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Single cancer stem–like cells (CSC) from colorectal cancers can be functionally identified by their ability to form large lumen-containing colonies in three-dimensional Matrigel cultures. These colonies contain the three types of differentiated colorectal epithelial cells, and single cells obtained from them can reproduce themselves and form tumors efficiently in immunodeficient mice. In this study, we show how hypoxia affects these CSC-derived lumens to control differentiation of stem-like cells and enterocytes via the homeobox gene CDX1. Lumens were identified by F-actin staining and they expressed many characteristics associated with normal differentiated intestinal epithelium, including brush border enzymes, polarization, and tight junctions. RNA interference–mediated silencing of CDX1 reduced lumen formation. Inhibitory effects of hypoxia on lumen formation and stem cell differentiation, including suppression of CDX1 expression, could be mimicked by inhibiting prolyl-hydroxylases that activate HIF1, suggesting that HIF1 is a critical mediator of the effects of hypoxia in this setting. Cell line–derived lumens were phenotypically indistinguishable from colorectal tumor glandular structures used by pathologists to grade tumor differentiation. Parallel results to those obtained with established cell lines were seen with primary cultures from fresh tumors. This in vitro approach to functional characterization of CSCs and their differentiation offers a valid model to study colorectal tumor differentiation and differentiation of colorectal CSCs, with additional uses to enable high-throughput screening for novel anticancer compounds. Cancer Res; 73(18); 5798–809. ©2013 AACR.

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PDF file, 287K, Lumen colonies express polarised enterocyte markers.

PDF file, 363K, Gallery of F-actin labelling of random colonies of various cell lines grown from single cells in Matrigel for 10 days.

PDF file, 266K, DMOG Induces Expression of Hif1alpha and hypoxia regulated genes Glut1 and Carbonic Anhydrase IX.

PDF file, 270K, Stromal alpha Smooth muscle actin labelled myofibroblasts co-stain with Beta-actin.

Objective The coronavirus pandemic has significantly disrupted the way we deliver healthcare worldwide. We have been flexible and creative in order to continue providing elective colorectal cancer operations and to restart services for benign cases during the recovery period of the pandemic. In this paper, we describe the impact of coronavirus on our elective services and how we have implemented new patient pathways to allow us to continue providing patient care. Methodology Data on major colorectal elective resections was prospectively collected in an Enhanced Recovery After Surgery (ERAS) database. Data on the number of proctology cases and telemed appointments were collected from the hospital theatre information management system and electronic patient record system respectively. Results During the pandemic, there was a complete shift towards cancer cases, with benign services and proctology cases being placed on hold. Hospital length of stay was reduced. We implemented earlier hospital discharge and more intense telephone follow up after elective major surgery. This has not resulted in an increase in post‐operative complications, nor any increase in readmission into hospital. During the recovery phase, we have introduced a higher proportion of telemed consultations, including one‐stop telemed proctology clinics, resulting in straight to tests or investigations. Conclusions We have created a streamlined multi‐disciplinary pathway to reinstate our elective colorectal services as soon as possible and to minimise potential harm caused to patients whose treatment have been delayed. We anticipate many of these changes will be permanently incorporated into our clinical practice once the pandemic is over.

Background Iatrogenic ureteric injury is a serious complication of colorectal surgery. Incidence is estimated to be between 0.3 and 1.5%. Of all ureteric injuries, 9% occur during colorectal procedures. Ureteric stents are utilised as a method to reduce the risk of injury; however, these are not without risk and do not guarantee prevention of injury. Fluorescence is a safe and effective alternative for intraoperative ureteric localisation. This proof of principle study aims to assess the use of methylene blue to fluoresce the ureter during colorectal surgery. Method Patients undergoing elective colorectal surgery were included in this open label, non-randomised study. Methylene blue was administered intravenously at varying doses (0.25–1 mg/kg) over 5 min, 10–15 min prior to entering ‘ureteric territory.’ Fluorescence was assessed using the PINPOINT Deep Red laparoscopic system at fixed time points by the surgeon and an independent observer. Results 42 patients received methylene blue; 2 patients were excluded from analysis. Of the 69 ureters assessed, 64 were seen under fluorescence. Of these, 14 were not visible under white light. 50 ureters were observed with both fluorescence and white light with 14 of these being seen earlier with fluorescence. In ten cases, fluorescence revealed the ureter to be in a different location than suspected. Conclusion Fluorescence is a promising method to allow visualisation of the ureter, where it is not identified easily under standard operative conditions, thereby improving safety and reducing operative time and difficulty. Electronic supplementary material The online version of this article (10.1007/s00464-018-6219-8) contains supplementary material, which is available to authorized users.

Background Standard surgical practice for colorectal cancer involves resection of the primary lesion and all draining lymph nodes. Accurate intraoperative assessment of nodal status could allow stratified resectional extent. One-step nucleic acid (OSNA) can provide a rapid method of interrogating nodal tissue, whilst near-infrared (NIR) laparoscopy together with indocyanine green (ICG) can identify relevant nodal tissue intraoperatively. Methods ICG was administered around the tumour endoscopically prior to the operation. Fluorescent nodes identified by NIR were marked and submitted for whole-node OSNA analysis. Further fresh lymph nodes dissected from the standard resection specimen were examined and analysed by both conventional histology and OSNA. In addition, the status of the fluorescent nodes was compared to that of non-ICG nodes to assess their predictive value. Results Sixteen patients were recruited with a total final lymph node count of 287. 78 fresh lymph nodes were identified on fresh dissection for both histological and OSNA assessment with an analytical concordance rate of 98.7% (77/78). OSNA sensitivity was 1 (0.81–1, 95% CI) and specificity 0.98 (0.91–1, 95% CI). Six patients had a total of nine nodes identified intraoperatively by ICG fluorescence. Of these nine nodes, one was positive for metastasis on OSNA. OSNA analysis of the ICG-labelled node matched the final histological nodal stage in 3/6 patients (two being N0 and one N1). The final pathological nodal stage of the other three was N1 or N2, while the ICG nodes were negative. Conclusion OSNA is highly concordant with standard histology, although only a minority of nodes identifiable by full pathological analysis were found for OSNA on fresh dissection. OSNA can be combined with NIR and ICG lymphatic mapping to provide intraoperative assessment of nodal tissue in patients with colorectal cancer.

Background: Our study aimed to identify the search engines and terms commonly used by inflammatory bowel disease (IBD) patients and aimed to assess the quality and readability of these resources.Methods: Patients attending IBD clinic were surveyed, regarding search engines, terms and number of websites viewed. Websites according to these predetermined criteria were identified. Website content was described and quality was assessed using DISCERN. Readability was graded using the Flesch Reading Ease Score (FRES).Results: From 33 survey responses, Google was universally utilised. Forty-two websites met the inclusion criteria (19 for Crohn’s disease (CD), 23 for ulcerative colitis (UC). Only one website originated from Australia. Websites were infrequently updated (CD 21%, UC 17%) within the previous 12 months. Overall readability was poor with a mean FRES of 35.8 (11.8, range 15.7-57.7) for CD and 35.7 (11.3, range 19.4-54.3) for UC websites. Quality was moderate to poor with only five (12%) websites being rated as high quality (2 CD, 3 UC).Conclusions: There is very little Australian based web information available on IBD surgery and overall, it is of a low readability and poor content quality. There is need for the development of patient targeted health literature to help these patients.

Background and aims Anastomotic dehiscence is one of the most feared complications in colorectal surgery leading to significant morbidity and mortality. Progressively lower anastomoses are associated with a greater leak rate. One of the key factors is the perfusion of the bowel to be joined. Presently, surgeons rely on a variety subjective measures to determine anastomotic perfusion and mechanical integrity however these have shortcomings. The aim of this paper is to appraise the literature on the use of fluorescence angiography (FA) in laparoscopic rectal surgery. Materials and methods A Pubmed search was undertaken using terms ‘fluorescence angiography’ and ‘rectal surgery’. The search was expanded using the related articles function. Studies were included if they used FA specifically for rectal surgery. Outcomes of interest including anastomotic leak rate, change of operative strategy and time taken for FA were recorded. Results Eleven papers detailing the use of FA in rectal surgery are outlined demonstrating that this technique may change operative strategy and lead to a reduction in anastomotic leak rate. Conclusion In this paper, we discuss assessment of colorectal blood supply using FA and how this technique holds great potential to detect insufficiently perfused bowel. In so doing, the operator can adjust their operative strategy to mitigate these affects with the aim of reducing the complications of anastomotic leak and stenosis. However, it is highlighted that there is a clear need for randomised controlled trials in order to determine this definitively.

The application of primary organoid cultures containing epithelial and mesenchymal elements to cancer modeling holds promise for combining the accurate multilineage differentiation and physiology of in vivo systems with the facile in vitro manipulation of transformed cell lines. Here we used a single air-liquid interface culture method without modification to engineer oncogenic mutations into primary epithelial and mesenchymal organoids from mouse colon, stomach and pancreas. Pancreatic and gastric organoids exhibited dysplasia as a result of expression of Kras carrying the G12D mutation (Kras(G12D)), p53 loss or both and readily generated adenocarcinoma after in vivo transplantation. In contrast, primary colon organoids required combinatorial Apc, p53, Kras(G12D) and Smad4 mutations for progressive transformation to invasive adenocarcinoma-like histology in vitro and tumorigenicity in vivo, recapitulating multi-hit models of colorectal cancer (CRC), as compared to the more promiscuous transformation of small intestinal organoids. Colon organoid culture functionally validated the microRNA miR-483 as a dominant driver oncogene at the IGF2 (insulin-like growth factor-2) 11p15.5 CRC amplicon, inducing dysplasia in vitro and tumorigenicity in vivo. These studies demonstrate the general utility of a highly tractable primary organoid system for cancer modeling and driver oncogene validation in diverse gastrointestinal tissues.

Single cancer stem–like cells (CSC) from colorectal cancers can be functionally identified by their ability to form large lumen-containing colonies in three-dimensional Matrigel cultures. These colonies contain the three types of differentiated colorectal epithelial cells, and single cells obtained from them can reproduce themselves and form tumors efficiently in immunodeficient mice. In this study, we show how hypoxia affects these CSC-derived lumens to control differentiation of stem-like cells and enterocytes via the homeobox gene CDX1. Lumens were identified by F-actin staining and they expressed many characteristics associated with normal differentiated intestinal epithelium, including brush border enzymes, polarization, and tight junctions. RNA interference–mediated silencing of CDX1 reduced lumen formation. Inhibitory effects of hypoxia on lumen formation and stem cell differentiation, including suppression of CDX1 expression, could be mimicked by inhibiting prolyl-hydroxylases that activate HIF1, suggesting that HIF1 is a critical mediator of the effects of hypoxia in this setting. Cell line–derived lumens were phenotypically indistinguishable from colorectal tumor glandular structures used by pathologists to grade tumor differentiation. Parallel results to those obtained with established cell lines were seen with primary cultures from fresh tumors. This in vitro approach to functional characterization of CSCs and their differentiation offers a valid model to study colorectal tumor differentiation and differentiation of colorectal CSCs, with additional uses to enable high-throughput screening for novel anticancer compounds. Cancer Res; 73(18); 5798–809. ©2013 AACR.

Purpose The cancer stem cell hypothesis predicts that only a subpopulation of cells within a tumor is responsible for driving growth. If this hypothesis were true, it would have a significant impact on our current treatment of cancer because conventional chemotherapy and radiotherapy target rapidly proliferating cells making up the bulk of the tumor, not specifically cancer stem cells. The aims of this review are to highlight the current evidence supporting the existence of cancer stem cells in colorectal cancer, to consider the relative merits of current cancer stem cell markers, and to discuss the implications of this on our current treatment of cancer. Methods Published scientific articles were selected by searching the PubMed database by use of the terms "colorectal," "cancer," and "stem cells," and by use of the bibliographies of extracted articles. Results and conclusion CD133, a glycosylated cell surface protein, has been demonstrated to isolate for a subpopulation of colorectal tumor cells enriched in cancer stem cells. However, only 1 in 262 CD133+ cells are able to initiate tumors. Other cancer stem cell markers have been investigated, but an overall need exists to identify more specific markers to allow further characterization of these cancer stem cells. We discuss how increased understanding of the distribution and behavior of cancer stem cells within tumors could have significant implications for the management of colorectal cancer, including screening, resection margins, sentinel node biopsy, determination of prognosis, and the development of novel therapeutic targets.

Cancer stem cells (CSCs) are the subpopulation of cells within a tumor that can self-renew, differentiate into multiple lineages, and drive tumor growth. Here we describe a two-pronged approach for the identification and characterization of CSCs from colorectal cancer cell lines, using a Matrigel-based differentiation assay, and cell surface markers CD44 and CD24. About 20 to 30% of cells from the SW1222 cell line form megacolonies in Matrigel that have complex 3D structures resembling colonic crypts. The megacolonies’ capacity to self-renew in vitro is direct evidence that they contain the CSCs. Furthermore, just 200 cells from SW1222 megacolonies initiate tumors in NOD/SCID mice. We also showed that CD44 + CD24 + cells enriched for colorectal CSCs in the HT29 and SW1222 cell lines, which can self-renew and reform all four CD44/CD24 subpopulations, are the most clonogenic in vitro and can initiate tumors in vivo. A single SW1222 CD44 + CD24 + CSC, when grown in Matrigel, can form large megacolonies that differentiate into enterocyte, enteroendocrine, and goblet cell lineages. The HCT116 line does not differentiate or express CDX1, nor does it contain subpopulations of cells with greater tumor-forming capacity, suggesting that HCT116 contains mainly CSCs. However, forced expression of CDX1 in HCT116 leads to reduced clonogenicity and production of differentiating crypt-containing colonies, which can explain the selection for reduced CDX1 expression in many colorectal cancers. In summary, colorectal cancer cell lines contain subpopulations of CSCs, characterized by their cell surface markers and colony morphology, which can self-renew and differentiate into multiple lineages.

Background The Internet is a vast resource available for patients to obtain health information. Objective This study examines the quality of Web sites that provide information on diverticular disease, treatment options, and surgery. Design Two search engines (Google and Yahoo) and the search terms "surgery and diverticular disease" and "surgery and diverticulitis" were used. The first 50 sites of each search were assessed. Sites that fulfilled the inclusion criteria were evaluated for content and scored by using the DISCERN instrument, which evaluates the quality of health information on treatment choices. Results Two hundred sites were examined, of which 60 (30%) provided patient-orientated information. 50 sites (25%) were duplicated, 7 (3.5%) were links, 10 (5%) were advertisements, 14 (7%) were resources for clinicians, 9 (4.5%) were message forums, 27 (13.5%) were articles, and 15 (7.5%) were dead links. Of the 60 Web sites that provided patient information, only 10 (16.7%) had been updated within the past 2 years. Seventeen (28.3%) sites were affiliated with hospitals and clinics, but another 17 (28.3%) sites were associated with private companies with commercial interests. Although most Web sites contained information on symptoms, complications, investigations, and treatment options of diverticular disease, 20 (33.3%) did not describe any of the risks of surgery, and 45 (75%) did not provide information on the timescale of recovery postoperatively. Eighteen sites did not provide balanced information on treatment options; of these, 7 were biased toward medical treatment and 6 focused on laparoscopic surgery. Overall, only 22 (36.7%) were identified as being "good" or "excellent" with the use of the DISCERN criteria. Conclusions The quality of patient information on surgery for diverticular disease is highly variable, and Web sites that are sponsored by private companies may be biased in discussing treatment options. There is potential for the Internet to provide valuable information, and clinicians should guide patients to access high-quality Web sites.

Hypoxia is an important regulator of normal and cancer stem cell (CSC) differentiation. Colorectal CSCs from SW1222, LS180, and CCK81 colorectal cancer-derived cell lines are able to differentiate into complex 3D lumen-containing structures in normoxia, whereas in hypoxia, they form undifferentiated dense colonies that have reduced expression of the enterocyte differentiation marker CDX1, lack goblet cell formation, and have increased expression of BMI1 and activated Notch1. Hypoxia increases the clonogenicity of CSCs, which is cumulative as each round of hypoxia enriches for more CSCs. The hypoxic phenotype is reversible, because cells from hypoxic-dense colonies are able to reform differentiated structures when regrown in normoxia. We show that CDX1 is able to stimulate the generation of lumens even in hypoxia and has a negative feedback on BMI1 expression. Knockdown of CDX1 reduces lumen formation but does not affect goblet cell formation, suggesting that enterocytes and goblet cells form from different progenitor cells. Notch inhibition by dibenzazepine (DBZ) allowed CSCs to form goblet cells in both normoxia and hypoxia. Finally, we show that Hif1α, but not CA9, is an important mediator of the effects of hypoxia on the clonogenicity and differentiation of CSCs. In summary, hypoxia maintains the stem-like phenotype of colorectal cell line-derived CSCs and prevents differentiation of enterocytes and goblet cells by regulating CDX1 and Notch1, suggesting that this regulation is an important component of how hypoxia controls the switch between stemness and differentiation in CSCs.

Colorectal surgery is one of the most common procedures performed around the world with more than 600,000 operations each year in the United States, and more than a million worldwide. In the past two decades, there has been a clear trend toward minimal access and surgeons have embraced this evolution. Widespread adoption of advanced minimally invasive procedures is often limited by procedural complexity and the need for specific technical skills. Furthermore, the loss of 3D vision, limited overview of the surgical field, and diminished tactile sensation make major colorectal procedures more challenging and have an impact on the surgeons' learning curves. New technologies are emerging that can compensate for some of the sensory losses associated with laparoscopy. High-definition picture acquisition, 3D camera systems, and the use of biomarkers will allow improved identification of the target structures and help differentiate them from surrounding tissues. In this article, we describe some of the new technologies available and, in particular, focus on the possible implications of biomarkers and fluorescent laparoscopic imaging.

We describe the case of an 81-year-old woman with large bowel-obstruction caused by an impacted gallstone. An 81-year-old, Caucasian, fully independent woman without significant comorbidities presented with absolute constipation, faecal vomiting and abdominal pain. Abdominal radiography revealed dilated small bowel, and a subsequent contrast CT demonstrated a 2.5 cm gallstone in the sigmoid colon. This is believed to have entered the transverse colon via a cholecyst-colonic fistula, and then migrated to a section of sigmoid colon affected by diverticular disease, where it became impacted. Two sigmoidoscopic removals were attempted but were unsuccessful as the gallstone9s size prevented removal with an endoscopic basket. A laparotomy was performed and the stone extracted via a sigmoid enterotomy. No covering stoma was formed, and following 48 h on intensive trauma unit and a short ward-based stay for rehabilitation, the patient was discharged home and is currently doing well.

Introduction Haemorrhoids are the most common benign condition seen by colorectal surgeons. At clinic appointments, advice given about lifestyle modification or surgical interventions may not be understood fully by patients. Patients may use the internet for further research into their condition. However, the quality of such information has not been investigated before. This study assessed the quality of patient information on surgical treatment of haemorrhoids on the internet. Methods Four searches were carried out using the search terms ‘surgery for haemorrhoids’ and ‘surgery for piles’ on two search engines (Google and Yahoo). The first 50 results for each search were assessed. Sites were evaluated using the DISCERN instrument. Results In total, 200 websites were assessed, of which 144 fulfilled the inclusion criteria. Of these, 63 (44%) were sponsored by herbal remedies for haemorrhoids. Eighty-nine (62%) mentioned conservative treatment options but eleven (8%) did not include surgery in their treatment options. Only 38 sites (27%) mentioned recurrence of haemorrhoids following surgery and 28 sites (20%) did not list any complications. Overall, 19 websites (14%) were judged as being of high quality, 66 (45%) as moderate quality and 58 (40%) as low quality. Conclusions The quality of information on the internet is highly variable and a significant proportion of websites assessed are poor. The majority of websites are sponsored by private companies selling alternative treatments for haemorrhoids. Clinicians should be prepared to advise their patients which websites can provide high-quality information on the surgical treatment of haemorrhoids.

Background: Myofibroblasts have an important role in regulating the normal colorectal stem cell niche. While the activation of myofibroblasts in primary colorectal cancers has been previously described, myofibroblast activation in lymph node metastases has not been described before. Methods: Paraffin-embedded lymph node sections from patients with macrometastases, micrometastases and isolated tumour cells were stained to identify myofibroblasts and to characterise the distribution of different cell types in tumour-containing lymph nodes. The extent of myofibroblast presence was quantified and compared with the size of the metastasis and degree of proliferation and differentiation of the cancer cells. Results: We show substantial activation of myofibroblasts in the presence of colorectal metastases in lymph nodes, which is intimately associated with glandular structures, both in micro- and macrometastases. The degree of activation is positively associated with the size of the metastases and the proportion of Ki67+ve cancer cells, and negatively associated with the degree of enterocyte differentiation as measured by CK20 expression. Conclusion: The substantial activation of myofibroblasts in tumour-containing lymph nodes strongly suggests that these metastatic cancer cells are still significantly dependent on their microenvironment. Further understanding of these epithelial–mesenchymal interactions could lead to the development of new therapies in metastatic disease.

The gastrointestinal epithelium is a highly organised tissue that is constantly being renewed. In order to maintain homeostasis, the balance between intestinal stem cell (ISC) self-renewal and differentiation must be carefully regulated. In this review, we describe how the intestinal stem cell niche provides a unique environment to regulate self-renewal and differentiation of ISCs. It has traditionally been believed that the mesenchymal myofibroblasts play an important role in the crosstalk between ISCs and the niche. However, recent evidence in Drosophila and in vertebrates suggests that epithelial cells also contribute to the niche. We discuss the multiple signalling pathways that are utilised to regulate stemness within the niche, including members of the Wnt, BMP and Hedgehog pathways, and how aberrations in these signals lead to disruption of the normal crypt-villus axis. Finally, we also discuss how CDX1 and inhibition of the Notch pathway are important in specifying enterocyte and goblet cell differentiation respectively. © Springer Basel AG 2011.

Background Spinal cord injury is a serious and debilitating condition, affecting millions of people worldwide. Long seen as a permanent injury, recent advances in stem cell research have brought closer the possibility of repairing the spinal cord. One such approach involves injecting oligodendrocyte progenitor cells, derived from human embryonic stem cells, into the injured spinal cord in the hope that they will initiate repair. A phase I clinical trial of this therapy was started in mid 2010 and is currently underway. Discussion The theory underlying this approach is that these myelinating progenitors will phenotypically replace myelin lost during injury whilst helping to promote a repair environment in the lesion. However, the importance of demyelination in the pathogenesis of human spinal cord injury is a contentious issue and a body of literature suggests that it is only a minor factor in the overall injury process. Summary This review examines the validity of the theory underpinning the on-going clinical trial as well as analysing published data from animal models and finally discussing issues surrounding safety and purity in order to assess the potential of this approach to successfully treat acute human spinal cord injury.

4124 Background: Cancer stem cells are defined as cells within a tumour that are able to self-renew and differentiate into all cell lineages within that tumour. With our extensive panel of colorectal cell lines, our aims are: 1) To characterise and isolate cancer stem cells based on stem cell markers, morphological appearances and the ability to form multiple lineages; 2) To understand how cancer stem cells drive tumour growth and progression. Methods: 1) Fluorescent Activated Cell Sorting (FACS); 2) In vitro soft agar clonogenic and Matrigel differentiation assays; 3) In vivo tumourigenic NOD/SCID mice assay; 4) Confocal immunofluorescence imaging. Results: 1) A subpopulation of cells can differentiate into crypt-like megacolonies, retaining the ability to self-renew and differentiate. SW1222 cell line forms heterogeneous colonies when single cells are plated in Matrigel. Megacolonies can both self-renew and form terminally differentiated small colonies, whereas small colonies cannot form megacolonies. Megacolonies develop crypt-like structures and increase their expression of differentiation markers (CDX-1, CK-20) over time. Experiments are currently under way to confirm that cells from megacolonies are able to initiate tumours in NOD/SCID mice. Some cell lines retain the ability to differentiate into both neuroendocrine and epithelial lineages. 2) CD44+CD24+ enriches for the cancer stem cell population. Colorectal cancer cell lines HCT116, HT29, LS180, LS174T and SW1222 express both CD44 and CD24. The CD44+CD24+ subpopulation is the most clonogenic. In SW1222, CD44+CD24+ cells enrich for megacolonies and can reform all four CD44/CD24 subpopulations. 3) Hypoxia reduces differentiation, increases stem-like phenotype and enhances clonogenicity. Hypoxia increases the proportion of undifferentiated colorectal cancer cells when plated on Matrigel and increases clonogenicity. Conclusions: 1) Colorectal cancer cell lines contain subpopulations of cells that have the ability to self-renew, differentiate and drive tumour growth, and may be characterised by their cell surface markers and colony morphology. 2) CD44+CD24+ can be used as markers for colorectal cancer stem cells. 3) Hypoxia increases the stem-like phenotype of cancer cells, reduces differentiation and increases clonogenicity. No significant financial relationships to disclose.