BACKGROUND AND AIMS:Endoscopic submucosal dissection (ESD) enables en bloc resection of large colorectal neoplasms, but the recurrence risk with positive or indeterminate horizontal margins (HM1/x) remains debated. This meta-analysis evaluated the local recurrence rate after en bloc ESD with HM1/x margins. METHODS:A systematic search of PubMed, EMBASE, Web of Science, and Cochrane Library through June 2024 identified studies reporting local recurrence after colorectal en bloc ESD with HM1/x margins. Cases with positive or indeterminate vertical margins or histologic high-risk features were excluded. Cumulative incidences and odds ratios were pooled using mixed-effects logistic regression. RESULTS:Eleven studies with 441 HM1/x primary resections (331 benign, 12 invasive, 98 unknown) were included. The pooled recurrence rate was 4.3% (20/441; 95% CI, 2.5%-7.5%). Median time to recurrence was 14 months (range, 6-71). Histologic details were available for 16 recurrences (3 invasive, 13 benign). Two invasive recurrences originated from invasive lesions; the third from a high-grade dysplasia (HGD) lesion. Treatment of recurrence was reported for 10 cases, with 7 undergoing endoscopic resection (all benign) and 3 surgery (1 benign inflammatory bowel disease-related lesion, 2 invasive). The recurrence risk was significantly greater for HM1/x margins compared with negative horizontal margin cases (5/1655 vs 15/403; odds ratio, 8.04; 95% CI, 2.77-23.30). CONCLUSIONS:En bloc ESD with HM1/x margins is associated with a significantly greater risk of local recurrence compared with negative horizontal margin resection. However, the absolute risk remains low and most recurrences are benign. Because invasive recurrences seem to originate from initially invasive or high-grade dysplasia lesions, intensive surveillance should be prioritized for HM1/x cases with these characteristics, rather than all HM1/x cases.

BACKGROUND AND AIMS:Rehabilitation of physical activity is an important functional outcome after endoscopic surgery. Our aim was to quantitatively assess recovery after endoscopic submucosal dissection (ESD) and transanal minimally invasive surgery (TAMIS). METHODS:In the TRIASSIC study (Netherlands Trial Registry: NL7083), patients with rectal polyps >20 mm were randomized between ESD and TAMIS. This ancillary study used smartwatches to track activity data for a 14-day preoperative baseline period and a 28-day postoperative recovery period. The primary end point for noninferiority was the mean time to recovery (≥90% of baseline step count for 2 consecutive days), assessed by means of Weibull regression with a 7-day noninferiority margin. RESULTS:Forty patients were included: 20 ESD and 20 TAMIS procedures. Median lesion size was 42.5 mm (interquartile range [IQR], 25-50), with 17.5% pT1RCs and 82.5% nonmalignant rectal polyps. Compliance with smartwatch measurements was 98.4% (IQR, 94.2-100). Within the 28-day timespan, 17 patients (85%) in the ESD group recovered and 15 (75%) in the TAMIS group (P = .43). Mean recovery times were 13.9 days for ESD and 21.0 days for TAMIS, indicating noninferiority of ESD (95% confidence interval of difference, -3.41 to 20.25). Recovery as measured by smartwatch significantly correlated with self-reported recovery (Spearman rho, 0.644; P < .001). Moderate to severe pain scores (≥4 out of 10) were reported by 15 patients (42.9%): in 27.8% of the ESD group and 58.9% of the TAMIS-group (P = .06). Increased pain scores were significantly associated with decreased physical activity (P < .01). CONCLUSIONS:In terms of mean time to physical recovery, ESD was noninferior to TAMIS. Post-procedural pain was significantly associated with reduced physical activity.

BACKGROUND AND AIMS:The indication for primary surgery on suspected deep submucosal invasive colorectal carcinoma (d-SMIC) is debatable. Consequently, local excision techniques, such as endoscopic submucosal dissection (ESD), are increasingly attempted in such patients. This study retrospectively evaluated the effectiveness of ESD in obtaining a free vertical margin (VM)-R0 in suspected d-SMIC compared with suspected superficial SMIC (s-SMIC). METHODS:ESDs for suspected T1 colorectal cancer (CRC) in treatment-naïve polyps were included between 2011 and 2022 in 5 European tertiary referral centers. Based on optical assessment, lesions were categorized into suspected d-SMIC or s-SMIC. Main outcomes were the VM-R0 rate, en bloc resection rate, and adverse event rate. An adjusted risk ratio for VM-R1 resections within the suspected d-SMIC group was calculated. RESULTS:In the suspected s-SMIC group (n = 1063), en bloc resection rate, VM-R0 rate, proportion of pT1 CRC, and adverse even rate were 90.5% (95% CI, 88-92), 90.6% (95% CI, 88-92), 18.0%, and 3.6% (IQR, 2-5), respectively. In the suspected d-SMIC group (n = 139), these values were 61.9% (IQR, 54-70), 55.4% (IQR, 47-63), 74.8%, and 5.8% (IQR, 2-10), respectively. Compared with suspected s-SMIC cases, the VM-R0 rate of suspected d-SMIC cases particularly decreased for pT1Sm2-3 (75.9% vs 55.7%). None of the investigated features (age, sex, polyp location, size, morphology, and Hiroshima classification) predicted a VM-R1 resection in suspected d-SMIC cases. CONCLUSIONS:ESD performed on polyps with suspected d-SMIC showed lower VM-R0 rates for pT1Sm2-3 cases compared with suspected s-SMIC cases. This should be taken into account when selecting the optimal resection technique for suspected d-SMIC cases.

Fatty acid omega hydroxylase P450s consist of enzymes that hydroxylate various chain-length saturated and unsaturated fatty acids (FAs) and bioactive eicosanoid lipids. The human cytochrome P450 gene 4 family (CYP4) consists of 12 members that are associated with several human diseases. However, their role in the progression of metabolic dysfunction-associated fatty liver disease (MASLD) remains largely unknown. It has long been thought that the induction of CYP4 family P450 during fasting and starvation prevents FA-related lipotoxicity through FA metabolism to dicarboxylic acids that are chain-shortened in peroxisomes and then transported to the mitochondria for complete oxidation. Several studies have revealed that peroxisome succinate transported to the mitochondria is used for gluconeogenesis during fasting and starvation, and recent evidence suggests that peroxisome acetate can be utilized for lipogenesis and lipid droplet formation as well as epigenetic modification of gene transcription. In addition, omega hydroxylation of the bioactive eicosanoid arachidonic acid to 20-Hydroxyeicosatetraenoic acid (20-HETE) is essential for activating the GPR75 receptor, leading to vasoconstriction and cell proliferation. Several mouse models of diet-induced MASLD have revealed the induction of selective CYP4A members and the suppression of CYP4F during steatosis and steatohepatitis, suggesting a critical metabolic role in the progression of fatty liver disease. Thus, to further investigate the functional roles of CYP4 genes, we analyzed the differential gene expression of 12 members of CYP4 gene family in datasets from the Gene Expression Omnibus (GEO) from patients with steatosis, steatohepatitis, fibrosis, cirrhosis, and hepatocellular carcinoma. We also observed the differential expression of various CYP4 genes in the progression of MASLD, indicating that different CYP4 members may have unique functional roles in the metabolism of specific FAs and eicosanoids at various stages of fatty liver disease. These results suggest that targeting selective members of the CYP4A family is a viable therapeutic approach for treating and managing MASLD.

Quantification of pSmad western blots and endoglin konock down in endothelial cells

Endoglin is expressed by CAFs, does not correlate to stage-III CRC patient survival and TCGA database analysis

Tumor formation and angiogenesis after co-injection with WT/ENG KO MEFs

Abstract Background Differentiating high‐grade dysplasia (HGD) and T1 colorectal cancer (T1CRC) from low‐grade dysplasia (LGD) in colorectal polyps can be challenging. Incorrect recognition of HGD or T1CRC foci can lead to a need for additional treatment after local resection, which might not have been necessary if it was recognized correctly. Tumor‐targeted fluorescence‐guided endoscopy might help to improve recognition. Objective Selecting the most suitable HGD and T1CRC‐specific imaging target from a panel of well‐established biomarkers: carcinoembryonic antigen (CEA), c‐mesenchymal‐epithelial transition factor (c‐MET), epithelial cell adhesion molecule (EpCAM), folate receptor alpha (FRα), and integrin alpha‐v beta‐6 (αvβ6). Methods En bloc resection specimens of colorectal polyps harboring HGD or T1CRC were selected. Immunohistochemistry on paraffin sections was used to determine the biomarker expression in normal epithelium, LGD, HGD, and T1CRC (scores of 0–12). The differential expression in HGD‐T1CRC components compared to surrounding LGD and normal components was assessed, just as the sensitivity and specificity of each marker. Results 60 specimens were included (21 HGD, 39 T1CRC). Positive expression (score >1) of HGD‐T1CRC components was found in 73.3%, 78.3%, and 100% of cases for CEA, c‐MET, and EpCAM, respectively, and in <40% for FRα and αvβ6. Negative expression (score 0–1) of the LGD component occurred more frequently for CEA (66.1%) than c‐MET (31.6%) and EpCAM (0%). The differential expression in the HGD‐T1CRC component compared to the surrounding LGD component was found for CEA in 66.7%, for c‐MET in 43.1%, for EpCAM in 17.2%, for FRα in 22.4%, and for αvβ6 in 15.5% of the cases. Moreover, CEA showed the highest combined sensitivity (65.0%) and specificity (75.0%) for the detection of an HGD‐T1CRC component in colorectal polyps. Conclusion Of the tested targets, CEA appears the most suitable to specifically detect HGD and T1 cancer foci in colorectal polyps. An in vivo study using tumor‐targeted fluorescence‐guided endoscopy should confirm these findings.

Endoglin is not expressed on NF in vivo, CAF marker protein expression, total Smad2 and endoglin expression on mCAFs

Supplementary table 1 and 2

BACKGROUND & AIMS: Improving clinical management of early stage colorectal cancers (T1CRCs) requires a better understanding of their underlying biology. Accumulating evidence shows that cancer-associated fibroblasts (CAFs) are important determinants of tumor progression in advanced colorectal cancer (CRC), but their role in the initial stages of CRC tumorigenesis is unknown. Therefore, we investigated the contribution of T1CAFs to early CRC progression. METHODS: Primary T1CAFs and patient-matched normal fibroblasts (NFs) were isolated from endoscopic biopsy specimens of histologically confirmed T1CRCs and normal mucosa, respectively. The impact of T1CAFs and NFs on tumor behavior was studied using 3-dimensional co-culture systems with primary T1CRC organoids and extracellular matrix (ECM) remodeling assays. Whole-transcriptome sequencing and gene silencing were used to pinpoint mediators of T1CAF functions. RESULTS: In 3-dimensional multicellular cultures, matrix invasion of T1CRC organoids was induced by T1CAFs, but not by matched NFs. Enhanced T1CRC invasion was accompanied by T1CAF-induced ECM remodeling and up-regulation of CD44 in epithelial cells. RNA sequencing of 10 NF-T1CAF pairs revealed 404 differentially expressed genes, with significant enrichment for ECM-related pathways in T1CAFs. Cathepsin H, a cysteine-type protease that was specifically up-regulated in T1CAFs but not in fibroblasts from premalignant lesions or advanced CRCs, was identified as a key factor driving matrix remodeling by T1CAFs. Finally, we showed high abundance of cathepsin H-expressing T1CAFs at the invasive front of primary T1CRC sections. CONCLUSIONS: Already in the earliest stage of CRC, cancer cell invasion is promoted by CAFs via direct interactions with epithelial cancer cells and stage-specific, cathepsin H-dependent ECM remodeling. RNA sequencing data of the 10 NF-T1CAF pairs can be found under GEO accession number GSE200660.

Antibodies for immunohistochemistry, immunofluorescence and western blot.

Supplementary Figures 1-5, Tables 1-2 from Inhibition of NOTCH Signaling by Gamma Secretase Inhibitor Engages the RB Pathway and Elicits Cell Cycle Exit in T-Cell Acute Lymphoblastic Leukemia Cells

Characterization of patient-derived NFs and CAFs

T1 colorectal cancers (T1CRC) are increasingly being treated by endoscopic submucosal dissection (ESD). After ESD of a T1CRC, completion surgery is indicated in a subgroup of patients. Currently, the influence of ESD on surgical morbidity and mortality is unknown. The aim of this study was to compare 90-day morbidity and mortality of completion surgery after ESD to primary surgery. The completion surgery group consisted of suspected T1CRC patients from a multicenter prospective ESD database (2014–2020). The primary surgery group consisted of pT1CRC patients from a nationwide surgical registry (2017–2019). Patients with rectal or sigmoidal cancers were selected. Patients receiving neoadjuvant therapy were excluded. Propensity score adjustment was used to correct for confounders. In total, 411 patients were included: 54 in the completion surgery group (39 pT1, 15 pT2) and 357 in the primary surgery group with pT1CRC. Adverse event rate was 24.1% after completion surgery and 21.3% after primary surgery. After completion surgery 90-day mortality did not occur, though one patient died in the primary surgery group. After propensity score adjustment, lymph node yield did not differ significantly between the groups. Among other morbidity-related outcomes, stoma rate (OR 1.298 95%-CI 0.587-2.872, p = 0.519) and adverse event rate (OR 1.162; 95%-CI 0.570-2.370, p = 0.679) also did not differ significantly. A subgroup analysis was performed in patients undergoing rectal surgery. In this subgroup (37 completion and 136 primary surgery), these morbidity outcomes also did not differ significantly. In conclusion, this study suggests that ESD does not compromise morbidity or 90-day mortality of completion surgery.

HEK293 invasion, BMP-9 expression by CRC cells and CAFs, invasion assays