Liver metastasis is a primary cause of death in colorectal cancer, yet the role of immune cells called neutrophils in this process remains complex. In this study, we identify three distinct neutrophil subtypes: anti-tumor high-density neutrophils, pro-tumor mature low-density neutrophils, and immature low-density neutrophils. Here we show that tumor-derived GM-CSF converts anti-tumor neutrophils into a pro-tumor state via the AKT-NF-κB-NFAT5 signaling axis. These activated neutrophils, when further stimulated by CXCR2 ligands, release web-like structures known as neutrophil extracellular traps which promote cancer spread by enhancing cancer cell growth, suppressing immune responses, and preparing the liver for metastasis. Importantly, we demonstrate that using antibodies depleting neutrophils or drugs inhibiting their pro-tumor activity effectively reduces liver metastasis in mice. Our work defines specific neutrophil subsets as key drivers of cancer progression and highlights their therapeutic potential in preventing metastatic spread.

The microenvironment of distant organs affects the colonization and growth of disseminated tumor cells. It remains unclear how tumor-associated neutrophils are influenced by the microenvironment of distant organs. Here, we demonstrate that mature low-density neutrophils in colorectal cancer patients abnormally accumulate neutral lipids and induce the reactivation of dormant tumor cells, a process regulated by hepatic stellate cells. Mechanistically, activated hepatic stellate cells increased DGAT1/2-dependent lipid droplet synthesis in low-density neutrophils through the secretion of IL33, thereby maintaining the survival and immunosuppressive function of these neutrophils. The uptake of lipids from lipid-laden low-density neutrophils drives dormant tumor cell reactivation through the potentiation of β-oxidation and the stimulation of protumorigenic eicosanoid synthesis. In mouse models, targeting IL33 blocked neutrophil lipid synthesis, decreased the colonization of colorectal cancer cells in the liver, and enhanced the efficacy of immunotherapy. Overall, our study revealed that lipid accumulation in mature low-density neutrophils regulates the growth of dormant tumor cells and antitumor immunity to facilitate colorectal cancer liver metastasis. Targeting IL33 could be a promising therapeutic approach for colorectal cancer liver metastases.

Background In laparoscopic low anterior resection for rectal cancer surgery, there has been controversy to whether the inferior mesenteric artery (IMA) should be ligated at the origin of its aorta (high ligation (HL)) or below the branches of the left colonic artery (LCA) (low ligation (LL)). This study was intended to clarify oncological outcome and long-term prognosis of retrospective analysis. Methods Analyzed the cases who underwent laparoscopic low anterior resection (LAR) in Shanghai Ruijin Hospital from January 2015 to December 2016, 357patients scheduled into 2 groups according to the level of IMA ligation: HL ( n = 247) versus LL ( n = 110). Results The primary endpoint is long-term outcomes, and the secondary endpoint is the incidence rate of major postoperative complications. There were no significant differences in 5-year overall survival ( P = 0.92) and 5-year disease-free survival ( P = 0.41). There were no differences between the clinical baseline levels in each group. The incidence of low anterior resection syndrome (LARS) in the two groups was statistically significant ( P = 0.037). No significant differences were observed in operative time ( P = 0.092) and intraoperative blood loss ( P = 0.118). In the HL group, 6 cases (2.4%) had additional colonic excision due to poor anastomotic blood supply; none of the colonic anastomosis in the low ligation group had ischemic manifestations, and length from the proximal margin ( P = 0.076), length from the distal margin ( P = 0.184), the total number of lymph nodes excised ( P = 0.065), and anastomotic leakage incidence ( P = 0.33). Conclusion Low ligation of the IMA which reserved LCA with vascular root lymph node dissection in laparoscopic low anterior resection for rectal cancer surgery may help protect the blood supply of the anastomosis, and will not increase postoperative complications while enhance recovery, without compromising radical excision and long-term prognosis.

Locally advanced colorectal cancer requires preoperative chemotherapy to reduce local recurrence and metastasis rates, but it remains difficult to predict the tumor will be sensitive to which treatments. The patient-derived organoids (PDOs) are considered an effective platform for predicting tumor drug responses in precision oncology. However, it has the limitation of being time-consuming in practical applications, especially in neoadjuvant treatment. Here we used cancer tissue-originated spheroids (CTOS) method to establish organoids from a heterogeneous population of colorectal cancer specimens, and evaluated the capacity of CTOS to predict clinical drug responses. By analyzing the relationship of the activities of drug-treated CTOS, drug targets and target-related pathways, tumor intrinsic effective-target-related pathways can be identified. These pathways were highly matched to the abnormal pathways indicated by whole-exome sequencing. Based on this, we used half effective concentration gradients to classify CTOS as sensitive or resistant to chemotherapy regimens within a week, for predicting neoadjuvant treatment outcomes for colorectal cancer patients. The drug sensitivity test results are highly matched to the clinical responses to treatment in individual patients. Thus, our data suggested that CTOS models can be effectively screened ex vivo to identify pathways sensitive to chemotherapies. These data also supported organoid research for personalized clinical medication guidance immediately after diagnosis in patients with advanced colorectal cancer.

Forkhead box D1 (FOXD1) serves a critical role in colorectal cancer (CRC). FOXD1 expression is an independent prognostic factor in patients with CRC; however, the molecular mechanism and signaling pathway of FOXD1 that regulates cell stemness and chemoresistance has not been fully characterized. The aim of the present study was to further validate the effect of FOXD1 on the proliferation and migration of CRC cells, and to delve into the possible potential of FOXD1 in the clinical treatment of CRC. The effect of FOXD1 on cell proliferation was assessed using Cell Counting Kit 8 (CCK-8) and colony formation assays. The effect of FOXD1 on cell migration was assessed by wound-healing and Transwell assays. The effect of FOXD1 on cell stemness was assessed by spheroid formation in vitro and limiting dilution assays in vivo. The expression of stemness associated proteins, leucine rich repeat containing G protein-coupled receptor 5 (LGR5), OCT4, Sox2 and Nanog, and epithelial-mesenchymal transition associated proteins, E-cadherin, N-cadherin and vimentin, were detected by western blotting. Proteins interrelationships were assessed by a co-immunoprecipitation assay. Oxaliplatin resistance was assessed using CCK-8 and apoptosis assays in vitro, and using a tumor xenograft model in vivo. By constructing FOXD1 overexpression and knockdown stably transfected strains of colon cancer cells, it was revealed that the overexpression of FOXD1 increased CRC cell stemness and chemoresistance. By contrast, knockdown of FOXD1 produced the opposite effects. These phenomena were caused by the direct interaction between FOXD1 and beta-catenin, thus promoting its nuclear translocation and the activation of downstream target genes, such as LGR5 and Sox2. Notably, inhibition of this pathway with a specific beta-catenin inhibitor (XAV-939) could impair the effects induced by the overexpression of FOXD1. In summary, these results indicated that FOXD1 may promote cell stemness and the chemoresistance of CRC by binding directly to beta-catenin and enhancing beta-catenin nuclear localization; therefore, it may be considered a potential clinical target.

Background The surgical procedure for laparoscopic right colectomy (LRC) is not standardized. Some published studies show the superiority of ileocolic anastomosis (IIA), but the evidence so far is insufficient. This study aimed to investigate the potential advantages in postoperative recovery and safety of IIA in LRC. Methods A total of 114 patients who underwent LRC with IIA ( n = 58) or extracorporeal ileocolic anastomosis (EIA, n = 56) between January 2019 and September 2021 were enrolled. We collected certain factors as clinical features, intraoperative characteristics, oncological outcomes, postoperative recovery, and short-term outcomes. Our primary outcome was time to gastrointestinal (GI) function recovery. Secondary outcomes were postoperative complications within 30 days, postoperative pain, and length of hospital stay. Results Faster GI recovery and less postoperative pain were observed in patients with IIA compared to EIA [time to first flatus: (2.4 ± 0.7) vs (2.8 ± 1.0) days, p < 0.01; time to liquid intake: (3.5 ± 0.7) vs (4.0 ± 1.1) days, p = 0.01; postoperative visual analogue scale score: (3.9 ± 1.0) vs (4.3 ± 0.6), p = 0.02]. No significant differences were detected in oncological outcomes or postoperative complications. IIA, rather than EIA, tended to be performed in patients with higher body mass index [(23.93 ± 3.52) vs (22.36 ± 2.87) kg/m 2 , p = 0.01]. Conclusions IIA is associated with faster GI function recovery and less postoperative pain and may be more favorable for obese patients.

IntroductionThe role of tumour secretory cytokines and peripheral circulatory cytokines in tumour progression has received increasing attention; however, the role of tumour-related inflammatory cytokines in colorectal cancer (CRC) remains unclear. In this study, the concentrations of various cytokines in the peripheral blood of healthy controls and patients with CRC at different stages were compared.MethodsPeripheral blood samples from 4 healthy participants and 22 colorectal cancer patients were examined. Luminex beads were used to evaluate concentration levels of 40 inflammatory cytokines in peripheral blood samples.ResultsIn peripheral blood, compared with healthy controls and early stage (I + II) CRC patients, advanced CRC (III + IV) patients had increased concentrations of mononuclear/macrophage chemotactic-related proteins (CCL7, CCL8, CCL15, CCL2, and MIF), M2 polarization-related factors (IL-1β, IL-4), neutrophil chemotactic and N2 polarization-related cytokines (CXCL2, CXCL5, CXCL6, IL-8), dendritic cells (DCs) chemotactic-related proteins (CCL19, CCL20, and CCL21), Natural killer (NK) cell related cytokines (CXCL9, CXCL10), Th2 cell-related cytokines (CCL1, CCL11, CCL26), CXCL12, IL-2, CCL25, and CCL27, and decreased IFN-γ and CX3CL1 concentrations. The differential upregulation of cytokines in peripheral blood was mainly concentrated in CRC patients with distant metastasis and was related to the size of the primary tumour; however, there was no significant correlation between cytokine levels in peripheral blood and the propensity and mechanism of lymph node metastasis.DiscussionDifferent types of immune cells may share the same chemokine receptors and can co-localise in response to the same chemokines and exert synergistic pro-tumour or anti-tumour functions in the tumour microenvironment. Chemokines and cytokines affect tumour metastasis and prognosis and may be potential targets for treatment.

Background Interleukin-17A (IL-17A), a proinflammatory cytokine primarily secreted by Th17 cells, γδT cells and natural killer T (NKT) cells, performs essential roles in the microenvironment of certain inflammation-related tumours by regulating cancer growth and tumour elimination proved in previous literature. In this study, the mechanism of IL-17A that induces mitochondrial dysfunction promoted pyroptosis has been explored in colorectal cancer cells. Method The records of 78 patients diagnosed with CRC were reviewed via the public database to evaluate clinicopathological parameters and prognosis associations of IL-17A expression. The colorectal cancer cells were treated with IL-17A, and the morphological characteristics of those cells were indicated by scanning electron microscope and transmission electron microscope. After IL-17A treatment, mitochondrial dysfunction was tested by mitochondrial membrane potential (MMP) and reactive oxygen species (ROS). The expression of pyroptosis associated proteins including cleaved caspase-4, cleaved gasdermin-D (GSDMD), IL-1β, receptor activator of nuclear NOD-like receptor family pyrin domain containing 3 (NLRP3), apoptosis-associated speck like protein containing a card (ASC), and factor-kappa B was measured through western blotting. Results Positive IL-17A protein expression was observed in CRC compared to the non-tumour tissue. IL-17A expression indicates a better differentiation, earlier stage, and better overall survival in CRC. IL-17A treatment could induce mitochondrial dysfunction and stimulate intracellular reactive oxygen species (ROS) production. Furthermore, IL-17A could promote pyroptosis of colorectal cancer cells and significantly increase the secretion of inflammatory factors. Nevertheless, the pyroptosis induced by IL-17A could be inhibited through the pre-treatment with Mito-TEMPO (a mitochondria-targeted superoxide dismutase mimetic with superoxide and alkyl radical scavenging properties) or Z-LEVD-FMK (caspase-4 inhibitor, fluoromethylketone). Additionally, after being treated with IL-17A, an increasing number of CD8 + T cells showed in mouse-derived allograft colon cancer models. Conclusion IL-17A, as a cytokine mainly secreted by γδT cells in the colorectal tumour immune microenvironment, can regulate the tumour microenvironment in multiple ways. IL-17A could induce mitochondrial dysfunction and pyroptosis through the ROS/NLRP3/caspase-4/GSDMD pathway, and promote intracellular ROS accumulation. In addition, IL-17A can promote the secretion of inflammatory factors such as IL-1β、IL-18 and immune antigens, and recruit CD8 + T cells to infiltrate tumours. Graphical Abstract

KRAS is one of the leading mutations reported in colon cancer. However, there are few studies on the application of KRAS related signature in predicting prognosis and drug sensitivity of colon cancer patient. We identified KRAS related differentially expressed genes (DEGs) using The Cancer Genome Atlas (TCGA) database. A signature closely related to overall survival was recognized with Kaplan-Meier survival analysis and univariate cox regression analysis. Then we validated this signature with overall expression score (OE score) algorithm using both scRNA-seq and bulk RNA-seq data. Based on this signature, we performed LASSO cox regression to establish a prognostic model, and corresponding scores were calculated. Differences in genomic alteration, immune microenvironment, drug sensitivity between high- and low-KRD score groups were investigated. A KRAS related signature composed of 80 DEGs in colon cancer were recognized, among which 19 genes were selected to construct a prognostic model. This KRAS related signature was significantly correlated with worse prognosis. Furthermore, patients who scored lower in the prognostic model presented a higher likelihood of responding to chemotherapy, targeted therapy and immunotherapy. Furthermore, among the 19 selected genes in the model, SPINK4 was identified as an independent prognostic biomarker. Further validation in vitro indicated the knockdown of SPINK4 promoted the proliferation and migration of SW48 cells. In conclusion, a novel KRAS related signature was identified and validated based on clinical and genomic information from TCGA and GEO databases. The signature was proved to regulate genomic alteration, immune microenvironment and drug sensitivity in colon cancer, and thus might serve as a predictor for individual prognosis and treatment.

Dear Editor, The existing preclinical drug models to screen cancer drug candidates are inefficient in achieving satisfactory results for clinical applications, and chemotherapy resistance remains a major problem in colorectal cancer. To search for effective therapeutic drugs, we innovatively employed a cancer tissue-originated spheroids (CTOS)-based screening method to identify personalised treatments and predict drug responses for neoadjuvant therapy. Testing of new drug candidates is predominantly performed in cancer cell lines, but these cell lines do not retain the characteristics of the original tumours they were derived from, in part due to the numerous passages.1 To overcome the low efficacy of therapeutic screening and take inter-patient variations into account, we established colorectal cancer patient-derived organoids through CTOS methods.2, 3 After tissue dissociation, the cell aggregates with diameters of 40–100 μm were collected for further culture (Figure 1A). CTOS appeared as irregular fragments immediately after filtration and then formed transparent and smooth spheres after culture (Figure S1A). Inter- and intratumour heterogeneity was observed in these CTOS, each retaining different differentiation degrees of the original adenocarcinomas with poorly differentiated lesions appearing as solid growths and well-differentiated ones as crypt-like (Figure S1B,C). The mutations of KRAS, BRAF, TP53 and PIK3CA matched well between CTOS and the original tumours (Figure 1B, Table S4). Histological characteristics, epithelial marker expression (EpCAM), proliferation level (Ki67) and mucin-producing ability (MUC2) were confirmed to have strong concordance with the respective tumours (Figure 1C,D). The extracted cell aggregates consisted of almost pure tumour cells with few immune or vascular endothelial cells (Figures 1E and S1D,E). Since cancer stem cells have an intrinsic mechanisms of drug resistance,4 the expression of the stemness markers CD133, CD166 (Figure 1F,G) and Lgr5 (Figure S1F) was evaluated, and they were slightly elevated during culture. It remained to be clarified, however, the availability of our CTOS model in the screening of a drug candidate, the detection of drug-induced cell death and its role in the immune response. Initially, a panel of 60 drugs (Table S2) was tested in our CTOS-based screening assay (Figure 2A). The after-treatment morphological changes of the spheroids directly indicated the drug effects. The signs of damaged cells included low transparency, disorganised shape with an irregular surface and disaggregation into single cells (Figure S1C). Our method of cell viability measurement proved to have similar inspection efficiency as evaluations by adenosine triphosphate (ATP) levels (Figures S1G,H). We identified four hit inhibitors, obatoclax mesylate (OM), BI 2536 (BI), (S)-(+)-camptothecin (CPT) and bortezomib (BTZ), showing marked growth suppression of the CTOS (Figure 2B–E). All agents demonstrated effective growth inhibition in a dose- and time-dependent manner and marked treatment-induced apoptosis (Figure S2). However, apoptotic cancer cells incapable of killing themselves tend to gain other mutations, leading to genetic instability and diversity of malignant cells.5 If different types of cell death operate synergistically, the toxicity of the agent may be reduced. Interestingly, when examining caspase-3 activity, we noticed that drug-damaged cells showed balloon-like bubbles distinct from classic apoptotic blebbing (Figure S2D). This was reminiscent of the typical pyroptotic morphology. We observed their appearance in culture, and the colorectal cancer cells, especially the suspension ones, were swelled and exhibited large bubbles of the plasma membrane after treatment (Figure 3A). We verified the existence of gasdermin E (GSDME) expression in SW1463 and SW620 cells (Figure S3A). The release of lactate dehydrogenase (LDH) and high mobility group proteins B1 (HMGB1) in culture medium was elevated in cells treated with the four focused drugs (Figure 3B,C), indicating cell membrane rupture and leakage. Cells undergoing necroptosis have a similar swelling phenotype and change in membrane permeability.6 To distinguish which pathway OM, BI, CPT and BTZ induced cell death through, we examined relative protein levels by western blot. Cleavage of both caspase-3 and poly (ADP-ribose) polymerase 1 (PARP1, markers of apoptosis) was observed in BI- and CPT-treated colorectal cancer cells, and GSDME, though not gasdermin D (GSDMD), was cleaved to generate N-terminal fragments (markers of pyroptosis, Figures 3D and S3E). No changes in mixed lineage kinase domain-like protein (MLKL) phosphorylation (a marker of necroptosis) appeared after OM, BI or CPT exposure (Figure 3E), while BTZ-treated cells underwent necroptosis instead of pyroptosis (Figure 3D,E). Moreover, when treated, the peripheral cells of CTOS first exhibited large bubbles of the membrane, and single cells dissociated from the spheroids (Figure 3F). We then employed siRNA to verify whether BI- and CPT-induced GSDME-mediated pyroptosis was dependent on caspase-3.7 After GSMDE knockdown, the pyroptotic phenotype was diminished without an influence of caspase-3 activation (Figure 3G,J). After the knockdown of caspase-3, the expression of both cleaved caspase-3 and the N-terminal fragments of GSDME were downregulated (Figure 3J), suggesting that the activation of caspase-3 regulates the cleavage of GSDME in BI- and CPT-induced pyroptosis of colorectal cancer cells. The release of LDH and HMGB1 was markedly decreased after the knockdown of GSDME or caspase-3 (Figure 3H,I). The above results were also confirmed with a caspase-3 inhibitor, Z-DEVD-FMK (DEVD) (Figure S3G–K), implying the pyroptosis was secondary to the activation of caspase-3. In vivo drug responses were validated with the following strategy. BLAB/c-nude and BLAB/c mice bearing subcutaneous tumours were treated with dimethyl sulfoxide (DMSO), OM, BI, CPT or BTZ via intraperitoneal and intratumoural injection every other day four times (Figure 4A). After the treatments, the growth rates of the tumours in either immunocompetent mice or immunodeficient mice were robustly inhibited compared with the vehicle control values (Figure 4B,C). In addition, no significant toxicity to the liver or kidney was visible, according to their morphology (Figure S4A,B). We also confirmed the drug efficacy in the patient-derived xenograft model (Figure S4C–E). Moreover, we surprisingly found that mice with an intact immune system exhibited better therapeutic effectiveness than nude mice (Figure 4B). Thus, we speculated that these small molecule drugs altered the immune response. As depicted in Figure 4E,F, CD4+ and CD8+ T cells were clustered in the tumour sites after treatment. In conclusion, using our strategy of CTOS-based drug screening, four drug candidates for colorectal cancer treatment were found. Among them, two small molecule inhibitors, BI and CPT, were further verified to induce GSDME-mediated pyroptosis concurrent with caspase-3-dependent apoptosis, and they showed both in vitro and in vivo antitumoural activity (Figure 4G). Pyroptosis induced by BI and CPT is an unrecognised mechanism that may feasibly enhance the efficacy of immunotherapy. We are grateful to all patients enrolled in the study. This work was supported by National Nature Science Foundation of China grant 81871933 (Aiguo Lu), Science and Technology Commission of Shanghai Municipality grant 20Y11909000 (Aiguo Lu), Shanghai Translational Medicine Collaborative Innovation Center grant TM202002 (Aiguo Lu) and National Nature Science Foundation of China grant 81802326 (Jingkun Zhao). The authors have declared that no conflicts of interest exists. Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.

Colorectal cancer (CRC) is a worldwide disease with worse survival. Our objective is to identify previously unrecognized prognostic factors to better evaluate disease progression. Seven GEO datasets were collected and analysed using R software, followed by KEGG enrichment analysis and TFs network construction. LASSO‐COX analysis was performed to select the most useful prognostic features. COX model was used to analyse prognostic factors associated with OS. The survival curve was constructed using Kaplan–Meier analysis. A Nomogram model was also constructed to predict prognosis. A total of 3559 differentially expressed genes (DEGs) and 66 differentially expressed transcription factors were identified. FOXD1 was identified as the most differentially expressed factor of TFs covering the most downstream DEGs and independent risk prognostic factor. Next, FOXD1 expression was detected using immunohistochemical staining in 131 CRC patients’ tissue and the association between FOXD1 expression and clinicopathologic features was analysed. High expression of FOXD1 was correlated with TNM stage and pathological differentiation. Multivariate COX regression analyses confirmed that FOXD1 high‐expression, TNM stage and tumour differentiation were independent prognostic risk factor of OS and DFS. Patients with high expression of FOXD1 were more likely to have poor overall survival and disease‐free survival. The combination of FOXD1 and Plk2 which we have previously reported allowed us to predict the survival of post‐surgical CRC patients more accurately, adding to the former prognostic model based on the TNM Stage. The results showed that patients with high expression of both FOXD1 and Plk2 have the worst survival. A combination of FOXD1 and Plk2 can better evaluate patients’ survival.

CXCL5 is overexpressed in colorectal cancer (CRC) and promotes distant metastasis and angiogenesis of tumors; however, the underlying mechanism that mediates CXCL5 overexpression in CRC remains unclear. Here, we successfully extracted and identified primary mesenchymal stromal cells (MSCs) and verified the promoting effects of tumor-associated MSCs on CRC proliferation and metastasis in vivo and in vitro. We found that MSCs not only promoted the expression of CXCL5 by secreting CCL7 but also secreted TGF-beta to inhibit this process. After secretion, CCL7/CCR1 activated downstream CBP/P300 to acetylate KLF5 to promote CXCL5 transcription, while TGF-beta reversed the effect of KLF5 on transcription activation by regulating SMAD4. Taken together, our results indicate that MSCs in the tumor microenvironment promoted the progression and metastasis of CRC and regulated the expression of CXCL5 in CRC cells by secreting CCL7 and TGF-beta. KLF5 is the key site of these processes and plays a dual role in CXCL5 regulation. MSCs and their secreted factors may serve as potential therapeutic targets in the tumor environment.

Oxaliplatin resistance is a major challenge in the treatment of colorectal cancer (CRC). Many molecular targeted drugs for refractory CRC have been developed to solve CRC drug resistance, but their effectiveness and roles in the progression of CRC and oxaliplatin resistance remain unclear. Here, we successfully constructed CRC PDOs and selected the Kruppel-like factor 5 (KLF5) inhibitor ML264 as the research object based on the results of the in vitro drug screening assay. ML264 significantly restored oxaliplatin sensitivity in CRC PDOs by restoring the apoptotic response, and this effect was achieved by inhibiting the KLF5/Bcl-2/caspase3 signaling pathway. Chromatin immunoprecipitation (ChIP) and luciferase reporter assays verified that KLF5 promoted the transcription of Bcl-2 in CRC cells. KLF5 inhibition also overcame oxaliplatin resistance in xenograft tumors. Taken together, our study demonstrated that ML264 can restore oxaliplatin sensitivity in CRC PDOs by restoring the apoptotic response. KLF5 may be a potential therapeutic target for oxaliplatin-resistant CRC. PDOs have a strong potential for evaluating inhibitors and drug combination therapy in a preclinical environment.

Colorectal cancer (CRC) is the third most common form of cancer, and the incidence of sporadic young-onset colorectal cancer (yCRC) has been increasing. Microbiota residing in the tumor microenvironment are emerging tumor components. The colonic microbiome differs between patients with CRC and healthy controls; however, few studies have investigated the role of the tumor microbiota in disease diagnosis and tumorigenesis of yCRC. We performed 16S rRNA sequencing analysis to identify the microbiome in CRC and found that tumor microbial diversity decreased in yCRC. Proteobacteria and Firmicutes were the most abundant phyla in all CRC samples, and Actinomyces and Schaalia cardiffensis were the key microbiota in the yCRC group. Correlation analysis revealed that Actinomyces co-occurred with various pro-tumor microbial taxa, including Bacteroidia, Gammaproteobacteria, and Pseudomonas . An independent cohort was used to validate the results. The Actinomyces in CRC was co-localized with cancer-associated fibroblasts and activated the TLR2/NF-κB pathway and reduces CD8+ T lymphocyte infiltration in CRC microenvironment. This study suggests that tumoral microbiota plays an important role in promoting tumorigenesis and therefore has potential as a promising non-invasive tool and intervention target for anti-tumor therapy.

BACKGROUND Neoadjuvant therapy (NAT) is becoming increasingly important in locally advanced rectal cancer. Hence, such research has become a problem. AIM To evaluate the downstaging effect of NAT, its impact on postoperative complications and its prognosis with different medical regimens. METHODS Seventy-seven cases from Shanghai Ruijin Hospital affiliated with Shanghai Jiaotong University School of Medicine were retrospectively collected and divided into the neoadjuvant radiochemotherapy (NRCT) group and the neoadjuvant chemotherapy (NCT) group. The differences between the two groups in tumor regression, postoperative complications, rectal function, disease-free survival, and overall survival were compared using the χ2 test and Kaplan-Meier analysis. RESULTS Baseline data showed no statistical differences between the two groups, whereas the NRCT group had a higher rate of T4 (30/55 vs 5/22, P < 0.05) than the NCT groups. Twelve cases were evaluated as complete responders, and 15 cases were evaluated as tumor regression grade 0. Except for the reduction rate of T stage (NRCT 37/55 vs NCT 9/22, P < 0.05), there was no difference in effectiveness between the two groups. Preoperative radiation was not a risk factor for poor reaction or anastomotic leakage. No significant difference in postoperative complications and disease-free survival between the two groups was observed, although the NRCT group might have better long-term overall survival. CONCLUSION NAT can cause tumor downstaging preoperatively or even complete remission of the primary tumor. Radiochemotherapy could lead to better T downstaging and promising overall survival without more complications.

Platelets promote tumor growth and metastasis in several tumor types. Recent research has found platelets can extravasate and infiltrate into the tumor stroma and interact with the tumor microenvironment. The prognostic role of platelet infiltration in colorectal cancer (CRC) remains controversial. A pan-cancer survival analysis was performed to find the potential prognostic value of platelet infiltration in patients with cancer. A survival analysis and a nomogram prognostic model were established to further confirm the results with data from our center. The correlations between patient outcomes and tumor-infiltrating platelets (TIPs) were identified by immunohistochemical staining for CD42b. The prognostic accuracy and discriminative ability of the nomogram were determined by the concordance index (C-index) and a calibration curve. The pan-cancer survival analysis showed platelet infiltration can lead to a poor prognosis in patients with several types of cancers, including CRC. Platelet infiltration was associated with overall survival (OS) and disease-free survival (DFS) in both primary and validation cohorts. The C-index values of the nomogram for predicting OS and DFS were 0.774 and 0.769, respectively, which were higher than that of the TNM staging system alone. Our study found platelet infiltration has a potential prognostic value regarding postsurgical survival in CRC patients. The proposed nomogram resulted in a more accurate prognostic prediction for postsurgical CRC patients.

BACKGROUND AND OBJECTIVES:There is controversy regarding whether the inferior mesenteric artery (IMA) should be ligated at its origin from the aorta (high ligation, HL) or below the branch of the left colic artery (low ligation, LL) during surgery for rectal cancer. METHODS:This prospective study randomized 95 patients with histologically proven rectal cancer (clinical stages I-III based on the 8th American Joint Committee on Cancer guidelines) to undergo HL (n = 47) or LL with lymph node dissection at the root of the IMA (n = 48). RESULTS:Only two intraoperative adverse events were observed (two HL patients experienced anastomotic ischemia and underwent extended bowel excision and splenic flexure mobilization). The LL group had a significantly shorter time to first flatus (p < .0001). No significant differences were observed in operative time (p = .14), intraoperative blood loss (p = .21), distance from the upper margin (p = .77), distance from the lower margin (p = .35), harvested lymph nodes (p = .33), or anastomotic leakage (p = .44), 2-year overall survival (p = .97), or 2-year disease-free survival (p = .42). CONCLUSION:During laparoscopic low anterior resection, a combination of LL at the IMA and vascular root lymph node dissection may help protect the blood supply of the anastomosis, reduce postoperative complications, and enhance recovery, without compromising radical excision.

Background and Objectives To investigate the effects of different suture reinforcement methods for anastomotic leakage and other postoperative complications after the use of a laparoscopic double stapling technique (DST). Methods We collected the data of 124 patients who underwent laparoscopic radical resection of colorectal cancer from July 2017 to September 2018 at our institution. Patients were divided into three groups according to the suture reinforcement methods: intermittent, continuous suture reinforcement, and non-reinforcement (n = 41, 41, and 42, respectively). One-way analysis of variance, chi(2), Fisher's exact, and nonparametric tests were used for statistical analysis. Results Among the 124 patients, there were no statistically significant differences in operation times, intraoperative blood loss, postoperative hospital stays and recovery of bowel movement. Nine patients were diagnosed with anastomotic leakage (AL). The incidences of serious AL in the intermittent and continuous suture reinforcement groups were lower than that in the control group, with lower reoperation rate, shorter average lengths of stay and lower treatment costs of two experimental groups. Conclusion Intermittent and continuous sutures after laparoscopic DST is effective, safe, and feasible on anastomotic leakage prevention. These procedures could be popularized in rectal surgery on patients with high risk of AL.

Objective:To primarily investigate the application value of glasses-free three-dimensional (3D) laparoscopy system in laparoscopic radical resection of gastrointestinal malignant tumors.Methods:The retrospective cohort study was conducted. The clinical data of 165 patients with gastrointestinal malignant tumors who were admitted to the Ruijin Hospital of Shanghai Jiaotong University School of Medicine between October 2018 and May 2019 were collected. There were 99 males and 66 females, aged from 28 to 86 years, with a median age of 63 years. There were 68 of 165 patients with gastric cancer and 97 with colorectal cancer. Sixteen patients with gastric cancer who underwent laparoscopic radical gastrectomy using the glasses-free 3D laparoscopy system were divided into glasses-free 3D gastric cancer group, and 52 patients with gastric cancer who underwent laparoscopic radical gastrectomy using the polarized glasses 3D laparoscopy system were divided into polarized glasses 3D gastric cancer group. Nineteen patients with colorectal cancer who underwent laparoscopic radical colectomy or proctectomy using the glasses-free 3D laparoscopy system were divided into glasses-free 3D colorectal cancer group, and 78 patients with colorectal cancer who underwent laparoscopic radical colectomy or proctectomy using the polarized glasses 3D laparoscopy system were divided into polarized glasses 3D colorectal cancer group. Observation indicators: (1) operative situations of patients with gastric cancer; (2) postoperative recovery of patients with gastric cancer; (3) postoperative pathological examination results of patients with gastric cancer; (4) operative situations of patients with colorectal cancer; (5) postoperative recovery of patients with colorectal cancer; (6) postoperative pathological examination results of patients with colorectal cancer; (7) follow-up. Follow-up was conducted by outpatient examination or telephone interview to detect complications and survival of patients up to the postoperative 30th day. Measurement data with skewed distribution were represented as M ( P25, P75) or M (range), and comparison between groups was analyzed by the Mann-Whitney U test. Count data were represented as absolute numbers or percentages, and comparison between groups was analyzed using the chi-square test or Fisher exact probability. Results:(1) Operative situations of patients with gastric cancer: all the 68 gastric cancer patients received successfully laparoscopic radical gastrectomy, without intraoperative complication or conversion to laparotomy. Cases with distal gastrectomy or total gastrectomy(surgical methods) , operation time, volume of intraoperative blood loss were 11, 5, 195 minutes(169 minutes, 214 minutes), 20 mL (10 mL, 90 mL) in patients of the glasses-free 3D gastric cancer group, and 31, 21, 196 minutes(173 minutes, 222 minutes), 40 mL(20 mL, 100 mL) in patients of the polarized glasses 3D gastric cancer group, with no significant difference between the two groups ( χ2=0.432, Z=-0.362, -1.065, P>0.05). (2) Postoperative recovery of patients with gastric cancer: the time to first flatus, time to initial semi-fluid diet intake, duration of postoperative hospital stay were 2 days(2 days, 3 days), 6 days(5 days, 7 days), 10 days(9 days, 14 days) in patients of the glasses-free 3D gastric cancer group, and 2 days(2 days, 3 days), 6 days(5 days, 6 days), 11 days(9 days, 14 days) in patients of the polarized glasses 3D gastric cancer group, with no significant difference between the two groups ( Z=-0.163, -1.870, -0.570, P>0.05). The postoperative complication incidence of the glasses-free 3D gastric cancer group was 12.5%(2/16), including 1 case with duodenal stump fistula, 1 case with anastomotic bleeding. The postoperative complication incidence of the polarized glasses 3D gastric cancer group was 17.3%(9/52), including 2 cases with duodenal stump fistula, 2 cases with delayed gastric emptying, 1 case with pulmonary infection, 1 case with abdominal bleeding, 1 case with anastomotic leakage, 1 case with chylous fistula, 1 case with intestinal obstruction. All the patients with complications were cured after symptomatic supportive treatment. There was no significant difference in the complication incidence between the two groups ( χ2=0.209, P>0.05). (3) Postoperative pathological examination results of patients with gastric cancer: the tumor diameter, cases in stage T1, T2, T3, T4 (tumor T staging), cases with vascular invasion, cases with nerve invasion, cases with tumor nodule, cases in stage N0, N1, N2, N3 (tumor N staging), the number of positive lymph node, the number of lymph node dissected, cases with qualified lymph node dissected, cases in stage Ⅰ, Ⅱ, Ⅲ, Ⅳ (TNM clinical staging) were 3.0 cm(2.0 cm, 5.0 cm), 5, 3, 1, 7, 8, 5, 2, 3, 2, 4, 7, 6(1, 15), 28(22, 43), 15, 4, 3, 9, 0 in patients of the glasses-free 3D gastric cancer group, and 3.5 cm(2.0 cm, 6.0 cm), 13, 10, 4, 25, 19, 23, 2, 26, 6, 7, 13, 1(0, 7), 29(21, 39), 43, 21, 10, 20, 1 in patients of the polarized glasses 3D gastric cancer group. There was no significant difference in the tumor diameter, tumor T staging, cases with vascular invasion, cases with nerve invasion, cases with tumor nodule, the number of lymph node dissected, cases with qualified lymph node dissected, TNM clinical staging between the two groups ( Z=-0.570, -0.434, χ2 =0.926, 0.851, 1.655, Z=-0.579, χ2=1.193, Z=-1.134, P>0.05). There were significant differences in the tumor N staging and the number of positive lymph node between the two groups ( Z=-2.167, -2.283, P<0.05). (4) Operative situations of patients with colorectal cancer: all the 97 colorectal cancer patients received successfully laparoscopic radical colectomy or proctectomy, without intraoperative complications or conversion to laparotomy. Cases with radical colectomy or proctectomy (surgical methods), operation time, volume of intraoperative blood loss were 7, 12, 132 minutes(97 minutes, 156 minutes), 20 mL(10 mL, 50 mL) in patients of the glasses-free 3D colorectal cancer group, and 40, 38, 124 minutes(110 minutes, 159 minutes), 25 mL(15 mL, 65 mL) in patients of the polarized glasses 3D colorectal cancer group, with no significant difference between the two groups ( χ2 =1.276, Z=-0.141, -0.863, P>0.05). (5) Postoperative recovery of patients with colorectal cancer: the time to first flatus, time to initial semi-fluid diet intake, duration of postoperative hospital stay were 2 days(1 days, 3 days), 5 days(5 days, 6 days), 8 days(7 days, 10 days) in patients of the glasses-free 3D colorectal cancer group, and 2 days(1 days, 3 days), 5 days(4 days, 6 days), 8 days(6 days, 10 days) in patients of the polarized glasses 3D colorectal cancer group, with no significant difference between the two groups ( Z=-0.678, -1.751, -1.674, P>0.05). The complication incidence of the glasses-free 3D colorectal cancer group was 15.8%(3/19), including 1 case with anastomotic leakage after low anterior proctectomy, 1 case with incision infection, 1 case with urinary tract infection. The complication incidence of the polarized glasses 3D colorectal cancer group was 14.1%(11/78), including 3 cases with anastomotic leakage after low anterior proctectomy, 2 cases with intestinal obstruction, 2 cases with urinary tract infection, 2 cases with incision infection, 1 case with anastomotic bleeding, 1 case with pulmonary infection. One of the 3 cases with anastomotic leakage after low anterior proctectomy in the polarized glasses 3D colorectal cancer group was cured after remedial terminal ileostomy. The other patients with complications were cured after symptomatic supportive treatment. There was no significant difference in the complication incidence between the two groups ( χ2=0.035, P>0.05). (6) Postoperative pathological examination results of patients with colorectal cancer: the tumor diameter, cases in stage T1, T2, T3, T4 (tumor T staging), cases with vascular invasion, cases with nerve invasion, cases with tumor nodule, cases in stage N0, N1-N2 (tumor N staging), the number of positive lymph node, the number of lymph node dissected, cases with qualified lymph node dissected, cases in stage Ⅰ, Ⅱ, Ⅲ, Ⅳ (TNM clinical staging) were 5.0 cm(3.0 cm, 6.0 cm), 3, 2, 7, 7, 3, 2, 1, 8, 11, 0(0, 4), 17(14, 23), 18, 2, 3, 12, 2 in patients of the glasses-free 3D colorectal cancer group, and 4.0 cm(3.0 cm, 5.0 cm), 7, 16, 43, 12, 14, 12, 7, 46, 32, 0(0, 1), 16(13, 19), 74, 14, 17, 40, 7 in patients of the polarized glasses 3D colorectal cancer group, with no significant difference between the two groups ( Z=-0.768, -1.135, χ2 =0.049, 0.292, 0.278, 1.762, Z=-0.694, -1.349, χ2=0.001, Z=-1.011, P>0.05). (7) Follow-up: 165 patients received follow-up, with out short-term reoperation or postoperative death in the postoperative 30 days. Conclusion:There is no significant difference in the efficacy between glasses-free 3D laparoscopic surgery and polarized glasses 3D laparoscopic surgery for radical resection of gastrointestinal malignant tumors, of which the clinical value requires further study.

目的·研究腹腔镜直肠双吻合术吻合口不同加固方式对吻合口漏的影响.方法·收集2017年7月—2018年9月上海交通大学医学院附属瑞金医院胃肠外科(上海市微创外科临床医学中心)收治的接受腹腔镜下直肠双吻合术的患者资料,根据吻合口加固方式的不同分为间断缝合加固组(n=41)、连续缝合加固组(n=41)、不加固组(对照组,n=42),比较3组术中和术后的情况.采用单因素方差分析、χ2检验或Fisher's精确概率法、非参数检验进行统计学分析.结果·共纳入124例患者,间断缝合加固组、连续缝合加固组的手术时间、术中失血量、术后住院天数、术后胃肠道排气时间、术后进食流质时间等指标与对照组比较,差异无统计学意义.9例被诊断为吻合口漏.间断缝合加固组、连续缝合加固组的严重吻合口漏发生率较对照组低.间断缝合加固组吻合口漏患者的平均住院天数为31(19～42)d,连续缝合加固组为41(37～43)d,均低于对照组的64(54～74)d.间断缝合加固组的平均治疗费用为(71142.6±2849.3)元,连续缝合加固组为(71360.1±2072.3)元,均少于对照组的(91386.0±9151.7)元.结论·腹腔镜直肠双吻合术后,应用间断缝合法和连续缝合法加固吻合口能够降低严重吻合口漏发生率、缩短住院时间、降低治疗费用,未增加手术时间及术后并发症风险.