Cd) retention capacity of the blueberry (Vaccinium ashei Reade). Firstly, we investigated the stress response of blueberry to different soil Cd concentrations (1, 5, 10, 15, 20 mg/kg) to assess its potential for phytoremediation by pot experiments. The results showed that the blueberry biomass exposed to 10 and 15 mg/kg Cd was significantly increased compared with the control (1 mg/kg Cd); the blueberry crown increased by 0.40% and 0.34% in 10 and 15 mg/kg Cd-contaminated soil, respectively, compared with control; the blueberry heigh did not even change significantly in each treatment group; the total chlorophyll content, peroxidase and catalase activity of blueberry were enhanced in 5-20 mg/kg Cd treatments. Furthermore, the Cd contents of blueberry in the root, stem and leaf increased significantly as the Cd concentration of soil increased. We found that more Cd accumulated in blueberry root: the bioaccumulation concentration factor was root > stem > leaf for all groups; the residual-Cd (Cd speciation) in soil increased by 3.83%-411.11% in blueberry-planted versus unplanted groups; blueberry improved the Cd-contaminated soil micro-ecological environment including soil organic matter, available K and P, as well as microbial communities. Then, to investigate the effect of blueberry cultivation on Cd migration, we developed a bioretention model and revealed that soil Cd transport along the model slope was significantly weakened by blueberry cultivation, especially at the bottom of the model. In a word, this research suggests a promising method for the phytoremediation of Cd-contaminated soil and the reduction of Cd migration in mining areas.