Cancer stem cell-immune cell crosstalk in the tumor microenvironment for liver cancer progression.

Crosstalk between cancer cells and the immune microenvironment is determinant for liver cancer progression. A tumor subpopulation called liver cancer stem cells (CSCs) significantly accounts for the initiation, metastasis, therapeutic resistance, and recurrence of liver cancer. Emerging evidence demonstrates that the interaction between liver CSCs and immune cells plays a crucial role in shaping an immunosuppressive microenvironment and determining immunotherapy responses. This review sheds light on the bidirectional crosstalk between liver CSCs and immune cells for liver cancer progression, as well as the underlying molecular mechanisms after presenting an overview of liver CSCs characteristic and their microenvironment. Finally, we discuss the potential application of liver CSCs-targeted immunotherapy for liver cancer treatment.

Seed and Soil: Consensus Molecular Subgroups (CMS) and Tumor Microenvironment Features Between Primary Lesions and Metastases of Different Organ Sites in Colorectal Cancer.

Purpose: Consensus molecular subtypes (CMS) are mainly used for biological interpretability and clinical stratification of colorectal cancer (CRC) in primary tumors (PT) but few in metastases. The heterogeneity of CMS distribution in metastases and the concordance of CMS between PT and metastases still lack sufficient study. We used CMS to classify CRC metastases and combine it with histopathological analysis to explore differences between PT and distant metastases. Patients and Methods: We obtained gene expression profiles for 942 PT samples from TCGA database (n=376) and GEO database (n=566), as well as 442 metastasis samples from GEO database. Among these, 765 PT samples and 442 metastasis samples were confidently identified with CMS using the "CMS classifier" and enrolled for analysis. Clinicopathological manifestation and CMS classification of CRC metastases were assessed with data from GEO, TCGA, and cBioPortal. Overall, 105 PT-metastasis pairs were extracted from 10 GEO datasets to assess CMS concordance. Tumor microenvironment (TME) features between PT and metastases were analyzed by immune-stromal infiltration with ESTIMATE and xCell algorithms. Finally, TME features were validated with multiplex immunohistochemistry in 27 PT-metastasis pairs we retrospectively collected. Results: Up to 64% of CRC metastases exhibited concordant CMS groups with matched PT, and the TME of metastases was similar to that of PT. For most common distant metastases, liver metastases were predominantly CMS2 and lung and peritoneal metastases were mainly CMS4, highlighting "seed" of tumor cells of different CMS groups had a preference for metastasis to "soil" of specific organs. Compared with PT, cancer-associated fibroblasts (CAF) reduced in liver metastases, CD4+T cells and M2-like macrophages increased in lung metastases, and M2-like macrophages and CAF increased in peritoneal metastases. Conclusion: Our findings underscore the importance of CMS-guided specific organ monitoring and treatment post-primary tumor surgery for patients. Differences in immune-stromal infiltration among different metastases provide targeted therapeutic opportunities for metastatic CRC.

TLR2-dependent and independent pyroptosis in dTHP-1 cells induced by Actinomyces oris MG-1.

In the immune system, the detection of pathogens through various mechanisms triggers immune responses. Several types of specific programmed cell deaths play a role in the inflammatory reaction. This study emphasizes the inflammatory response induced by Actinomycetes. Actinomyces spp. are resident bacteria in human oral plaque and often serve as a bridge for pathogenic bacteria, which lack affinity to the tooth surface, aiding their colonization of the plaque. We aim to investigate the potential role of Actinomyces oris in the early stages of oral diseases from a new perspective. Actinomyces oris MG-1 (A. oris) was chosen for this research. Differentiated THP-1 (dTHP-1) cells were transiently treated with A. oris to model the inflammatory reaction. Cell viability, as well as relative gene and protein expression levels of dTHP-1 cells, were assessed using CCK-8, quantitative real-time polymerase chain reaction (RT-qPCR), enzyme-linked immunosorbent assay (ELISA), and Western blot assay. The treatment decreased cell viability and increased the expression of inflammatory genes such as IL-1R1 and NLRP3. It was also observed to significantly enhance the release of IL-1ß/IL-18 into the supernatant. Immunoblot analysis revealed a notable increase in the expression of N-gasdermin D persisting up to 24 h. Conversely, in models pre-treated with TLR2 inhibitors, N-gasdermin D was detectable only 12 h post-treatment and absent at 24 h. These results suggest that Actinomyces oris MG-1 induces pyroptosis in dTHP-1 cells via TLR2, but the process is not solely dependent on TLR2.

TET2-mediated tumor cGAS triggers endothelial STING activation to regulate vasculature remodeling and anti-tumor immunity in liver cancer.

Induction of tumor vascular normalization is a crucial measure to enhance immunotherapy efficacy. cGAS-STING pathway is vital for anti-tumor immunity, but its role in tumor vasculature is unclear. Herein, using preclinical liver cancer models in Cgas/Sting-deficient male mice, we report that the interdependence between tumor cGAS and host STING mediates vascular normalization and anti-tumor immune response. Mechanistically, TET2 mediated IL-2/STAT5A signaling epigenetically upregulates tumor cGAS expression and produces cGAMP. Subsequently, cGAMP is transported via LRRC8C channels to activate STING in endothelial cells, enhancing recruitment and transendothelial migration of lymphocytes. In vivo studies in male mice also reveal that administration of vitamin C, a promising anti-cancer agent, stimulates TET2 activity, induces tumor vascular normalization and enhances the efficacy of anti-PD-L1 therapy alone or in combination with IL-2. Our findings elucidate a crosstalk between tumor and vascular endothelial cells in the tumor immune microenvironment, providing strategies to enhance the efficacy of combinational immunotherapy for liver cancer.

One-Step Surface-to-Bulk Modification of High-Voltage and Long-Life LiCoO2 Cathode with Concentration Gradient Architecture.

Raising the charging cut-off voltage of layered oxide cathodes can improve their energy density. However, it inevitably introduces instabilities regarding both bulk structure and surface/interface. Herein, exploiting the unique characteristics of high-valence Nb5+ element, a synchronous surface-to-bulk-modified LiCoO2 featuring Li3 NbO4 surface coating layer, Nb-doped bulk, and the desired concentration gradient architecture through one-step calcination is achieved. Such a multifunctional structure facilitates the construction of high-quality cathode/electrolyte interface, enhances Li+ diffusion, and restrains lattice-O loss, Co migration, and associated layer-to-spinel phase distortion. Therefore, a stable operation of Nb-modified LiCoO2 half-cell is achieved at 4.6 V (90.9% capacity retention after 200 cycles). Long-life 250 Wh kg-1 and 4.7 V-class 550 Wh kg-1 pouch cells assembled with graphite and thin Li anodes are harvested (both beyond 87% after 1600 and 200 cycles). This multifunctional one-step modification strategy establishes a technological paradigm to pave the way for high-energy density and long-life lithium-ion cathode materials.

Inhibition of ACLY overcomes cancer immunotherapy resistance via polyunsaturated fatty acids peroxidation and cGAS-STING activation.

Adenosine 5'-triphosphate citrate lyase (ACLY) is a cytosolic enzyme that converts citrate into acetyl-coenzyme A for fatty acid and cholesterol biosynthesis. ACLY is up-regulated or activated in many cancers, and targeting ACLY by inhibitors holds promise as potential cancer therapy. However, the role of ACLY in cancer immunity regulation remains poorly understood. Here, we show that ACLY inhibition up-regulates PD-L1 immune checkpoint expression in cancer cells and induces T cell dysfunction to drive immunosuppression and compromise its antitumor effect in immunocompetent mice. Mechanistically, ACLY inhibition causes polyunsaturated fatty acid (PUFA) peroxidation and mitochondrial damage, which triggers mitochondrial DNA leakage to activate the cGAS-STING innate immune pathway. Pharmacological and genetic inhibition of ACLY overcomes cancer resistance to anti-PD-L1 therapy in a cGAS-dependent manner. Furthermore, dietary PUFA supplementation mirrors the enhanced efficacy of PD-L1 blockade by ACLY inhibition. These findings reveal an immunomodulatory role of ACLY and provide combinatorial strategies to overcome immunotherapy resistance in tumors.

Copper in hepatocellular carcinoma： A double-edged sword with therapeutic potentials.

Copper is a necessary cofactor vital for maintaining biological functions, as well as participating in the development of cancer. A plethora of studies have demonstrated that copper is a double-edged sword, presenting both benefits and detriments to tumors. The liver is a metabolically active organ, and an imbalance of copper homeostasis can result in deleterious consequences to the liver. Hepatocellular carcinoma (HCC), the most common primary liver cancer, is a highly aggressive malignancy with limited viable therapeutic options. As research advances, the focus has shifted towards the relationships between copper and HCC. Innovatively, cuproplasia and cuproptosis have been proposed to depict copper-related cellular growth and death, providing new insights for HCC treatment. By summarizing the constantly elucidated molecular connections, this review discusses the mechanisms of copper in the pathogenesis, progression, and potential therapeutics of HCC. Additionally, we aim to tentatively provide a theoretical foundation and gospel for HCC patients.

Simple transanal total mesorectal resection versus laparoscopic transabdominal total mesorectal resection for the treatment of low rectal cancer: a single-center retrospective case-control study.

Aim: To evaluate the efficacy and safety of simple TaTNE in the treatment of low rectal cancer compared with laparoscopic transabdominal TME. Methods: We collected patients with low rectal cancer admitted to our hospital between January 2019 and November 2021 who received simple TaTME or laparoscopic transabdominal TME. The main outcome was the integrity of the TME specimen. Secondary outcomes were the number of lymph nodes dissected, intraoperative blood loss, operative time, surgical conversion rate, Specimen resection length, circumferential margin (CRM), and distal resection margin (DRM), complication rate. In addition, the Wexner score and LARS score of fecal incontinence were performed in postoperative follow-up. Results: Pathological tissues were successfully resected in all patients. all circumferential margins of the specimen were negative. Specimen resection length was not statistically significant (9.94 ± 2.85 vs. 8.90 ± 2.49, P > 0.05). The incidence of postoperative complications in group A (n = 0) was significantly lower than that in group B (n = 3) (P > 0.05). There was no significant difference in operation time between group A and group B (296 ± 60.36 vs. 305 ± 58.28, P > 0.05). Among the patients with follow-up time less than 1 year, there was no significant difference in Wexner score and LARS score between group A and group B (P > 0.05). However, in patients who were followed up for more than 1 year, the Wexner score in group A (9.25 ± 2.73) was significantly lower than that in group B (17.36 ± 10.95) and was statistically significant (P < 0.05). Conclusion: For radical resection of low rectal cancer, Simple TaTME resection may be as safe and effective as laparoscopic transabdominal TME, and the long-term prognosis may be better.

Virtual biopsy using CT radiomics for evaluation of disagreement in pathology between endoscopic biopsy and postoperative specimens in patients with gastric cancer: a dual-energy CT generalizability study.

PURPOSE: To develop a noninvasive radiomics-based nomogram for identification of disagreement in pathology between endoscopic biopsy and postoperative specimens in gastric cancer (GC). MATERIALS AND METHODS: This observational study recruited 181 GC patients who underwent pre-treatment computed tomography (CT) and divided them into a training set (n = 112, single-energy CT, SECT), a test set (n = 29, single-energy CT, SECT) and a validation cohort (n = 40, dual-energy CT, DECT). Radiomics signatures (RS) based on five machine learning algorithms were constructed from the venous-phase CT images. AUC and DeLong test were used to evaluate and compare the performance of the RS. We assessed the dual-energy generalization ability of the best RS. An individualized nomogram combined the best RS and clinical variables was developed, and its discrimination, calibration, and clinical usefulness were determined. RESULTS: RS obtained with support vector machine (SVM) showed promising predictive capability with AUC of 0.91 and 0.83 in the training and test sets, respectively. The AUC of the best RS in the DECT validation cohort (AUC, 0.71) was significantly lower than that of the training set (Delong test, p = 0.035). The clinical-radiomic nomogram accurately predicted pathologic disagreement in the training and test sets, fitting well in the calibration curves. Decision curve analysis confirmed the clinical usefulness of the nomogram. CONCLUSION: CT-based radiomics nomogram showed potential as a clinical aid for predicting pathologic disagreement status between biopsy samples and resected specimens in GC. When practicability and stability are considered, the SECT-based radiomics model is not recommended for DECT generalization. CRITICAL RELEVANCE STATEMENT: Radiomics can identify disagreement in pathology between endoscopic biopsy and postoperative specimen.

Single-cell and spatial transcriptome analysis reveals the cellular heterogeneity of liver metastatic colorectal cancer.

In this study, we comprehensively charted the cellular landscape of colorectal cancer (CRC) and well-matched liver metastatic CRC using single-cell and spatial transcriptome RNA sequencing. We generated 41,892 CD45- nonimmune cells and 196,473 CD45+ immune cells from 27 samples of six CRC patients, and found that CD8_CXCL13 and CD4_CXCL13 subsets increased significantly in liver metastatic samples that exhibited high proliferation ability and tumor-activating characterization, contributing to better prognosis of patients. Distinct fibroblast profiles were observed in primary and liver metastatic tumors. F3+ fibroblasts enriched in primary tumors contributed to worse overall survival by expressing protumor factors. However, MCAM+ fibroblasts enriched in liver metastatic tumors might promote generation of CD8_CXCL13 cells through Notch signaling. In summary, we extensively analyzed the transcriptional differences of cell atlas between primary and liver metastatic tumors of CRC by single-cell and spatial transcriptome RNA sequencing, providing different dimensions of the development of liver metastasis in CRC.

WD repeat protein 54-mediator of ErbB2-driven cell motility 1 axis promotes bladder cancer tumorigenesis and metastasis and impairs chemosensitivity.

One of the most abundant protein-protein interaction domains in the human proteome is the WD40 repeat (WDR) domain. A Gene Expression Omnibus dataset revealed 37 differentially expressed WDR domain genes in bladder cancer (BC). WD repeat domain 54 (WDR54), an upregulated WDR domain gene, was selected for further investigation. Sixty pairs of frozen BC tumor and non-malignant bladder tissues and 83 paraffin-embedded BC tissue specimens were obtained. Loss-/gain-of-function experiments were carried out using BC and xenograft tumor models. WDR54 was overexpressed in BC cells, and its high expression was linked to tumor stage and lymph node metastases in patients. WDR54 contributed to the tumorigenesis and metastasis of BC and impaired its chemosensitivity. WDR54 prevented the degradation and ubiquitination of the mediator of ErbB2-driven cell motility 1 (MEMO1). WDR54 also promoted the interaction between MEMO1 and insulin receptor substrate 1 (IRS1) and activated the IRS1/AKT/ß-catenin pathway in BC cells. Particularly, WDR54 depended on MEMO1 to exert its biological functions. Our study demonstrated the relevance of WDR54 in BC and provides insight into the molecular mechanism underlying BC.

Improving the Oxygen Evolution Activity of Layered Double-Hydroxide via Erbium-Induced Electronic Engineering.

Layered double-hydroxide (LDH) has been considered an important class of electrocatalysts for the oxygen evolution reaction (OER), but the adsorption-desorption behaviors of oxygen intermediates on its surface still remain unsatisfactory. Apart from transition-metal doping to solve this electrocatalytic problem of LDH, rare-earth (RE) species have sprung up as emerging dopants owing to their unique 4f valence-electronic configurations. Herein, the Er is chosen as a RE model to improve OER activity of LDH via constructing nickel foam supported Er-doped NiFe-LDH catalyst (Er-NiFe-LDH@NF). The optimal Er-NiFe-LDH@NF exhibits a low overpotential (191 mV at 10 mA cm-2 ), high turnover frequency (0.588 s-1 ), and low activation energy (36.03 kJ mol-1 ), which are superior to Er-free sample. Electrochemical in situ Raman spectra reveal the facilitated transition of Ni-OH into Ni-OOH for promoted OER kinetics through the Er doping effect. Theoretical calculations demonstrate that the introduction of Er facilitates the spin crossover of valence electrons by optimizing the d band center of NiFe-LDH, which leads to the GO -GHO closer to the optimal activity of the kinetic OER volcano by balancing the bonding strength of *O and *OH. Moreover, the Er-NiFe-LDH@NF presents high practicability in electrochemical water-splitting devices with a low driving potential of and a well-extended driving period.

Plasma exosomal miR-122 regulates the efficacy of metformin via AMPK in type 2 diabetes and hepatocellular carcinoma.

Metformin is a drug that has been applied in clinical use for many years for the treatment of type 2 diabetes mellitus (T2DM). It achieves its function through multiple targets and modulation of multiple signaling pathways. To date, the mechanism of the action of metformin is still not fully understood. Along with glycemic control, metformin has shown good inhibitory effects on the development of many tumors. Here, we elucidated that plasma exosomal microRNA-122-5p (miR-122) is closely related to the mechanism of metformin. MiR-122 regulates glycogen-glucose metabolism in hepatocytes or hepatocellular carcinoma cells (HCC) by inhibiting the phosphorylation of AMPK. Since miR-122 and metformin regulate glucose metabolism homeostasis through similar mechanisms, miR-122 can antagonize the effects of metformin. MiR-122 expression increases the sensitivity of hepatocytes or HCC to metformin. Conversely, decreased expression of miR-122 results in hepatocyte insensitivity to metformin. Therefore, significantly elevated levels of miR-122 in plasma exosomes of hepatocellular carcinoma patients could enhance their sensitivity to metformin. The results of the present study revealed a key regulatory role of plasma exosomal miR-122 on the molecular mechanism of metformin. The regulation of key molecules of related signaling pathways by miR-122 may lead to similar glycemic lowering and tumor suppression therapeutic effects as metformin. This provides new ideas for the development of new therapeutic strategies for hepatocellular carcinoma based on the mechanism of miR-122 and metformin.

Diagnostic value of dual-energy CT and clinicopathological and imaging feature analysis of mixed endometrial stromal and smooth muscle tumors with intracardiac extension.

Objective: To describe the clinicopathological and imaging features of mixed endometrial stromal and smooth muscle tumors with intracardiac extension and to explore the diagnostic value of dual-energy computed tomography (DECT) for this rare entity. Materials and methods: This retrospective study analyzed the clinicopathological data and images of a 41-year-old female patient with pathologically documented mixed endometrial stromal and smooth muscle tumors with intracardiac extension who had undergone DECT examination. Seven virtual monoenergetic images (VMIs) in 10-kiloelectron volt (keV) intervals (range = 40-100 keV), iodine density (ID) maps, and Z effective (Zeff) maps were reconstructed, and lesion conspicuity was assessed. Tumor homology was analyzed using quantitative DECT parameters and energy spectrum attenuation curve. Results: The patient complained of a 10-day history of bilateral lower extremity edema. Computed tomography showed a hypoattenuating filling defect located within the paracervical vein that extended into the right atrium to the ventricle through the right iliac veins and inferior vena cava (IVC). Intracardiac and intravenous lesions mainly demonstrated moderate progressive enhancement, with localized non-enhancing necrotic areas on contrast-enhanced CT. Multiple nodules showing progressive enhancement (long-T1 signal, long-T2 signal) were observed at the fundus of the uterus on dynamic contrast-enhanced magnetic resonance imaging (MRI), which were deemed the primary lesions of the tumor. Overall, the tumor was characterized by a small primary lesion with extensive vascular extension. In addition, the 40 keV VMIs reconstructions were found to provide best visualization for the early detection of tumors. Conclusion: Although a definitive diagnosis of MESSMT with intracardiac extension requires confirmation by histopathological examination, imaging examination can be used to characterize the extent of the lesion. The dual-energy dataset facilitates tumor visualization and homology evaluation.

Molecular profiling of human non-small cell lung cancer by single-cell RNA-seq.

BACKGROUND: Lung cancer, one of the most common malignant tumors, exhibits high inter- and intra-tumor heterogeneity which contributes significantly to treatment resistance and failure. Single-cell RNA sequencing (scRNA-seq) has been widely used to dissect the cellular composition and characterize the molecular properties of cancer cells and their tumor microenvironment in lung cancer. However, the transcriptomic heterogeneity among various cancer cells in non-small cell lung cancer (NSCLC) warrants further illustration. METHODS: To comprehensively analyze the molecular heterogeneity of NSCLC, we performed high-precision single-cell RNA-seq analyses on 7364 individual cells from tumor tissues and matched normal tissues from 19 primary lung cancer patients and 1 pulmonary chondroid hamartoma patient. RESULTS: In 6 of 16 patients sequenced, we identified a significant proportion of cancer cells simultaneously expressing classical marker genes for two or even three histologic subtypes of NSCLC-adenocarcinoma (ADC), squamous cell carcinoma (SCC), and neuroendocrine tumor (NET) in the same individual cell, which we defined as mixed-lineage tumor cells; this was verified by both co-immunostaining and RNA in situ hybridization. These data suggest that mixed-lineage tumor cells are highly plastic with mixed features of different types of NSCLC. Both copy number variation (CNV) patterns and mitochondrial mutations clearly showed that the mixed-lineage and single-lineage tumor cells from the same patient had common tumor ancestors rather than different origins. Moreover, we revealed that patients with high mixed-lineage features of different cancer subtypes had worse survival than patients with low mixed-lineage features, indicating that mixed-lineage tumor features were associated with poorer prognosis. In addition, gene signatures specific to mixed-lineage tumor cells were identified, including AKR1B1. Gene knockdown and small molecule inhibition of AKR1B1 can significantly decrease cell proliferation and promote cell apoptosis, suggesting that AKR1B1 plays an important role in tumorigenesis and can serve as a candidate target for tumor therapy of NSCLC patients with mixed-lineage tumor features. CONCLUSIONS: In summary, our work provides novel insights into the tumor heterogeneity of NSCLC in terms of the identification of prevalent mixed-lineage subpopulations of cancer cells with combined signatures of SCC, ADC, and NET and offers clues for potential treatment strategies in these patients.

The Effect of Mesenchymal Stem Cells, Adipose Tissue Derived Stem Cells, and Cellular Stromal Vascular Fraction on the Repair of Acute Anal Sphincter Injury in Rats.

BACKGROUND: Anal sphincter incontinence (ASI) can cause a serious decline in the quality of life and can cause a socioeconomic burden. Studies have shown that bone marrow mesenchymal stem cells (MSC) have significant therapeutic effects on ASI, but the cost and risk of MSC harvest limit their further application. In contrast, adipose tissue derived stem cells (ADSC) and cellular stromal vascular fraction (CSVF) as stem cell sources have multipotency and the advantage of easy harvest. OBJECTIVE: Here we aim to investigate the effects of ADSC and CSVF on treating ASI and compare them to that of bone marrow MSC. METHODS: Bone marrow MSC, ADSC, and CSVF were obtained and labeled with green fluorescent protein (GFP), and CSVF was labeled with DIL. Sprague Dawley (SD) rats were divided into 5 groups. Four groups were injected with 0.2 mL phosphate buffer saline (PBS), 1 × 107/0.2 mL of MSC, ADSC, or CSVF, respectively, after model establishment. The control group received no treatment. The repair was assessed by anal functional tests and immunostaining on day 5 and day 10 after injection. RESULTS: MSC, ADSC, and CSVF significantly promoted tissue repair and the recovery of muscle contraction and electromyographic activity in ASI. The generation of myosatellite cells by injected MSC, ADSC, and CSVF was found in the wounded area. On day 5, CSVF showed highest therapeutic effect, while on day 10, MSC and ADSC showed higher therapeutic effects than CSVF. When comparing the effects of MSC and ADSC, ADSC was slightly better than MSC in the indexes of anal pressure, etc. Conclusion: ADSC and CVSF are alternative stem cell sources for ASI repair.

Similarity-Based Method with Multiple-Feature Sampling for Predicting Drug Side Effects.

Drugs can treat different diseases but also bring side effects. Undetected and unaccepted side effects for approved drugs can greatly harm the human body and bring huge risks for pharmaceutical companies. Traditional experimental methods used to determine the side effects have several drawbacks, such as low efficiency and high cost. One alternative to achieve this purpose is to design computational methods. Previous studies modeled a binary classification problem by pairing drugs and side effects; however, their classifiers can only extract one feature from each type of drug association. The present work proposed a novel multiple-feature sampling scheme that can extract several features from one type of drug association. Thirteen classification algorithms were employed to construct classifiers with features yielded by such scheme. Their performance was greatly improved compared with that of the classifiers that use the features yielded by the original scheme. Best performance was observed for the classifier based on random forest with MCC of 0.8661, AUROC of 0.969, and AUPR of 0.977. Finally, one key parameter in the multiple-feature sampling scheme was analyzed.

The Effect of Tissue Stromal Vascular Fraction as Compared to Cellular Stromal Vascular Fraction to Treat Anal Sphincter Incontinence.

BACKGROUND: The long-term prognosis of current treatments for anal sphincter incontinence (ASI) is poor. Here, we explored the efficacy of tissue adipose stromal vascular fraction SVF (tSVF) on ASI and compared it to that of cellular SVF (cSVF). We then investigated possible mechanisms. METHODS: Rat cSVF and tSVF were isolated and labeled with DIL. One day after modeling, three groups received phosphate-buffered saline (PBS), cSVF, tSVF, respectively. The control group received nil modeling nor any treatments. The effect was assessed by function test for anal pressure and electromyography, and staining for fiber content, proliferation and differentiation at day 5 and day 10. RESULTS: cSVF injection resulted in faster healing than tSVF. The cSVF group showed significant improvement on anal pressure on day 10. For the electromyography test, cSVF showed significant improvement for the frequencies on day 10, and for the peak values on both time points, while tSVF showed significant improvement for the peak values on day 10. The two SVF both alleviated fibrosis. Immunofluorescence tracing identified differentiation of some injected cells towards myosatellite cells and smooth muscle cells in both SVF groups. For all the tests, the tSVF group tends to have similar or lower effects than the cSVF group with no significant difference. CONCLUSION: cSVF and tSVF are both safe and effective in treating ASI, while the effect of cSVF is slighter higher than tSVF.

Enzymatic deamination of the epigenetic nucleoside N6-methyladenosine regulates gene expression.

N6-methyladenosine (m6A) modification is the most extensively studied epigenetic modification due to its crucial role in regulating an array of biological processes. Herein, Bsu06560, formerly annotated as an adenine deaminase derived from Bacillus subtilis 168, was recognized as the first enzyme capable of metabolizing the epigenetic nucleoside N6-methyladenosine. A model of Bsu06560 was constructed, and several critical residues were putatively identified via mutational screening. Two mutants, F91L and Q150W, provided a superiorly enhanced conversion ratio of adenosine and N6-methyladenosine. The CRISPR-Cas9 system generated Bsu06560-knockout, F91L, and Q150W mutations from the B. subtilis 168 genome. Transcriptional profiling revealed a higher global gene expression level in BS-F91L and BS-Q150W strains with enhanced N6-methyladenosine deaminase activity. The differentially expressed genes were categorized using GO, COG, KEGG and verified through RT-qPCR. This study assessed the crucial roles of Bsu06560 in regulating adenosine and N6-methyladenosine metabolism, which influence a myriad of biological processes. This is the first systematic research to identify and functionally annotate an enzyme capable of metabolizing N6-methyladenosine and highlight its significant roles in regulation of bacterial metabolism. Besides, this study provides a novel method for controlling gene expression through the mutations of critical residues.

The Clinical Significance of Mesenteric Lymphocytes in Human Colorectal Cancer.

OBJECTIVE: The mesentery is a potential site of residual tumor in patients with colorectal cancer (CRC). However, the mesenteric immune microenvironment remains unclear. In this study, we investigated the immune landscape of the mesentery, particularly the role of lymphocytes and its association with the clinicopathological characteristics of CRC. METHODS: Flow cytometry was used to detect lymphocytes in the paired mesenteric tissue specimens adjacent to the colorectal tumors and normal mesenteric tissue specimens 10 cm away from the colorectal tumor edge and preoperative peripheral blood samples obtained from patients with CRC who underwent surgery. T-distributed stochastic neighbor embedding was utilized to analyze multiparameter flow cytometry data. Multiplex immunohistochemistry was performed to evaluate T cells subsets in the paired mesentery adjacent to the colorectal tumors and normal mesentery. The Fisher's exact test and non-parametric Wilcoxon's matched-pairs tests were used for statistical analysis. The non-parametric Mann-Whitney U test was used to determine associations between percentage data and clinical parameters of patients with CRC. RESULTS: We found that immune cells in the normal mesentery were mainly of lymphoid lineage. Compared with peripheral blood, the normal mesentery showed decreased NK cells and the CD4/CD8 ratio and increased CD3+ CD56+, memory CD4+ T, memory CD8+ T, CD4+ tissue-resident memory T (TRM), and CD8+ TRM cells. Compared with the normal mesentery, the mesentery adjacent to the colorectal tumor showed increased B and regulatory T cells and decreased NK, CD3+ CD56+, CD4+ TRM, and CD8+ TRM cells. Moreover, memory CD8+ T cells and plasmablasts are negatively correlated with the depth of invasion of CRC. Increased memory CD4+ T cells are associated with distant metastasis of CRC and high preoperative serum carcinoembryonic antigen levels. CONCLUSION: The mesentery shows a specific immune microenvironment, which differs from that observed in peripheral blood. CRC can alter the mesenteric immune response to promote tumor progression.