Tumor cell-based liquid biopsy using high-throughput microfluidic enrichment of entire leukapheresis product.

Circulating Tumor Cells (CTCs), interrogated by sampling blood from patients with cancer, contain multiple analytes, including intact RNA, high molecular weight DNA, proteins, and metabolic markers. However, the clinical utility of tumor cell-based liquid biopsy has been limited since CTCs are very rare, and current technologies cannot process the blood volumes required to isolate a sufficient number of tumor cells for in-depth assays. We previously described a high-throughput microfluidic prototype utilizing high-flow channels and amplification of cell sorting forces through magnetic lenses. Here, we apply this technology to analyze patient-derived leukapheresis products, interrogating a mean blood volume of 5.83 liters from patients with metastatic cancer, with a median of 2,799 CTCs purified per patient. Isolation of many CTCs from individual patients enables characterization of their morphological and molecular heterogeneity, including cell and nuclear size and RNA expression. It also allows robust detection of gene copy number variation, a definitive cancer marker with potential diagnostic applications. High-volume microfluidic enrichment of CTCs constitutes a new dimension in liquid biopsies.

Unusual presentation of a neuroendocrine tumor in the ileostomy specimen after rectal cancer treatment: a case report.

Background: Neuroendocrine tumors of the small intestine are uncommon, but at the same time they are the most frequent subtype of neuroendocrine tumor in the gastrointestinal system. They originate from enterochromaffin cells, which are involved in the creation of serotonin. This asymptomatic characteristic in the initial presentation is usually why these tumors are discovered at a late stage, sometimes in association with symptomatic metastatic disease. Case Description: We present a case-report of a 52-year-old gentleman with a suggestive family history of hereditary cancer syndrome (mother with lung cancer and maternal uncle with colon cancer at the age of 40 years old). The patient was diagnosed with rectal cancer and he received neoadjuvant chemotherapy with short-course radiotherapy followed by a robotic low anterior resection with diverting loop ileostomy. Following closure of his ileostomy, the pathology report of the ileostomy resection specimen showed a 1.1 cm neuroendocrine tumor with negative margins. Conclusions: This extraordinary unusual presentation could be very fortuity for the patient, who in every other opportunity just found this neuroendocrine tumor after advanced or maybe metastatic diseases.

Justin H. Kelly and his procedure for bladder exstrophy and epispadias.

The operations involved in the repair of complete bladder exstrophy (CBE), familiarly known as the Kelly procedure, evolved over more than 100 years. Through repeated cycles of trial and error, some of the most prominent urologists in the world developed techniques that addressed each of the formidable surgical challenges presented by CBE and epispadias. A key figure is Justin H. Kelly of the Royal Children's Hospital, Melbourne, who made surgery for CBE his life's work. He took the lessons of his surgical predecessors, giants like: Friedrich Trendelenburg, Hugh Hampton Young, John Dees, and Guy Leadbetter, applied techniques for anorectal anomalies from his contemporary Alberto Peña, and saw his procedures improved by the next generation of leaders in paediatric urology that included Phillip Ransley, Peter Cuckow, Patrick Duffy, and John Gearhart. Over his long career, Mr Kelly modified and perfected his eponymous procedure patient-by-patient through a painstaking process of trial-and-error, bearing with his young patients and their families through every heart-breaking complication, and gradually creating the standard operation for children with CBE and epispadias.

Sepsis.

This article provides an overview of the history of the sepsis definitions as well as an overview of the current understanding of the pathogenesis of sepsis. The evolution of the treatment bundles is also presented.

Transanal minimally invasive surgery for rectal cancer.

Due to the increased uptake of rectal cancer screening and the increasing rates of complete clinical response to chemoradiotherapy, more early-stage and down-staged rectal cancers are being treated. This has triggered surgeons to question the necessity for proctectomy and its associated morbidity and consider local excision and organ preservation in selected cases. Transanal minimally invasive surgery (TAMIS) has evolved as an oncologically safe yet cost-effective platform for local excision of rectal tumors using traditional laparoscopic instruments. This review highlights the recent advances and current role of TAMIS in the treatment of rectal cancer.

Root colonization and arbuscular mycorrhizal fungal community composition in a genetically modified maize, its non-modified isoline, and a landrace.

The use of genetically modified (GM) plants has increased in recent decades, but there are uncertainties about their effects on soil microbial communities. Aiming to quantify root colonization and characterize arbuscular mycorrhizal fungi (AMF) communities associated with roots and rhizosphere soil of different maize genotypes, a field trial was carried out in Southern Brazil with three maize genotypes as follows: a GM hybrid (DKB 240 VTPRO), its non-modified isoline (DKB 240), and a landrace (Pixurum). Soil samples were collected to evaluate the occurrence of AMF during the growth of corn genotypes at sowing and V3 (vegetative), R1 (flowering), and R3 (grain formation) stages of the crop. The occurrence of AMF was determined by the morphological identification of spores, and by analyzing AMF community composition in soil and roots of maize, using PCR-DGGE. The GM genotype of maize promoted lower mycorrhizal colonization in the vegetative stage and had lower sporulation at grain development than the conventional hybrid and the landrace maize. Twenty AMF morphotypes were identified and 13 were associated with all maize genotypes. The genera Acaulospora, Glomus, and Dentiscutata had the largest numbers of species. There were no differences in AMF community composition due to maize genotypes or genetic modification, but crop phenological stages affected AMF communities associated with maize roots.

Bifidobacterium animalis ssp. lactis BB-12 enumeration by quantitative PCR assay in microcapsules with full-fat goat milk and inulin-type fructans.

The current study was conducted to develop a quantitative polymerase chain reaction (qPCR) assay for Bifidobacterium animalis ssp. lactis BB-12 quantification in microcapsules matrix with full-fat goat milk and inulin-type fructans. DNA was isolated from milk, feed solutions (before spray drying) and microcapsules (after spray drying) using DNAzol. Two primer pairs targeting Bal-23S or Tuf sequences were evaluated by qPCR. The qPCR efficiency was higher (89.5%) using the Tuf primers than Bal-23S primers (84.8%). Tuf primer pair was able to selectively detect B. animalis ssp. lactis BB-12. After, quantification of bifidobacteria in the microcapsules matrix by Tuf qPCR assay was compared to conventional enumeration by plate counting. The analysis of probiotic feed solutions and microcapsules showed higher (P < 0.05) bacterial enumeration determined by Tuf qPCR assay compared to those obtained by plate counting. This qPCR assay was considered a rapid and sensitive alternative for the quantification of B. animalis ssp. lactis BB-12 in probiotic microcapsules compared to plate counting.

Achieving Opioid-Free Major Colorectal Surgery: Is It Possible?

INTRODUCTION: Opioid analgesia remains the mainstay of postoperative pain management strategies despite being associated with many adverse effects. A specific opioid-free protocol was designed to limit opioid usage. OBJECTIVE: The aim of the study was to audit the opioid-free rate within this protocol and to identify factors that might contribute to opioid-free surgery. METHODS: A retrospective study of all elective patients receiving abdominal colorectal surgery at the Center for Colon and Rectal Surgery at AdventHealth over 6 months was performed. Data on demographics, indications, perioperative management, outcomes, and inpatient and outpatient analgesic requirements were collected with subsequent analysis. RESULTS: A total of 303 consecutive patient records were analyzed. Approximately two-thirds (67.7%) of patients did not receive narcotics once they left the postanesthesia care unit as an inpatient. One-third of patients (32.0%) did not receive narcotic analgesia within 30 days of surgery as an outpatient. Patients in the opioid-free cohort were significantly older and had a malignant indication, less perioperative morbidity, and a shorter length of stay. CONCLUSIONS: Our study demonstrates that opioid-free analgesia is indeed possible in major colorectal surgery. Study limitations include its retrospective nature and that it is from a single institution. Despite these limitations, this study provides proof of concept that opioid-free colorectal surgery is possible within a specific protocol.

Defining the learning curve for transanal total mesorectal excision for rectal adenocarcinoma.

BACKGROUND: Early observational data suggest that this approach is safe and feasible, but it is technically challenging and the learning curve has not yet been determined. The objective of this study was to determine the number of cases required achieve proficiency in transanal total mesorectal excision (TA-TME) for rectal adenocarcinoma. METHODS: All TA-TME cases performed from 03/2012-01/2017 at a single high-volume tertiary care institution for rectal adenocarcinoma were included. A cumulative summation (CUSUM) analysis was performed to determine the number of cases required to reach proficiency, defined as high-quality TME (complete or near-complete mesorectal envelope, negative distal (DRM), and circumferential resection (> 1 mm; CRM) margin). The acceptable and unacceptable rates of good quality TME were defined based on the incidence of high-quality TME in laparoscopic (unacceptable rate = 81.7%) and open (acceptable rate = 86.9%) arms of the ACOSOG Z6051 trial. RESULTS: A total of 87 consecutive cases were included with mean tumor height 4.8 cm (SD 2.7) and 80% (70/87) received neoadjuvant chemoradiation. Post-operative morbidity occurred in 44% (38/87) of cases, including 21% (18/87) readmissions. Median length of stay was 4 days [IQR 3-8]. A good quality TME was performed in 95% (83/87) of cases including 98% (85/87) negative CRM, 99% (86/87) negative DRM, and 99% (86/87) complete or near-complete mesorectal envelope. CUSUM analysis reported that the good quality TME rate reaches an acceptable rate after 51 cases overall, and 45 cases if abdominoperineal resections are excluded. CONCLUSION: TA-TME is a complex technique that requires a minimum of 45-51 cases to reach an acceptable incidence of high-quality TME and lower operative duration.

Impact of residual nodal involvement after complete tumor response in patients undergoing neoadjuvant (chemo)radiotherapy for rectal cancer.

BACKGROUND: The management of patients with a complete clinical response after neoadjuvant therapy for rectal adenocarcinoma is controversial. Those who advocate for resection point out the inaccuracy of N-staging with current imaging modalities. The objective of this study is to determine the impact of residual nodal involvement after complete tumor regression after neoadjuvant (chemo)radiotherapy. METHODS: The 2004 to 2014 National Cancer Database was queried for patients undergoing proctectomy for nonmetastatic rectal adenocarcinoma who had received neoadjuvant (chemo)radiotherapy and with ypT0 on final pathology. Patients were grouped based on pathologic nodal stage: ypT0N- and ypT0N+. The main outcome was 5-year overall survival. RESULTS: There were 5,156 patients with ypT0N- and 527 with ypT0N+. Mean lymph node harvest was similar (ypT0N- 12.2 nodes [standard deviation 9.1] vs ypT0N+ 11.6 nodes [standard deviation 10.3]; P = .086). Patients with ypT0N+ were more likely to have had clinically involved nodes (P < .001) and earlier clinical T-stage (P = .002). Overall survival at 5 years was less for patients with ypT0N+ (80% vs 86%, log-rank P = .014). ypT0N+ was independently associated with worse overall survival (hazard ratio 1.74, 95% confidence interval 1.33-2.28). CONCLUSION: Residual nodal involvement despite complete tumor regression was associated with worse 5-year overall survival compared to complete pathologic response. Additional therapy should be considered in the presence of complete clinical tumor regression after neoadjuvant (chemo)radiotherapy.

Minimally invasive surgery for stage III colon adenocarcinoma is associated with less delay to initiation of adjuvant systemic therapy and improved survival.

BACKGROUND: Minimally invasive surgery (MIS) may improve surgical recovery and reduce time to adjuvant systemic therapy after colon cancer resection. The objective of this study was to determine the effect of MIS on the initiation of adjuvant systemic therapy and survival in patients with stage III colon cancer. METHODS: The 2010-2014 National Cancer Database was queried for patients with resected stage III colon adenocarcinoma, and divided into MIS, which included laparoscopic and robotic approaches, and open surgery. Propensity-score matching was used to balanced open and MIS groups. The main outcome measures were delayed initiation of adjuvant systemic therapy (defined as > 8 weeks after surgery) and 5-year overall survival (OS). Multiple Cox regression was performed to identify independent predictors for 5-year OS, including an interaction between delayed systemic therapy and MIS, and adjusted for clustering at the hospital level. RESULTS: There were 86,680 patients that were included in this study. Overall, 45% (38,713) underwent MIS colectomy, of which 93% underwent laparoscopic and 7% robotic surgery. After matching, 33,183 open patients were balanced to 33,183 MIS patients. Patient, tumor, and facility characteristics were similar in the matched cohort. More patients in the MIS group received adjuvant therapy within 8 weeks of surgery (49% vs. 42%, p < 0.001), and fewer MIS patients did not receive any systemic therapy (30% vs. 35%, p < 0.001). Delayed initiation of systemic therapy > 8 weeks was associated with worse 5-year OS (HR 1.27, 95%CI 1.19-1.36). MIS was independently associated with improved survival (HR 0.92, 95%CI 0.86-0.97). This relationship remained even if 90-day mortality was excluded. CONCLUSIONS: MIS approaches are associated with less delay to the initiation of adjuvant systemic therapy and improved survival in patients with stage III colon adenocarcinoma. Surgeons should favor MIS approaches for the treatment of stage III colon adenocarcinoma whenever possible.

Lower survival after right-sided versus left-sided colon cancers: Is an extended lymphadenectomy the answer?

INTRODUCTION: Tumour location may affect oncologic outcomes for colon adenocarcinoma due to different levels of vascular ligation and nodal harvest, but the data are equivocal. The objective of this study is to determine the effect of tumor location and lymph node yield on overall survival(OS) in stage I-III colon adenocarcinoma. METHODS: The 2004-2014 National Cancer Database was queried for colectomies for non-metastatic colon adenocarcinoma, excluding transverse colon and rectal cancer. Patients were grouped based on left/right tumor location. Main outcome measure was 5-year OS. Propensity score matching created balanced cohorts. Multilevel survival analysis determined the independent effect of tumor location and nodal harvest on OS. RESULTS: There were 504,958 patients (273,198 right; 231,760 left) in the entire cohort: 26.4% stage-I, 37.3% stage-II, and 36.3% stage-III (equal distribution left/right). After 1:1 matching(n = 297,080), right cancers were associated with worse 5-year overall survival for stage-II (66% vs. 70%, p < 0.001) and -III (56% vs. 60%, p < 0.001) despite similar nodal harvest and proportion receiving systemic therapy. On multivariate analysis, right-sided cancers (HR 1.12, 95%CI 1.06-1.19) had worse OS, independent of stage and nodal harvest. Nodal harvest ≥22 nodes had the highest OS (HR 0.71, 95%CI 0.68-0.75). There was an interaction between right-sided cancer and >22 lymph node harvest towards increased survival (HR 0.86, 95%CI 0.80-0.92). CONCLUSIONS: Right-sided cancers are associated with worse oncologic outcomes compared to left-sided tumors but a higher lymph node yield improves survival. These data provide indirect evidence for a higher lymphatic harvest to improve survival.

Transanal transabdominal TME: how far can we push it?

Over many decades, advances in surgical technology, such as the use of the electrocautery Bovie, development of minimally invasive and advanced endoscopic platforms and the ability to create and maintain pneumorectum have propelled surgical techniques forward to today, with development of the transanal total mesorectal excision TME (taTME) for en bloc resection of rectal cancers. The transanal platform offers, for now, a viable alternative to perform safe and oncologically sound TME, especially favorable in cases of low rectal lesions in a narrow pelvis post neoadjuvant treatment. The aspiration of the colorectal community remains to continue to push the operative boundaries whilst maintaining safe oncological principals with the best possible functional outcomes for patients. In this article we review this evolving technique and focus on future directions.

Establishing the learning curve of transanal minimally invasive surgery for local excision of rectal neoplasms.

INTRODUCTION: Transanal minimally invasive surgery (TAMIS) is an endoscopic operating platform for local excision of rectal neoplasms. However, it may be technically demanding, and its learning curve has yet to be adequately defined. The objective of this study was to determine the number of TAMIS procedures for the local excision of rectal neoplasm required to reach proficiency. METHODS AND PROCEDURES: All TAMIS cases performed from 07/2009 to 12/2016 at a single high-volume tertiary care institution for local excision of benign and malignant rectal neoplasia were identified from a prospective database. A cumulative summation (CUSUM) analysis was performed to determine the number of cases required to reach proficiency. The main proficiency outcome was rate of margin positivity (R1 resection). The acceptable and unacceptable R1 rates were defined as the R1 rate of transanal endoscopic microsurgery (TEM-10%) and traditional transanal excision (TAE-26%), which was obtained from previously published meta-analyses. Comparisons of patient, tumor, and operative characteristics before and after TAMIS proficiency were performed. RESULTS: A total of 254 TAMIS procedures were included in this study. The overall R1 resection rate was 7%. The indication for TAMIS was malignancy in 57%. CUSUM analysis reported that TAMIS reached an acceptable R1 rate between 14 and 24 cases. Moving average plots also showed that the mean operative times stabilized by proficiency gain. The mean lesion size was larger after proficiency gain (3.0 cm (SD 1.5) vs. 2.3 cm (SD 1.3), p = 0.008). All other patient, tumor, and operative characteristics were similar before and after proficiency gain. CONCLUSIONS: TAMIS for local excision of rectal neoplasms is a complex procedure that requires a minimum of 14-24 cases to reach an acceptable R1 resection rate and lower operative duration.

Synchronous Upper Squamous and Lower Adenocarcinoma of the Oesophagus: A Rarely Reported Case Treated with Palliative Chemotherapy and Stenting.

Oesophageal cancer is divided into two main subtypes, squamous and adenocarcinoma. It is the eighth most common cancer in the world with squamous more common in the developing world and adenocarcinoma most prevalent in the developed world. Incidences of concomitant squamous carcinoma with adenocarcinoma are exceedingly rare with only a few documented occurrences in the form of case reports existing. Here we report a case of synchronous squamous and adenocarcinoma of the oesophagus occurring in an 81-year-old lady with dysphagia, weight loss, and no identifiable risk factors.

Chemoradiation and Local Excision for T2N0 Rectal Cancer Offers Equivalent Overall Survival Compared to Standard Resection: a National Cancer Database Analysis.

BACKGROUND: Local excision (LE) alone is associated with worse survival compared to radical surgery (RS) for T2 rectal cancer, but LE with additional chemoradiation (CRT) may improve outcomes. The objective of this study was to compare combined CRT and LE versus RS for T2 rectal cancer. METHODS: The 2004-2014 National Cancer Database was queried for patients with T2N0M0 rectal cancer undergoing LE with neoadjuvant(NA-CRT + LE) or adjuvant(LE + Adj-CRT) CRT, or RS. The main outcome was 5-year overall survival (OS). Cox proportional hazards was used to determine the independent effect of treatment on OS. RESULTS: A total of 4822 patients were included (4367 RS, 242 CRT + LE, 213 LE + Adj-CRT). Mean follow-up was 48.6 (SD28.5) months. There were no differences in patient characteristics, but more high-risk features in the LE + Adj-CRT group. There were no differences in 90-day mortality. Five-year OS was similar (RS 77.4% vs. CRT + LE 76.1% vs. LE + Adj-CRT 79.7%, p = 0.786). Older age, male gender, and higher Charlson score were independently associated with worse OS, whereas treatment type was not. If 90-day mortality was excluded, LE + Adj-CRT was independently associated with worse OS compared to RS. CONCLUSIONS: CRT with LE for T2N0M0 rectal cancer was not associated with worse OS compared to RS, and may be a viable treatment modality.

Communities of endophytic microorganisms in different developmental stages from a local variety as well as transgenic and conventional isogenic hybrids of maize.

The diversity of endophytic microorganisms may change due to the genotype of the host plant and its phenological stage. In this study we evaluated the effect of phenological stage, transgenes and genetic composition of maize on endophytic bacterial and fungal communities. The maize populations were composed of a local variety named Rosado (RS) and three isogenic hybrids. One isogenic hybrid was not genetically modified (NGM). Another hybrid (Hx) contained the transgenes cry1F and pat (T1507 event), which provide resistance to insects of the order Lepidoptera and tolerance to the glufosinate-ammonium herbicide, respectively. The third hybrid (Hxrr) contained the transgene cp4 epsps (NK603 event) combined with the transgenes cry1F and pat (T1507 event), which allow tolerance to the Roundup Ready herbicide, besides the characteristics of Hx. Evaluation of the foliar tissue was done through PCR-DGGE analysis, with specific primers for bacteria and fungi within four phenological stages of maize. The endophytic bacteria were only clustered by phenological stages; the structure of the fungal community was clustered by maize genotypes in each phenological stage. The fungal community from the local variety RS was different from the three hybrids (NGM, Hx and Hxrr) within the four evaluated stages. In the reproductive stage, the fungal community from the two transgenic hybrids (Hx and Hxrr) were separated, and the Hxrr was different from NGM, in the two field experiments. This research study showed that the genetic composition of the maize populations, especially the presence of transgenes, is the determining factor for the changes detected in the endophytic fungal community of maize leaves.

Myeloid-related protein 8 induces self-tolerance and cross-tolerance to bacterial infection via TLR4- and TLR2-mediated signal pathways.

Myeloid-related protein 8 (Mrp8) is the active component of Mrp8/14 protein complex released by phagocytes at the site of infection and stimulates inflammatory responses. However, it is unclear whether Mrp8 could induce self-tolerance and cross-tolerance to bacterial infection. Here we report that Mrp8 triggered TNF-α and IL-6 release via a Toll-like receptor 4 (TLR4)-dependent manner. Pre-stimulation of murine macrophages and human monocytes with Mrp8 induced self-tolerance to Mrp8 re-stimulation and cross-tolerance to lipopolysaccharide (LPS), bacterial lipoprotein (BLP), gram-negative and gram-positive bacterial challenges, with substantially attenuated TNF-α and IL-6 release. Moreover, Mrp8 tolerisation significantly reduced serum TNF-α and IL-6, increased polymorphonuclear neutrophil (PMN) recruitment and accelerated bacterial clearance, thus protecting mice against LPS-induced lethality and cecal ligation and puncture (CLP)-induced polymicrobial sepsis. In addition to TLR4, TLR2 also contributed to Mrp8-induced inflammatory response and tolerance. Down-regulation of phosphorylated p38 by Mrp8 pre-stimulation was predominantly responsible for the intracellular mechanism of Mrp8-induced tolerance. Thus, our findings of Mrp8-induced self-tolerance and cross-tolerance may provide a potential strategy for attenuating an overwhelming proinflammatory cascade and enhancing antimicrobial responses during microbial sepsis.

Metastatic breast cancer presenting as a gallstone ileus.

Metastatic breast cancer to the small bowel (SB) presenting as gallstone ileus and resulting in SB obstruction has not been described previously. A 76-year-old woman with previous metastatic breast cancer to the axial spine and hips presented with abdominal pain and bilious vomiting. CT scanning revealed SB obstruction consistent with gallstone ileus. The patient underwent two segmental SB resections for distal ileal strictures mimicking what appeared to be macroscopic Crohn's disease. The entero-biliary fistula was undisturbed. Pathological analysis revealed the dual pathologies of gallstone ileus and metastatic carcinoma from a breast primary causing luminal SB obstruction. Improvements in staging and treatment modalities have contributed to the increased overall long-term survival for breast cancer, compelling clinicians to consider metastatic breast cancer as a differential diagnosis in women presenting with new onset of gastrointestinal symptoms in order that appropriate treatment be administered in a timely fashion.