Colorectal Cancer Stem Cells Fuse with Monocytes to Form Tumour Hybrid Cells with the Ability to Migrate and Evade the Immune System.

BACKGROUND: The cancer cell fusion theory could be one of the best explanations for the metastasis from primary tumours. METHODS: Herein, we co-cultured colorectal cancer (CRC) stem cells with human monocytes and analysed the properties of the generated tumour hybrid cells (THCs). The presence of THCs in the bloodstream together with samples from primary and metastatic lesions and their clinical correlations were evaluated in CRC patients and were detected by both FACS and immunofluorescence methods. Additionally, the role of SIGLEC5 as an immune evasion molecule in colorectal cancer was evaluated. RESULTS: Our data demonstrated the generation of THCs after the in vitro co-culture of CRC stem cells and monocytes. These cells, defined as CD45+CD14+EpCAM+, showed enhanced migratory and proliferative abilities. The THC-specific cell surface signature allows identification in matched primary tumour tissues and metastases as well as in the bloodstream from patients with CRC, thus functioning as a biomarker. Moreover, SIG-LEC5 expression on in vitro generated THCs has shown to be involved in the mechanism for immune evasion. Additionally, sSIGLEC5 levels correlated with THC numbers in the prospective cohort of patients. CONCLUSIONS: Our results indicate the generation of a hybrid entity after the in vitro co-culture between CRC stem cells and human monocytes. Moreover, THC numbers present in patients are related to both prognosis and the later spread of metastases in CRC patients.

Intertwined leukocyte balances in tumours and peripheral blood as robust predictors of right and left colorectal cancer survival.

BACKGROUND: Colorectal cancer (CRC) accounts for 9.4% of overall cancer deaths, ranking second after lung cancer. Despite the large number of factors tested to predict their outcome, most patients with similar variables show big differences in survival. Moreover, right-sided CRC (RCRC) and left-sided CRC (LCRC) patients exhibit large differences in outcome after surgical intervention as assessed by preoperative blood leukocyte status. We hypothesised that stronger indexes than circulating (blood) leukocyte ratios to predict RCRC and LCRC patient outcomes will result from combining both circulating and infiltrated (tumour/peritumour fixed tissues) concentrations of leukocytes. AIM: To seek variables involving leukocyte balances in peripheral blood and tumour tissues and to predict the outcome of CRC patients. METHODS: Sixty-five patients diagnosed with colon adenocarcinoma by the Digestive Surgery Service of the La Paz University Hospital (Madrid, Spain) were enrolled in this study: 43 with RCRC and 22 with LCRC. Patients were followed-up from January 2017 to March 2021 to record overall survival (OS) and recurrence-free survival (RFS) after surgical interventions. Leukocyte concentrations in peripheral blood were determined by routine laboratory protocols. Paraffin-fixed samples of tumour and peritumoural tissues were assessed for leukocyte concentrations by immunohistochemical detection of CD4, CD8, and CD14 marker expression. Ratios of leukocyte concentration in blood and tissues were calculated and evaluated for their predictor values for OS and RFS with Spearman correlations and Cox univariate and multivariate proportional hazards regression, followed by the calculation of the receiver-operating characteristic and area under the curve (AUC) and the determination of Youden's optimal cutoff values for those variables that significantly correlated with either RCRC or LCRC patient outcomes. RCRC patients from the cohort were randomly assigned to modelling and validation sets, and clinician-friendly nomograms were developed to predict OS and RFS from the respective significant indexes. The accuracy of the model was evaluated using calibration and validation plots. RESULTS: The relationship of leukocyte ratios in blood and peritumour resulted in six robust predictors of worse OS in RCRC: CD8+ lymphocyte content in peritumour (CD8pt, AUC = 0.585, cutoff < 8.250, P = 0.0077); total lymphocyte content in peritumour (CD4CD8pt, AUC = 0.550, cutoff < 10.160, P = 0.0188); lymphocyte-to-monocyte ratio in peritumour (LMRpt, AUC = 0.807, cutoff < 3.185, P = 0.0028); CD8+ LMR in peritumour (CD8MRpt, AUC = 0.757, cutoff < 1.650, P = 0.0007); the ratio of blood LMR to LMR in peritumour (LMRb/LMRpt, AUC = 0.672, cutoff > 0.985, P = 0.0244); and the ratio of blood LMR to CD8+ LMR in peritumour (LMRb/CD8MRpt, AUC = 0.601, cutoff > 1.485, P = 0.0101). In addition, three robust predictors of worse RFS in RCRC were found: LMRpt (AUC = 0.737, cutoff < 3.185, P = 0.0046); LMRb/LMRpt (AUC = 0.678, cutoff > 0.985, P = 0.0155) and LMRb/CD8MRpt (AUC = 0.615, cutoff > 1.485, P = 0.0141). Furthermore, the ratio of blood LMR to CD4+ LMR in peritumour (LMRb/CD4MRpt, AUC = 0.786, cutoff > 10.570, P = 0.0416) was found to robustly predict poorer OS in LCRC patients. The nomograms showed moderate accuracy in predicting OS and RFS in RCRC patients, with concordance index of 0.600 and 0.605, respectively. CONCLUSION: Easily obtainable variables at preoperative consultation, defining the status of leukocyte balances between peripheral blood and peritumoural tissues, are robust predictors for OS and RFS of both RCRC and LCRC patients.

Implementation of an enhanced recovery after surgery program with robotic surgery in high-risk patients obtains optimal results after colorectal resections.

Enhanced recovery after surgery programs reduce postoperative complications and length of stay after laparoscopic colorectal surgery, but are still under evaluation after robotic colorectal surgery. To evaluate potential benefits in terms of length of stay and complications of an Enhanced recovery after surgery program in colorectal surgery. A subanalysis was performed to assess what combination of surgical approach and perioperative care had better outcomes. Prospective observational cohort study. 300 consecutive colorectal surgery patients: 150 were prospectively included in the enhanced recovery after Surgery program group and 150 retrospectively in the traditional care group, and subdivided according to the type of surgery, in Hospital Marques de Valdecilla, between 2013 and 2016. Postoperative complications decreased significantly (p = 0.002) from 46 to 28% (traditional care vs program group). The length of stay was decreased by 2 days (p < 0.001). Multivariate analysis indicated similar effect sizes after adjusting for age, gender, Charlson score, and type of surgery. Type of surgery was an independent predictive factor for postoperative complications and length of stay. Compared to open surgery, postoperative complications decreased by 50% (p < 0.001) after robotic surgery and by 40% (p = 0.01) after laparoscopic surgery, while the median length of stay decreased by three days (p < 0.001) after minimally invasive surgery. Enhanced recovery after surgery program and minimally invasive surgery were associated with decreased morbidity and length of stay after colorectal surgery compared to open surgery and traditional care. An enhanced recovery after surgery program with robotic surgery in high-risk patients might be beneficial.

Soluble SIGLEC5: A New Prognosis Marker in Colorectal Cancer Patients.

Colorectal cancer (CRC) is the second most deadly and third most commonly diagnosed cancer worldwide. There is significant heterogeneity among patients with CRC, which hinders the search for a standard approach for the detection of this disease. Therefore, the identification of robust prognostic markers for patients with CRC represents an urgent clinical need. In search of such biomarkers, a total of 114 patients with colorectal cancer and 67 healthy participants were studied. Soluble SIGLEC5 (sSIGLEC5) levels were higher in plasma from patients with CRC compared with healthy volunteers. Additionally, sSIGLEC5 levels were higher in exitus than in survivors, and the receiver operating characteristic curve analysis revealed sSIGLEC5 to be an exitus predictor (area under the curve 0.853; cut-off > 412.6 ng/mL) in these patients. A Kaplan-Meier analysis showed that patients with high levels of sSIGLEC5 had significantly shorter overall survival (hazard ratio 15.68; 95% CI 4.571-53.81; p ≤ 0.0001) than those with lower sSIGLEC5 levels. Our study suggests that sSIGLEC5 is a soluble prognosis marker and exitus predictor in CRC.

Surgical Wound Complications after Colorectal Surgery with Single-Use Negative-Pressure Wound Therapy Versus Surgical Dressing over Closed Incisions: A Randomized Controlled Trial.

BACKGROUND: Global studies indicate that surgical site infections (SSIs) are a major healthcare challenge within hospitals and can have a profound impact on patient quality of life and healthcare costs. Closed-incision negative-pressure therapy (ciNPT) has been reported to provide positive clinical benefits for patients with various incisions, including those following colorectal surgeries. METHODS: Investigators performed a prospective, randomized, multicenter trial to evaluate complications of surgical incisions in patients who received a ciNPT dressing versus a conventional surgical dressing (control) over their closed incision following colorectal surgery. The incidence of SSI was determined at 7, 15, and 30 days postsurgery. RESULTS: A total of 148 patients participated in the study. Results showed that the SSI rate on day 7 was lower in the ciNPT group versus the control group (10/75 [13.3%] vs 17/73 [23.3%]), but this difference was not statistically significant. On day 15, the SSI rate was 12/75 (16.0%) in the ciNPT group versus 21/73 (28.8%) in the control group; however, this difference was only marginally statistically significant (P = .0621). At 1 month, the SSI rate remained lower in the ciNPT group (13/75 [17.3%] vs 21/73 [28.8%], P = .0983) compared with the control group. CONCLUSIONS: Future studies with larger population sizes are necessary to determine the impact of ciNPT on patients' incisions after colorectal surgery.

Tumor stem cells fuse with monocytes to form highly invasive tumor-hybrid cells.

The 'cancer cell fusion' theory is controversial due to the lack of methods available to identify hybrid cells and to follow the phenomenon in patients. However, it seems to be one of the best explanations for both the origin and metastasis of primary tumors. Herein, we co-cultured lung cancer stem cells with human monocytes and analyzed the dynamics and properties of tumor-hybrid cells (THC), as well as the molecular mechanisms beneath this fusion process by several techniques: electron-microscopy, karyotyping, CRISPR-Cas9, RNA-seq, immunostaining, signaling blockage, among others. Moreover, mice models were assessed for in vivo characterization of hybrids colonization and invasiveness. Then, the presence of THCs in bloodstream and samples from primary and metastatic lesions were detected by FACS and immunofluorescence protocols, and their correlations with TNM stages established. Our data indicate that the generation of THCs depends on the expression of CD36 on tumor stem cells and the oxidative state and polarization of monocytes, the latter being strongly influenced by microenvironmental fluctuations. Highly oxidized M2-like monocytes show the strongest affinity to fuse with tumor stem cells. THCs are able to proliferate, colonize and invade organs. THC-specific cell surface signature CD36+CD14+PANK+ allows identifying them in matched primary tumor tissues and metastases as well as in bloodstream from patients with lung cancer, thus functioning as a biomarker. THCs levels in circulation correlate with TNM classification. Our results suggest that THCs are involved in both origin and spread of metastatic cells. Furthermore, they might set the bases for future therapies to avoid or eradicate lung cancer metastasis.

PD-L1/PD-1 crosstalk in colorectal cancer: are we targeting the right cells?

BACKGROUND: The analysis of tumour-infiltrating immune cells within patients' tumour samples in colorectal cancer (CRC) has become an independent predictor of patient survival. The tumour microenvironment and the immune checkpoints, such as PD-L1/PD-1, are relevant to the prognoses and also appear to be relevant for further CRC therapies. METHODS: We analysed the presence and features of the infiltrated monocyte/macrophage and lymphocyte populations in both tumour and peritumour samples from patients with CRC (n = 15). RESULTS: We detected a large number of CD14+ monocytes/macrophages with an alternative phenotype (CD64+CD163+) and CD4+ lymphocytes that infiltrated the tumour, but not the peritumour area. The monocytes/macrophages expressed PD-L1, whereas the lymphocytes were PD-1+; however, we did not find high PD-L1 levels in the tumour cells. Coculture of circulating naïve human monocytes/macrophages and lymphocytes with tumour cells from patients with proficient mismatch repair CRC induced both an alternative phenotype with higher expression of PD-L1 in CD14+ cells and the T-cell exhaustion phenomenon. The addition of an α-PD-1 antibody restored lymphocyte proliferation. CONCLUSION: These results emphasise the interesting nature of immune checkpoint shifting therapies, which have potential clinical applications in the context of colorectal cancer.

Fístula enteroatmosférica y uso del VAC: a propósito de un caso / Enteroatmospheric fistula and vaccum assited closure: report of a case

Las fístulas gastrointestinales asociadas con abdomen abierto posterior a cirugía abdominal mayor son una complicación grave. El manejo es extremadamente difícil y la mortalidad bastante alta a pesar de los modernos avances médicos. Aquellos pacientes que sobreviven al daño metabólico y fisiopatológico inicial, requieren en su mayoría cierre quirúrgico de la fístula lo cual es técnicamente complejo. Presentamos el caso de un paciente con una neoplasia de rectosigma que se abordó por laparoscopia y desarrolló una fístula enteroatmosférica sobre la incisión de Pfannestiel que se utilizó para la extracción de la pieza. Conclusión: El cierre asistido por vacío artesanal y el manejo nutricional adecuado permiten la mejoría en pacientes con fístulas complejas logrando las condiciones adecuadas para el cierre definitivo.

The gastrointestinal fistula associated to posterior open abdominal trauma or abdominal surgery implies severe complications. The handling of these cases is extremely hard and mortality is very high despite medical advances. Those patients who survive the initial metabolic and phisycopathological damages require, on most cases, a surgical closure of the fistula which is a very complex procedure technically. We describe the case of a patient with a rectosigmoid neoplasm that was addressed laparoscopically and enteroatmospheric fistula just developing on phannestiel incision was used to extract the neoplasm. Conclusion: The closure assisted by “hand made vacuum” and adequate nourishment allows recovering patients with complex fistulas to achieve adequate conditions for definitive closure.

Fístula enteroatmosférica y uso del VAC: a propósito de un caso / Enteroatmospheric fistula and vaccum assited closure: report of a case

Las fístulas gastrointestinales asociadas con abdomen abierto posterior a cirugía abdominal mayor son una complicación grave. El manejo es extremadamente difícil y la mortalidad bastante alta a pesar de los modernos avances médicos. Aquellos pacientes que sobreviven al daño metabólico y fisiopatológico inicial, requieren en su mayoría cierre quirúrgico de la fístula lo cual es técnicamente complejo. Presentamos el caso de un paciente con una neoplasia de rectosigma que se abordó por laparoscopia y desarrolló una fístula enteroatmosférica sobre la incisión de Pfannestiel que se utilizó para la extracción de la pieza. Conclusión: El cierre asistido por vacío artesanal y el manejo nutricional adecuado permiten la mejoría en pacientes con fístulas complejas logrando las condiciones adecuadas para el cierre definitivo. (AU)

The gastrointestinal fistula associated to posterior open abdominal trauma or abdominal surgery implies severe complications. The handling of these cases is extremely hard and mortality is very high despite medical advances. Those patients who survive the initial metabolic and phisycopathological damages require, on most cases, a surgical closure of the fistula which is a very complex procedure technically. We describe the case of a patient with a rectosigmoid neoplasm that was addressed laparoscopically and enteroatmospheric fistula just developing on phannestiel incision was used to extract the neoplasm. Conclusion: The closure assisted by ôhand made vacuumö and adequate nourishment allows recovering patients with complex fistulas to achieve adequate conditions for definitive closure. (AU)

Resección transanal a través de un trocar monopuerto. Una nueva aproximación al NOTES / Transanal resection using a single port trocar: a new approach to NOTES

Introducción La morbilidad y la mortalidad, así como las alteraciones funcionales que conllevan la cirugía radical del cáncer de recto, han hecho que se produzca un incremento en el interés por el tratamiento local del cáncer de recto en estadios precoces. La cirugía transanal convencional ofrece una tasa de recurrencia elevada por lo que en los últimos años la TEM se considera el tratamiento de elección en los adenomas rectales de gran tamaño y en el cáncer de recto precoz (T1 de bajo riesgo).Pacientes y método Hemos intervenido cinco pacientes, con diagnóstico de adenoma velloso de recto (4) y carcinoma in situ de recto (1), mediante este nuevo abordaje transanal, utilizando un dispositivo monopuerto. Resultados La localización de las lesiones fue en recto medio, a una distancia media del margen anal de 9cm (r: 6-10). Todas las lesiones estaban situadas en cara posterior. Los márgenes de resección en todos los casos fueron negativos. El tamaño medio de los adenomas fue de 4cm, siendo tres de forma sésil y uno con pedículo corto y grueso (> 1cm); el carcinoma tenía un tamaño de 3cm. El tiempo medio quirúrgico fue de 55 minutos. Todos los pacientes fueron dados de alta a las 48 horas del procedimiento quirúrgico. Conclusión Pensamos que el abordaje transanal endoscópico a través de un dispositivo monopuerto es un procedimiento sencillo, fácilmente reproductible y coste-efectivo comparado con la TEM (AU)

Introduction The morbidity and mortality, along with the functional changes that arise from radical surgery of rectal cancer, has led to an increasing interest in local treatment in the early stages of cancer of the rectum. Conventional transanal surgery has a high recurrence rate, for this reason transanal endoscopic microsurgery (TEM) is considered the treatment of choice in the last few years in large rectal adenomas and in early rectal cancer (low risk T1).Patients and method We have intervened five patients, four with a diagnosis of villous adenoma of the rectum, and one in situ rectal carcinoma, using this new transanal approach, with a single port device. Results The locations of the lesions were in the mid-rectum, at a mean distance of 9cm (range 6-10) from the anal margin. All of them were situated in the posterior side. The resection margins were negative in all cases. The mean size of the adenomas was 4cm, three being sessile shaped, and one with a short, thick pedicle (>1cm); the size of the carcinoma was 3cm. The mean surgical time was 55minutes. All the patients were discharged 48hours after the surgical procedure. Conclusion We believe that the transanal endoscopic approach with a single port device is a simple, easily reproducible and cost-effective procedure when compared to TEM (AU)

Efecto citoprotector del tacrolimus a dosis bajas sobre islotes de Langeerhans en cultivo sometidos a estímulos por citocinas del rechazo agudo / Cytoprotective effect of low-dose tacrolimus on islets of Langerhans in cultures subjected to stimulation by acute rejection cytokines

Introducción La mejoría de los resultados en el trasplante de islotes pancreáticos se debe en gran parte a la introducción de nuevos protocolos de inmunosupresión que incluyen, entre otros, tacrolimus a bajas dosis. Este fármaco tiene efectos antioxidantes y antiapoptóticos que podrían ser de utilidad en la prevención del rechazo primario. Objetivos Evaluar la respuesta in vitro a tacrolimus a bajas dosis en islotes de rata estimulados con citocinas proinflamatorias implicadas en el rechazo primario de islotes. Material y método Se cultivaron islotes de rata en medio RPMI determinándose producción de lipoperóxido (LPO) y óxido nítrico (NO) y marcadores de apoptosis (nucleosomas y Bcl-2) en presencia de IL-1 (50UI/ml) e IF-γ (1000UI/ml) y adición de tacrolimus (FK-506; 5ng/ml).Resultados Tras la estimulación se apreció un aumento muy significativo (p<0,01) de los marcadores de estrés oxidativo (LPO 10,1±1,16pmol/islote x 24; NO 19,1±3,28pmol/islote x 24h) y apoptosis (nucleosomas 0,24±0,04; Bcl-2 0,69±0,212). Dichos efectos fueron contrarrestados de manera significativa tras añadir tacrolimus, siendo la reversión completa (p NS frente a controles) en el caso de la producción de lipoperóxidos (1,58pmol/islote x 24h) y óxido nítrico (9,81pmol/islote x 24h) así como en el descenso de Bcl-2 (1,37±0,23Ui/islote).Conclusiones El efecto citoprotector in vitro del tacrolimus a bajas dosis sobre islotes estimulados con citocinas proinflamatorias consigue aminorar la generación de estrés oxidativo y la activación de la apoptosis, habitualmente implicados en el rechazo en las primeras 48h postimplante (AU)

Introduction The improvement in pancreatic islet transplantation results is due to immunosuppression protocols that include, among others, low-dose tacrolimus. Both anti-inflammatory and anti-oxidant effects of tacrolimus could be useful in preventing primary rejection. Aim To evaluate in vitro islet low-dose tacrolimus response after pro-inflammatory stimulation. Material and methodsIsolated rat islets were cultured in RPMI medium in the presence of IL-1 (50UI/mL) plus IF-γ (1000UI/mL) and tacrolimus (5ng/mL). The 24h production of lipoperoxide (LPO) and nitric oxide (NO) were measured as oxidative stress markers. Determination of apoptosis markers (nucleo some content and Bcl-2) was also performed. Results Oxidative stress (LPO 10.1±1.16pmol/islet x 24; NO 19.1±3.28pmol/islet x 24h) and apoptosis (nucleosome 0.24±0.04UI/islet; Bcl-2 0.69±0.212UI/islet) markers showed a very significant increase after cytokine stimulation (p<0.01). Both effects improved by adding tacrolimus to the medium. Protective effect was complete when lipoperoxide (1.58pmol/islet x 24h), nitric oxide (9.81pmol/islet x 24h) and Bcl-2 (1.37±0.23UI/islet) were determined. Conclusion In vitro cytoprotective effect of low-dose tacrolimus on isolated rat islets decreases both oxidative stress and apoptosis markers after stimulation of pro-inflammatory mediators (AU)

[Cytoprotective effect of low-dose tacrolimus on islets of Langerhans in cultures subjected to stimulation by acute rejection cytokines]. / Efecto citoprotector del tacrolimus a dosis bajas sobre islotes de Langeerhans en cultivo sometidos a estímulos por citocinas del rechazo agudo.

INTRODUCTION: The improvement in pancreatic islet transplantation results is due to immunosuppression protocols that include, among others, low-dose tacrolimus. Both anti-inflammatory and anti-oxidant effects of tacrolimus could be useful in preventing primary rejection. AIM: To evaluate in vitro islet low-dose tacrolimus response after pro-inflammatory stimulation. MATERIAL AND METHODS: Isolated rat islets were cultured in RPMI medium in the presence of IL-1 (50 UI/mL) plus IF-gamma (1000 UI/mL) and tacrolimus (5 ng/mL). The 24 h production of lipoperoxide (LPO) and nitric oxide (NO) were measured as oxidative stress markers. Determination of apoptosis markers (nucleosome content and Bcl-2) was also performed. RESULTS: Oxidative stress (LPO 10.1+/-1.16 pmol/islet x 24; NO 19.1+/-3.28 pmol/isletx24 h) and apoptosis (nucleosome 0.24+/-0.04 UI/islet; Bcl-2 0.69+/-0.212 UI/islet) markers showed a very significant increase after cytokine stimulation (p<0.01). Both effects improved by adding tacrolimus to the medium. Protective effect was complete when lipoperoxide (1.58 pmol/isletx24 h), nitric oxide (9.81 pmol/isletx24 h) and Bcl-2 (1.37+/-0.23 UI/islet) were determined. CONCLUSION: In vitro cytoprotective effect of low-dose tacrolimus on isolated rat islets decreases both oxidative stress and apoptosis markers after stimulation of pro-inflammatory mediators.