Association of gut microbiome with immune microenvironment in surgically treated colorectal cancer patients.

This study explored the relationship between faecal microbiota distribution and local or systemic immune response in patients with colorectal cancer (CRC). The study population included 114 surgically treated CRC patients. Faeces were analysed using 16S rRNA gene sequencing. The immune score in tumour microenvironment was evaluated using CD3 and CD8 immunohistochemistry. Genetic alterations, microsatellite instability status and five systemic inflammatory markers were also analysed. Thirty of 114 (26.3%) CRC patients were categorised as the 'immune type' with a high density of T-cells. The immune type CRC cases showed lower angiolymphatic invasion and longer overall survival. Of the 123 selected bacterial species, Bacteroides fragilis and Collinsella aerofaciens were prevalent in immune CRC cases, whereas Odoribacter splanchnicus and Phascolarctobacterium succinatutens were prevalent in non-immune CRC patients. Bacteroides fragilis was associated with shorter disease free survival in univariable and multivariable survival analyses. Regarding systemic immunity, a high prevalence of C. aerofaciens was associated with a high modified Glasgow prognostic score. This study revealed a potential relationship among the gut microbiome, immune microenvironment, and disease progression in patients with CRC. Our findings suggest that abundant B. fragilis in patients with CRC is associated with a 'cold immune' tumour microenvironment.

Surgical quality assessment for the prospective study of oncologic outcomes after laparoscopic modified complete mesocolic excision for nonmetastatic right colon cancer (PIONEER study).

BACKGROUND: The modified complete mesocolic excision (mCME) procedure for right-sided colon cancer is a tailored approach based on the original complete mesocolic excision (CME) methodology. Limited studies evaluated the safety and feasibility of laparoscopic mCME using objective surgical quality assessments in patients with right colon cancer. The objectives of the PIONEER study were to evaluate oncologic outcomes after laparoscopic mCME and to identify optimal clinically relevant endpoints and values for standardizing laparoscopic right colon cancer surgery based on short-term outcomes of procedures performed by expert laparoscopic surgeons. MATERIALS AND METHODS: This is an ongoing prospective, multi-institutional, single-arm study conducted at five tertiary colorectal cancer centers in South Korea. Study registrants included 250 patients scheduled for laparoscopic mCME with right-sided colon adenocarcinoma (from the appendix to the proximal half of the transverse colon). The primary endpoint was 3-year disease-free survival. Secondary outcomes included 3-year overall survival, incidence of morbidity in the first 4 weeks postoperatively, completeness of mCME, central radicality, and distribution of metastatic lymph nodes. Survival data will be available after the final follow-up date (June 2024). RESULTS: The postoperative complication rate was 12.9%, with a major complication rate of 2.7%. In 87% of patients, central radicality was achieved with dissection at or beyond the level of complete exposure of the superior mesenteric vein. Mesocolic plane resection with an intact mesocolon was achieved in 75.9% of patients, as assessed through photographs. Metastatic lymph node distribution varied by tumor location and extent. Seven optimal clinically relevant endpoints and values were identified based on the analysis of complications in low-risk patients. CONCLUSIONS: Laparoscopic mCME for right-sided colon cancer produced favorable short-term postoperative outcomes. The identified optimal clinically relevant endpoints and values can serve as a reference for evaluating surgical performance of this procedure.

Glycemic traits and colorectal cancer survival in a cohort of South Korean patients: A Mendelian randomization analysis.

BACKGROUND: Clinical diabetic traits have been reported to be associated with increased colorectal cancer (CRC) risk in observational studies. Using the Mendelian randomization (MR) analysis method, we examined the causal association between glycemic traits, such as fasting glucose (FG), fasting insulin (FI), and glycosylated hemoglobin A1c (HbA1c), and survival in a cohort of CRC patients. METHODS: We conducted a two-sample MR analysis among a cohort of patients with locally advanced CRC at Seoul National University Hospital. Single-nucleotide polymorphisms robustly associated (p < 5 × 10-8 ) with the three glycemic traits were obtained from the Meta-Analyses of Glucose and Insulin-related traits Consortium, Asian Genetic Epidemiology Network, and Korea Biobank Array. Three-year and 5-year overall survival (OS) and progression-free survival (PFS) were used as outcomes. Survival analysis was conducted using subgroup analysis by cancer stage and subsite in a multivariate Cox proportional hazards model adjusted for age and sex to examine whether glycemic traits affected survival. RESULTS: A total of 509 patients were included in our final analysis. MR analysis showed that HbA1c levels were associated with poor 3-year OS (ß = 4.20, p = 0.02). Sensitivity analyses did not show evidence of any violations of the MR assumptions. In the cancer subgroup analysis of the Cox proportional hazards model, pooled hazard ratios for FG were significantly associated with poor 3-year OS and PFS regardless of cancer stage. FI was not significantly associated with any 3-year survival endpoints. Among Stage III patients, three glycemic traits were significantly associated with both 5-year OS and PFS. Location-specific subgroup analysis showed a significant association between three glycemic traits and 5-year PFS in patients with left-sided colon cancer. FG was associated with poor 3-year survival for colon cancer but not rectal cancer. CONCLUSIONS: Our results suggest that FG and HbA1c could be used to predict prognosis in CRC patients. Lifestyle and/or pharmacological interventions targeting glycemic traits could help improve survival for CRC patients.

A multimodal virtual vision platform as a next-generation vision system for a surgical robot.

Robot-assisted surgery platforms are utilized globally thanks to their stereoscopic vision systems and enhanced functional assistance. However, the necessity of ergonomic improvement for their use by surgeons has been increased. In surgical robots, issues with chronic fatigue exist owing to the fixed posture of the conventional stereo viewer (SV) vision system. A head-mounted display was adopted to alleviate the inconvenience, and a virtual vision platform (VVP) is proposed in this study. The VVP can provide various critical data, including medical images, vital signs, and patient records, in three-dimensional virtual reality space so that users can access medical information simultaneously. An availability of the VVP was investigated based on various user evaluations by surgeons and novices, who executed the given tasks and answered questionnaires. The performances of the SV and VVP were not significantly different; however, the craniovertebral angle of the VVP was 16.35° higher on average than that of the SV. Survey results regarding the VVP were positive; participants indicated that the optimal number of displays was six, preferring the 2 × 3 array. Reflecting the tendencies, the VVP can be a neoconceptual candidate to be customized for medical use, which opens a new prospect in a next-generation surgical robot.

Assessment of Genetic Stability in Human Induced Pluripotent Stem Cell-Derived Cardiomyocytes by Using Droplet Digital PCR.

Unintended genetic modifications that occur during the differentiation and proliferation of human induced pluripotent stem cells (hiPSCs) can lead to tumorigenicity. This is a crucial concern in the development of stem cell-based therapies to ensure the safety and efficacy of the final product. Moreover, conventional genetic stability testing methods are limited by low sensitivity, which is an issue that remains unsolved. In this study, we assessed the genetic stability of hiPSCs and hiPSC-derived cardiomyocytes using various testing methods, including karyotyping, CytoScanHD chip analysis, whole-exome sequencing, and targeted sequencing. Two specific genetic mutations in KMT2C and BCOR were selected from the 17 gene variants identified by whole-exome and targeted sequencing methods, which were validated using droplet digital PCR. The applicability of this approach to stem cell-based therapeutic products was further demonstrated with associated validation according to the International Council for Harmonisation (ICH) guidelines, including specificity, precision, robustness, and limit of detection. Our droplet digital PCR results showed high sensitivity and accuracy for quantitatively detecting gene mutations, whereas conventional qPCR could not avoid false positives. In conclusion, droplet digital PCR is a highly sensitive and precise method for assessing the expression of mutations with tumorigenic potential for the development of stem cell-based therapeutics.

Risk Factor Analysis of Lymph Node Metastasis for Rectal Neuroendocrine Tumors: Who Needs a Radical Resection in Rectal Neuroendocrine Tumors Sized 1-2 cm?

BACKGROUND: Rectal neuroendocrine tumors (NETs) have malignant potential, and lymph node (LN) or distant metastases can occur; however, treatment of NETs 1-2 cm in size is controversial. OBJECTIVE: This study aimed to identify predictive factors for LN metastasis and prognostic factors for recurrence of rectal NETs, especially tumors 1‒2 cm in size. METHODS: Between October 2004 and November 2020, 453 patients underwent endoscopic or surgical treatment for rectal NETs in Seoul National University Hospital. The data on these patients were prospectively collected in our database and reviewed retrospectively. In cases of local excision, we evaluated LN metastasis with radiologic imaging, including computed tomography or magnetic resonance imaging before treatment and during the follow-up periods. RESULTS: LN metastasis was observed in 40 patients (8.8%). A higher rate of LN metastasis was observed in larger-sized tumors, advanced T stage, lymphovascular invasion (LVI), perineural invasion (PNI), and high tumor grade. In multivariable analysis, the significant risk factors for LN metastasis were tumor size (1 ≤ size < 2 cm: hazard ratio [HR] 64.07; size ≥2 cm: HR 102.37, p < 0.001) and tumor grade (G2: HR 3.63, p = 0.034; G3: HR 5.09, p = 0.044). In multivariable analysis for tumors 1-2 cm in size, the risk factor for LN metastasis was tumor grade (G2: HR 6.34, p = 0.013). CONCLUSIONS: Tumor grade and size are important predictive factors for LN metastasis. In NETs 2 cm in size, tumor grade is also important for LN metastasis, and radical resection should be considered.

Three-dimensional body composition parameters using automatic volumetric segmentation allow accurate prediction of colorectal cancer outcomes.

BACKGROUND: Parameters obtained from two-dimensional (2D) cross-sectional images have been used to determine body composition. However, data from three-dimensional (3D) volumetric body images reflect real body composition more accurately and may be better predictors of patient outcomes in cancer. This study aimed to assess the 3D parameters and determine the best predictive factors for patient prognosis. METHODS: Patients who underwent surgery for colorectal cancer (CRC) between 2010 and 2016 were included in this study. Preoperative computed tomography images were analysed using an automatic segmentation program. Body composition parameters for muscle, muscle adiposity, subcutaneous fat (SF) and abdominal visceral fat (AVF) were assessed using 2D images at the third lumbar (L3) level and 3D images of the abdominal waist (L1-L5). The cut-off points for each parameter were determined using X-tile software. A Cox proportional hazards regression model was used to identify the association between the parameters and the treatment outcomes, and the relative influence of each parameter was compared using a gradient boosting model. RESULTS: Overall, 499 patients were included in the study. At a median follow-up of 59 months, higher 3D parameters of the abdominal muscles and SF from the abdominal waist were found to be associated with longer overall survival (OS) and disease-free survival (all P < 0.001). Although the 3D parameters of AVF were not related to survival outcomes, patients with a high AVF volume and mass experienced higher rate of postoperative complications than those with low AVF volume (27.4% vs. 18.7%, P = 0.021, for mass; 27.1% vs. 19.0%, P = 0.028, for volume). Low muscle mass and volume (hazard ratio [HR] 1.959, P = 0.016; HR 2.093, P = 0.036, respectively) and low SF mass and volume (HR 1.968, P = 0.008; HR 2.561, P = 0.003, respectively), both in the abdominal waist, were identified as independent prognostic factors for worse OS. Along with muscle mass and volume, SF mass and volume in the abdominal waist were negatively correlated with mortality (all P < 0.001). Both AVF mass and volume in the abdominal waist were positively correlated with postoperative complications (P < 0.05); 3D muscle volume and SF at the abdominal waist were the most influential factors for OS. CONCLUSIONS: 3D volumetric parameters generated using an automatic segmentation program showed higher correlations with the short- and long-term outcomes of patients with CRC than conventional 2D parameters.

Exploring the Functional Heterogeneity of Directly Reprogrammed Neural Stem Cell-Derived Neurons via Single-Cell RNA Sequencing.

The therapeutic potential of directly reprogrammed neural stem cells (iNSCs) for neurodegenerative diseases relies on reducing the innate tumorigenicity of pluripotent stem cells. However, the heterogeneity within iNSCs is a major hurdle in quality control prior to clinical applications. Herein, we generated iNSCs from human fibroblasts, by transfecting transcription factors using Sendai virus particles, and characterized the expression of iNSC markers. Using immunostaining and quantitative real time -polymerase chain reaction (RT -qPCR), no differences were observed between colonies of iNSCs and iNSC-derived neurons. Unexpectedly, patch-clamp analysis of iNSC-derived neurons revealed distinctive action potential firing even within the same batch product. We performed single-cell RNA sequencing in fibroblasts, iNSCs, and iNSC-derived neurons to dissect their functional heterogeneity and identify cell fate regulators during direct reprogramming followed by neuronal differentiation. Pseudotime trajectory analysis revealed distinct cell types depending on their gene expression profiles. Differential gene expression analysis showed distinct NEUROG1, PEG3, and STMN2 expression patterns in iNSCs and iNSC-derived neurons. Taken together, we recommend performing a predictable functional assessment with appropriate surrogate markers to ensure the quality control of iNSCs and their differentiated neurons, particularly before cell banking for regenerative cell therapy.

Aromadendrin inhibits PMA-induced cytokine formation/NF-κB activation in A549 cells and ovalbumin-induced bronchial inflammation in mice.

Hyperproduction of immune cell-derived inflammatory molecules and recruitment of immune cells promote the development of allergic asthma (AA). Aromadendrin (ARO) has various biological properties including anti-inflammatory effects. In this study, we evaluated the ameliorative effects of ARO on the development of AA in vitro and in vivo. Phorbol 12-myristate 13-acetate (PMA, 100 nM) was used to induce inflammation in A549 airway epithelial cells. The cohesion of A549 and eosinophil EOL-1 cells was studied. Ovalbumin (30 or 60 µg)/Alum (3 mg) mixture was adapted for AA induction in mice. ARO (5 or 10 mg/kg, p. o.) was administered to mice to investigate its ameliorative effect on AA development. Enzyme-linked immunosorbent assay, western blotting, and hematoxylin and eosin/periodic acid Schiff staining were performed to study the ameliorative effect of ARO on bronchial inflammation. In PMA-stimulated A549 cells, the upregulation of cytokines (interleukin [IL]-1ß/IL-6/tumor necrosis factor alpha [TNF-α]/monocyte chemoattractant protein [MCP]-1]) and nuclear factor kappa B (NF-κB) activation was effectively reduced by ARO pretreatment. ARO suppressed the adhesion of A549 cells and eosinophils. In ovalbumin-induced AA mice, the levels of cells, such as eosinophils, Th2 cytokines, MCP-1 in bronchoalveolar lavage fluid, IgE in serum, and inducible nitric oxide synthase/cyclooxygenase-2 expression in the lung tissue were upregulated, which were all suppressed by ARO. In addition, the increase in cell inflow and mucus formation in the lungs of AA mice was reversed by ARO as per histological analysis. ARO also modulated NF-κB activation in the lungs of AA mice. Overall, the anti-inflammatory properties of ARO in vitro/in vivo studies of AA were notable. Thus, ARO has a modulatory effect on bronchial inflammation and may be a potential adjuvant for AA treatment.

Cystic lymphangioma causing intussusception of the right colon: a case report and review of current literature.

Among intraabdominal lymphangiomas, colonic lymphangiomas are rare. These cystic tumors are generally asymptomatic and incidentally found but may present with bleeding or obstructive symptoms. Intussusception by such tumors is scarcely reported, with only nine previously reported cases listed in Pubmed. We report a case of a 41-year-old female Asian patient who presented with acute abdomen and was diagnosed with colonic intussusception caused by lymphangioma. She received emergent right hemicolectomy, recovered well without complications, and was discharged on the 5th postoperative day.

Antioxidant and Anti-Inflammatory Mechanisms of Lipophilic Fractions from Polyscias fruticosa Leaves Based on Network Pharmacology, In Silico, and In Vitro Approaches.

Polyscias fruticosa leaf (PFL) has been used in food and traditional medicine for the treatment of rheumatism, ischemia, and neuralgia. However, the lipophilic components of PFL and their biological properties remain unknown. This study, integrating network pharmacology analysis with in silico and in vitro approaches, aimed to elucidate the antioxidant and anti-inflammatory capacities of lipophilic extracts from PFL. A total of 71 lipophilic compounds were identified in PFL using gas chromatography-mass spectrometry. Network pharmacology and molecular docking analyses showed that key active compounds, mainly phytosterols and sesquiterpenes, were responsible for regulating core target genes, such as PTGS2, TLR4, NFE2L2, PRKCD, KEAP1, NFKB1, NR1l2, PTGS1, AR, and CYP3A4, which were mostly enriched in oxidative stress and inflammation-related pathways. Furthermore, lipophilic extracts from PFL offered powerful antioxidant capacities, as evident in our cell-free antioxidant assays. These extracts also provided a protection against oxidative stress by inducing the expression of catalase and heme oxygenase-1 in lipopolysaccharide (LPS)-treated RAW 264.7 cells. Additionally, lipophilic fractions from PFL showed anti-inflammatory potential in downregulating the level of pro-inflammatory factors in LPS-treated macrophages. Overall, these findings provide valuable insights into the antioxidant and anti-inflammatory properties of lipophilic extracts from PFL, which can be used as a fundamental basis for developing nutraceuticals and functional foods.

A 3 mm Port Reduces Postoperative Pain After Laparoscopic Colon Cancer Surgery: A Case-control Matched Study.

BACKGROUND: Recently, smaller-size trocars and instruments have been developed for laparoscopic colon cancer surgery; however, their effectiveness and safety have not been elucidated. This study aimed to investigate whether 3 mm trocars and instruments have benefits compared with conventional trocars and instruments. PATIENTS AND METHODS: Patients with colon cancer who underwent laparoscopic anterior resection or right hemicolectomy were included. Patients who underwent combined resections of other organs and those with conversion to open surgery were excluded. In the 3 mm group, three 5 mm trocars were replaced by 3 mm trocars. The numeric rating scale (NRS) immediately postoperatively at 24, 48, and 72 hours, respectively, after surgery and the use of additional analgesics and perioperative outcomes were analyzed. Case-control matched analysis was used to reduce bias according to the type of surgery. RESULTS: A total of 207 patients (conventional: n = 158, 3 mm: n = 49) were included. Before matching, NRS 48 hours postoperatively ( P = 0.049), proportion of patients using additional intravenous (IV) analgesics ( P = 0.007), postoperative hospital stay ( P < 0.001), and blood loss ( P < 0.001) were lower in the 3 mm group. In multivariable analysis, trocar type significantly impacted the proportion of patients using additional IV analgesics (odds ratio: 0.330; 95% CI: 0.153-0.712; P = 0.005). After case-control matching, NRS immediately postoperatively ( P = 0.015) and 24 hours postsurgery ( P = 0.043), patients using additional IV analgesics ( P = 0.019), postoperative hospital stay ( P = 0.010), intraoperative blood loss ( P < 0.001), and postoperative complication rate ( P = 0.028) were significantly lower in the 3 mm group compared with the 5 mm group. CONCLUSIONS: The use of 3 mm trocars and instruments in laparoscopic colon cancer surgery can effectively reduce postoperative pain while maintaining perioperative safety.

Clinical Usefulness of Immune Profiling for Differential Diagnosis between Crohn's Disease, Intestinal Tuberculosis, and Behcet's Disease.

It is important to make a differential diagnosis between inflammatory diseases of the bowel with similar clinical and endoscopic features. The profiling of immune cells could be helpful for accurately diagnosing inflammatory bowel diseases. We compared immune marker expression between Crohn's disease (CD), intestinal Behcet's disease (BD), and intestinal tuberculosis (TB) and evaluated the usefulness of immune profiling in differentiating between these diseases. Biopsy specimens were acquired around ulcerations on the terminal ileum or cecum from five patients with each disease. Panel 1 included multiplex immunohistochemistry staining for CD8, CD4, Foxp3, CD20, programmed death-1, and granzyme B. CD56, CD68, CD163, CD11c, and HLA-DR were analyzed in panel 2. The differences in cytotoxic T cells (CD8+CD4-Fopx3-CD20-), helper T cells (CD8-CD4+Fopx3-CD20-), and regulatory T cells (CD8-CD4+Fopx3+CD20-) were also not significant. However, M1 macrophage (CD68+CD163-HLA-DR-) cell densities were significantly higher in intestinal BD than in other diseases. The expression level of dendritic cells (CD56-CD68-CD163-CD11c+HLA-DR+) was highest in intestinal TB and lowest in intestinal BD. The expression of immune cells, including M1 macrophages and dendritic cells, was different between CD, intestinal BD, and intestinal TB. Immune profiling can be helpful for establishing differential diagnoses of inflammatory bowel diseases.

Prognosis of Patients with Chronic Hepatitis C Genotype 1b Infection Treated Using Daclatasvir/Asunaprevir after Sustained Virologic Response: A 6-Year Multicenter Prospective Observational Study.

Aim and Objectives: Direct-acting antiviral (DAA) therapy can cure chronic hepatitis C (CHC), and daclatasvir (DCV)/asunaprevir (ASV) was the first interferon-free DAA therapy introduced in Korea. Patients who achieve sustained virologic response (SVR) after DAA treatment are expected to have good prognoses. Therefore, in this study, we aimed to investigate the prognosis of these patients. Materials and Methods: This multicenter prospective observational study included patients with CHC who achieved SVR after DCV/ASV treatment. The primary endpoint was hepatocellular carcinoma (HCC) occurrence, which was reviewed annually. Results: We included 302 patients (median follow-up duration: 38 [16.5-60.0] months; median age: 58 [49-67] years) in the study. Cirrhosis was observed in 103 patients (34.1%), and the median Child-Pugh score was 5.0. HCC occurred in 16 patients (5.3%) within six years post-SVR; these patients were older and had higher cirrhosis prevalence, alpha-fetoprotein levels, and fibrosis-4 index scores than did those without HCC development. Cox proportional hazards analysis revealed that age > 71 years (p = 0.005) and cirrhosis (p = 0.035) were significant risk factors for HCC occurrence. Conclusions: Although the prognoses of patients who achieved SVR with DCV/ASV therapy were generally good, the risk for HCC was present, especially in older patients and in those with cirrhosis. Hence, early treatment at younger ages and regular follow-up surveillance after achieving SVR are warranted.

Total neoadjuvant therapy with short-course radiotherapy Versus long-course neoadjuvant chemoradiotherapy in Locally Advanced Rectal cancer, Korean trial (TV-LARK trial): study protocol of a multicentre randomized controlled trial.

BACKGROUND: For locally advanced rectal cancer (LARC), total neoadjuvant therapy (TNT) may enhance tumour response, reduce recurrence, and improve patient compliance compared to upfront surgery. Recent studies have shown that chemoradiotherapy (CRT) followed by consolidation chemotherapy leads to higher rate of pathologic complete response (pCR) than induction chemotherapy followed by CRT. However, an optimal TNT regimen that maximise the pCR rate and minimise toxicity has not been established. Therefore, the aim of this trial was to investigate whether preoperative short-course radiotherapy followed by chemotherapy with four cycles of CAPOX can double the pCR rate compared to a standard schedule of long-course preoperative CRT in patients with LARC. METHODS: This is a multi-centre, prospective, open label, randomised controlled trial. Patients with clinical primary tumour stage 3 and higher or regional node-involved rectal cancer located within 10 cm from the anal verge were randomly assigned equally to short-course radiotherapy (25 Gy in 5 fractions over 1 week) followed by four cycles of CAPOX (intravenous oxaliplatin [130 mg/m2, once a day] on day 1 and capecitabine [1,000 mg/m2, twice a day] from days 1 to 14) (TNT) or CRT (50.4 Gy in 28 fractions over 5 weeks, concurrently with concomitant oral capecitabine 825 mg/m2 twice a day). After preoperative treatment, total mesorectal excision was performed 2-4 weeks in the TNT group and 6-10 weeks in the CRT group, followed by optional additional adjuvant chemotherapy. The primary endpoint is the pCR rate, and secondary endpoints include disease-related treatment failure, quality of life, and cost-effectiveness. Assuming a pCR rate of 28% and 15% in the TNT and CRT groups, respectively, and one-side alpha error rate of 0.025 and power of 80%, 348 patients will be enrolled considering 10% dropout rate. DISCUSSION: The TV-LARK trial will evaluate the superiority of employed TNT regimen against the standard CRT regimen for patients with LARC. We aimed to identify a TNT regimen that will improve the pCR rate and decrease systemic recurrence in these patients. TRIAL REGISTRATION: Cris.nih.go.kr ID: KCT0007169 (April 08, 2022). The posted information will be updated as needed to reflect the protocol amendments and study progress.

Peroxiredoxin V Protects against UVB-Induced Damage of Keratinocytes.

Ultraviolet B (UVB) irradiation generates reactive oxygen species (ROS), which can damage exposed skin cells. Mitochondria and NADPH oxidase are the two principal producers of ROS in UVB-irradiated keratinocytes. Peroxiredoxin V (PrxV) is a mitochondrial and cytosolic cysteine-dependent peroxidase enzyme that robustly removes H2O2. We investigated PrxV's role in protecting epidermal keratinocytes against UVB-induced ROS damage. We separated mitochondrial and cytosolic H2O2 levels from other types of ROS using fluorescent H2O2 indicators. Upon UVB irradiation, PrxV-knockdown HaCaT human keratinocytes showed higher levels of mitochondrial and cytosolic H2O2 than PrxV-expressing controls. PrxV depletion enhanced hyperoxidation-mediated inactivation of mitochondrial PrxIII and cytosolic PrxI and PrxII in UVB-irradiated keratinocytes. PrxV-depleted keratinocytes exhibited mitochondrial dysfunction and were more susceptible to apoptosis through decreased oxygen consumption rate, loss of mitochondrial membrane potential, cardiolipin oxidation, cytochrome C release, and caspase activation. Our findings show that PrxV serves to protect epidermal keratinocytes from UVB-induced damage such as mitochondrial dysfunction and apoptosis, not only by directly removing mitochondrial and cytosolic H2O2 but also by indirectly improving the catalytic activity of mitochondrial PrxIII and cytosolic PrxI and PrxII. It is possible that strengthening PrxV defenses could aid in preventing UVB-induced skin damage.

Predicting heterotrophic plate count exceedance in tap water: A binary classification model supervised by culture-independent data.

Culture-independent data can be utilized to identify heterotrophic plate count (HPC) exceedances in drinking water. Although HPC represents less than 1% of the bacterial community and exhibits time lags of several days, HPC data are widely used to assess the microbiological quality of drinking water and are incorporated into drinking water standards. The present study confirmed the nonlinear relationships between HPC, intact cell count (ICC), and adenosine triphosphate (ATP) in tap water samples (stagnant and flushed). By using a combination of ICC, ATP, and free chlorine data as inputs, we show that HPC exceedance can be classified using a 2-layer feed-forward artificial neural network (ANN). Despite the nonlinearity of HPC, the best binary classification model showed accuracies of 95%, sensitivity of 91%, and specificity of 96%. ICC and chlorine concentrations were the most important features for classifiers. The main limitations, such as sample size and class imbalance, were also discussed. The present model provides the ability to convert data from emerging measurement techniques into established and well-understood measures, overcoming culture dependence and offering near real-time data to help ensure the biostability and safety of drinking water.

Personalised circulating tumour DNA assay with large-scale mutation coverage for sensitive minimal residual disease detection in colorectal cancer.

BACKGROUND: Postoperative minimal residual disease (MRD) detection using circulating-tumour DNA (ctDNA) requires a highly sensitive analysis platform. We have developed a tumour-informed, hybrid-capture ctDNA sequencing MRD assay. METHODS: Personalised target-capture panels for ctDNA detection were designed using individual variants identified in tumour whole-exome sequencing of each patient. MRD status was determined using ultra-high-depth sequencing data of plasma cell-free DNA. The MRD positivity and its association with clinical outcome were analysed in Stage II or III colorectal cancer (CRC). RESULTS: In 98 CRC patients, personalised panels for ctDNA sequencing were built from tumour data, including a median of 185 variants per patient. In silico simulation showed that increasing the number of target variants increases MRD detection sensitivity in low fractions (<0.01%). At postoperative 3-week, 21.4% of patients were positive for MRD by ctDNA. Postoperative positive MRD was strongly associated with poor disease-free survival (DFS) (adjusted hazard ratio 8.40, 95% confidence interval 3.49-20.2). Patients with a negative conversion of MRD after adjuvant therapy showed significantly better DFS (P < 0.001). CONCLUSION: Tumour-informed, hybrid-capture-based ctDNA assay monitoring a large number of patient-specific mutations is a sensitive strategy for MRD detection to predict recurrence in CRC.

Gut microbiome associated with low anterior resection syndrome after rectal cancer surgery.

This study aimed to assess the likely association of gut microbiome with low anterior resection syndrome (LARS) symptoms. Postoperative stool samples from patients with minor or major LARS after sphincter-preserving surgery (SPS) for rectal cancer were collected and analyzed using 16S ribosomal RNA sequencing method. The symptom patterns of LARS were classified into two groups (PC1LARS, PC2LARS) using principal component analysis. The dichotomized sum of questionnaire items (sub1LARS, sub2LARS) was used to group patients according to the main symptoms. According to microbial diversity, enterotype, and taxa, PC1LARS and sub1LARS were associated with frequency-dominant LARS symptoms and patients, while PC2LARS and sub2LARS were grouped as incontinence-dominant LARS symptoms and patients. Butyricicoccus levels decreased while overall LARS scores increased. The α-diversity richness index Chao1 showed a significantly negative correlation in sub1LARS and a positive correlation in sub2LARS. In sub1LARS, the severe group showed a lower Prevotellaceae enterotype and higher Bacteroidaceae enterotype than the mild group. Subdoligranulum and Flavonifractor showed a negative and a positive correlation with PC1LARS, respectively, while showing a negative relationship with PC2LARS. Lactobacillus and Bifidobacterium were negatively correlated to PC1LARS. Frequency-dominant LARS had decreased diversity of gut microbiome and showed lower levels of lactic acid-producing bacteria.

Widespread somatic L1 retrotransposition in normal colorectal epithelium.

Throughout an individual's lifetime, genomic alterations accumulate in somatic cells1-11. However, the mutational landscape induced by retrotransposition of long interspersed nuclear element-1 (L1), a widespread mobile element in the human genome12-14, is poorly understood in normal cells. Here we explored the whole-genome sequences of 899 single-cell clones established from three different cell types collected from 28 individuals. We identified 1,708 somatic L1 retrotransposition events that were enriched in colorectal epithelium and showed a positive relationship with age. Fingerprinting of source elements showed 34 retrotransposition-competent L1s. Multidimensional analysis demonstrated that (1) somatic L1 retrotranspositions occur from early embryogenesis at a substantial rate, (2) epigenetic on/off of a source element is preferentially determined in the early organogenesis stage, (3) retrotransposition-competent L1s with a lower population allele frequency have higher retrotransposition activity and (4) only a small fraction of L1 transcripts in the cytoplasm are finally retrotransposed in somatic cells. Analysis of matched cancers further suggested that somatic L1 retrotransposition rate is substantially increased during colorectal tumourigenesis. In summary, this study illustrates L1 retrotransposition-induced somatic mosaicism in normal cells and provides insights into the genomic and epigenomic regulation of transposable elements over the human lifetime.