Long term in vitro expansion of epithelial stem cells enabled by pharmacological inhibition of PAK1-ROCK-Myosin II and TGF-β signaling

Summary: Despite substantial self-renewal capability in vivo, epithelial stem and progenitor cells located in various tissues expand for a few passages in vitro in feeder-free condition before they succumb to growth arrest. Here, we describe the EpiX method, which utilizes small molecules that inhibit PAK1-ROCK-Myosin II and TGF-β signaling to achieve over one trillion-fold expansion of human epithelial stem and progenitor cells from skin, airway, mammary, and prostate glands in the absence of feeder cells. Transcriptomic and epigenomic studies show that this condition helps epithelial cells to overcome stresses for continuous proliferation. EpiX-expanded basal epithelial cells differentiate into mature epithelial cells consistent with their tissue origins. Whole-genome sequencing reveals that the cells retain remarkable genome integrity after extensive in vitro expansion without acquiring tumorigenicity. EpiX technology provides a solution to exploit the potential of tissue-resident epithelial stem and progenitor cells for regenerative medicine. : Zhang et al. screen a small-molecule collection and find that pharmacologic inhibition of TGF-β and PAK1-ROCK-Myosin II, in low calcium conditions, supports extended expansion of epithelial stem cells in 2D format. This approach enhances the potential of tissue-resident epithelial stem cells for cell therapy. Keywords: epithelial stem and progenitor cells, cell culture method, TGF-β, PAK1/ROCK/Myosin II, feeder-free, regenerative medicine, cell therap

Liver transplant outcomes after ex vivo machine perfusion: a meta-analysis

Background: The pressure on liver-transplant programmes has expanded the usage of extended-criteria allografts. Machine perfusion may be better than conventional static cold storage (SCS) in alleviating ischaemia-reperfusion injury in this setting. Recipient outcomes with hypothermic or normothermic machine perfusion were assessed against SCS here. Methods: A search in MEDLINE, EMBASE and Scopus was conducted in February 2021. Primary studies investigating ex vivo machine perfusion were assessed for the following outcomes: morbidity, ICU and hospital stay, graft and patient survival rates and relative costs. Meta-analysis was performed to obtain pooled summary measures. Results: Thirty-four articles involving 1742 patients were included, of which 20 were used for quantitative synthesis. Odds ratios favoured hypothermic machine perfusion (over SCS) with less early allograft dysfunction, ischaemic cholangiopathy, non-anastomotic strictures and graft loss. Hypothermic machine perfusion was associated with a shorter hospital stay and normothermic machine perfusion with reduced graft injury. Two randomized clinical trials found normothermic machine perfusion reduced major complication risks. Conclusion: Machine perfusion assists some outcomes with potential cost savings

Longitudinal Bioluminescence Imaging of Primary Versus Abdominal Metastatic Tumor Growth in Orthotopic Pancreatic Tumor Models in NSG Mice

Objectives: The purpose of the present study was to develop and validate noninvasive bioluminescence imaging methods for differentially monitoring primary and abdominal metastatic tumor growth in mouse orthotopic models of pancreatic cancer. Methods: A semiautomated maximum entropy segmentation method was implemented for the primary tumor region of interest, and a rule-based method for manually drawing a region of interest for the abdominal metastatic region was developed for monitoring tumor growth in orthotopic models of pancreatic cancer. The 2 region-of-interest methods were validated by having 2 observers independently segment Panc-1 tumors, and the results were compared with the number of mesenteric lymph node nodules and histopathologic assessment of liver metastases. The findings were extended to orthotopic tumors of the more metastatic MIA PaCa-2 and AsPC-1 cells where separate groups of animals were implanted with different numbers of cells. Results: The results demonstrated that the segmentation methods were highly reliable, reproducible, and robust and allowed statistically significant discrimination in the growth rates of primary and abdominal metastatic tumors of different cell lines implanted with different numbers of cells. Conclusions: The present results demonstrate that primary tumors and abdominal metastatic foci in orthotopic pancreatic cancer models can be reliably quantified separately and noninvasively over time with bioluminescence imaging

Management of patients at the hepatopancreatobiliary unit of a London teaching hospital during the COVID-19 pandemic

To mitigate COVID-19-related shortage of treatment capacity, the hepatopancreatobiliary (HPB) unit of the Royal Free Hospital London (RFHL) transferred its practice to independent hospitals in Central London through the North Central London Cancer Alliance. The aim of this study was to critically assess this strategy and evaluate perioperative outcomes. Prospectively collected data were reviewed on all patients who were treated under the RFHL HPB unit in six hospitals between November 2020 and October 2021. A total of 1541 patients were included, as follows: 1246 (81%) at the RFHL, 41 (3%) at the Chase Farm Hospital, 23 (2%) at the Whittington Hospital, 207 (13%) at the Princess Grace Hospital, 12 (1%) at the Wellington Hospital and 12 (1%) at the Lister Hospital, Chelsea. Across all institutions, overall complication rate were 40%, major complication (Clavien-Dindo grade ≥ 3a) rate were 11% and mortality rates were 1.4%, respectively. In COVID-19-positive patients (n = 28), compared with negative patients, complication rate and mortality rates were increased tenfold. Outsourcing HPB patients, including their specialist care, to surrounding institutions was safe and ensured ongoing treatment with comparable outcomes among the institutions during the COVID-19 pandemic. Due to the lack of direct comparison with a non-pandemic cohort, these results can strictly only be applied within a pandemic setting

Elective Cancer Surgery in COVID-19-Free Surgical Pathways During the SARS-CoV-2 Pandemic: An International, Multicenter, Comparative Cohort Study.

PURPOSE: As cancer surgery restarts after the first COVID-19 wave, health care providers urgently require data to determine where elective surgery is best performed. This study aimed to determine whether COVID-19-free surgical pathways were associated with lower postoperative pulmonary complication rates compared with hospitals with no defined pathway. PATIENTS AND METHODS: This international, multicenter cohort study included patients who underwent elective surgery for 10 solid cancer types without preoperative suspicion of SARS-CoV-2. Participating hospitals included patients from local emergence of SARS-CoV-2 until April 19, 2020. At the time of surgery, hospitals were defined as having a COVID-19-free surgical pathway (complete segregation of the operating theater, critical care, and inpatient ward areas) or no defined pathway (incomplete or no segregation, areas shared with patients with COVID-19). The primary outcome was 30-day postoperative pulmonary complications (pneumonia, acute respiratory distress syndrome, unexpected ventilation). RESULTS: Of 9,171 patients from 447 hospitals in 55 countries, 2,481 were operated on in COVID-19-free surgical pathways. Patients who underwent surgery within COVID-19-free surgical pathways were younger with fewer comorbidities than those in hospitals with no defined pathway but with similar proportions of major surgery. After adjustment, pulmonary complication rates were lower with COVID-19-free surgical pathways (2.2% v 4.9%; adjusted odds ratio [aOR], 0.62; 95% CI, 0.44 to 0.86). This was consistent in sensitivity analyses for low-risk patients (American Society of Anesthesiologists grade 1/2), propensity score-matched models, and patients with negative SARS-CoV-2 preoperative tests. The postoperative SARS-CoV-2 infection rate was also lower in COVID-19-free surgical pathways (2.1% v 3.6%; aOR, 0.53; 95% CI, 0.36 to 0.76). CONCLUSION: Within available resources, dedicated COVID-19-free surgical pathways should be established to provide safe elective cancer surgery during current and before future SARS-CoV-2 outbreaks

Elective cancer surgery in COVID-19-free surgical pathways during the SARS-CoV-2 pandemic: An international, multicenter, comparative cohort study

PURPOSE As cancer surgery restarts after the first COVID-19 wave, health care providers urgently require data to determine where elective surgery is best performed. This study aimed to determine whether COVID-19–free surgical pathways were associated with lower postoperative pulmonary complication rates compared with hospitals with no defined pathway. PATIENTS AND METHODS This international, multicenter cohort study included patients who underwent elective surgery for 10 solid cancer types without preoperative suspicion of SARS-CoV-2. Participating hospitals included patients from local emergence of SARS-CoV-2 until April 19, 2020. At the time of surgery, hospitals were defined as having a COVID-19–free surgical pathway (complete segregation of the operating theater, critical care, and inpatient ward areas) or no defined pathway (incomplete or no segregation, areas shared with patients with COVID-19). The primary outcome was 30-day postoperative pulmonary complications (pneumonia, acute respiratory distress syndrome, unexpected ventilation). RESULTS Of 9,171 patients from 447 hospitals in 55 countries, 2,481 were operated on in COVID-19–free surgical pathways. Patients who underwent surgery within COVID-19–free surgical pathways were younger with fewer comorbidities than those in hospitals with no defined pathway but with similar proportions of major surgery. After adjustment, pulmonary complication rates were lower with COVID-19–free surgical pathways (2.2% v 4.9%; adjusted odds ratio [aOR], 0.62; 95% CI, 0.44 to 0.86). This was consistent in sensitivity analyses for low-risk patients (American Society of Anesthesiologists grade 1/2), propensity score–matched models, and patients with negative SARS-CoV-2 preoperative tests. The postoperative SARS-CoV-2 infection rate was also lower in COVID-19–free surgical pathways (2.1% v 3.6%; aOR, 0.53; 95% CI, 0.36 to 0.76). CONCLUSION Within available resources, dedicated COVID-19–free surgical pathways should be established to provide safe elective cancer surgery during current and before future SARS-CoV-2 outbreaks

Mortality and pulmonary complications in patients undergoing surgery with perioperative SARS-CoV-2 infection: an international cohort study

Background: The impact of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) on postoperative recovery needs to be understood to inform clinical decision making during and after the COVID-19 pandemic. This study reports 30-day mortality and pulmonary complication rates in patients with perioperative SARS-CoV-2 infection. Methods: This international, multicentre, cohort study at 235 hospitals in 24 countries included all patients undergoing surgery who had SARS-CoV-2 infection confirmed within 7 days before or 30 days after surgery. The primary outcome measure was 30-day postoperative mortality and was assessed in all enrolled patients. The main secondary outcome measure was pulmonary complications, defined as pneumonia, acute respiratory distress syndrome, or unexpected postoperative ventilation. Findings: This analysis includes 1128 patients who had surgery between Jan 1 and March 31, 2020, of whom 835 (74·0%) had emergency surgery and 280 (24·8%) had elective surgery. SARS-CoV-2 infection was confirmed preoperatively in 294 (26·1%) patients. 30-day mortality was 23·8% (268 of 1128). Pulmonary complications occurred in 577 (51·2%) of 1128 patients; 30-day mortality in these patients was 38·0% (219 of 577), accounting for 81·7% (219 of 268) of all deaths. In adjusted analyses, 30-day mortality was associated with male sex (odds ratio 1·75 [95% CI 1·28–2·40], p\textless0·0001), age 70 years or older versus younger than 70 years (2·30 [1·65–3·22], p\textless0·0001), American Society of Anesthesiologists grades 3–5 versus grades 1–2 (2·35 [1·57–3·53], p\textless0·0001), malignant versus benign or obstetric diagnosis (1·55 [1·01–2·39], p=0·046), emergency versus elective surgery (1·67 [1·06–2·63], p=0·026), and major versus minor surgery (1·52 [1·01–2·31], p=0·047). Interpretation: Postoperative pulmonary complications occur in half of patients with perioperative SARS-CoV-2 infection and are associated with high mortality. Thresholds for surgery during the COVID-19 pandemic should be higher than during normal practice, particularly in men aged 70 years and older. Consideration should be given for postponing non-urgent procedures and promoting non-operative treatment to delay or avoid the need for surgery. Funding: National Institute for Health Research (NIHR), Association of Coloproctology of Great Britain and Ireland, Bowel and Cancer Research, Bowel Disease Research Foundation, Association of Upper Gastrointestinal Surgeons, British Association of Surgical Oncology, British Gynaecological Cancer Society, European Society of Coloproctology, NIHR Academy, Sarcoma UK, Vascular Society for Great Britain and Ireland, and Yorkshire Cancer Research

Reducing the environmental impact of surgery on a global scale: systematic review and co-prioritization with healthcare workers in 132 countries

Abstract Background Healthcare cannot achieve net-zero carbon without addressing operating theatres. The aim of this study was to prioritize feasible interventions to reduce the environmental impact of operating theatres. Methods This study adopted a four-phase Delphi consensus co-prioritization methodology. In phase 1, a systematic review of published interventions and global consultation of perioperative healthcare professionals were used to longlist interventions. In phase 2, iterative thematic analysis consolidated comparable interventions into a shortlist. In phase 3, the shortlist was co-prioritized based on patient and clinician views on acceptability, feasibility, and safety. In phase 4, ranked lists of interventions were presented by their relevance to high-income countries and low–middle-income countries. Results In phase 1, 43 interventions were identified, which had low uptake in practice according to 3042 professionals globally. In phase 2, a shortlist of 15 intervention domains was generated. In phase 3, interventions were deemed acceptable for more than 90 per cent of patients except for reducing general anaesthesia (84 per cent) and re-sterilization of ‘single-use’ consumables (86 per cent). In phase 4, the top three shortlisted interventions for high-income countries were: introducing recycling; reducing use of anaesthetic gases; and appropriate clinical waste processing. In phase 4, the top three shortlisted interventions for low–middle-income countries were: introducing reusable surgical devices; reducing use of consumables; and reducing the use of general anaesthesia. Conclusion This is a step toward environmentally sustainable operating environments with actionable interventions applicable to both high– and low–middle–income countries

Functional Profiling of CFTR-Directed Therapeutics Using Pediatric Patient-Derived Nasal Epithelial Cell Models

Functional profiling of CFTR-directed therapeutics offers the potential to provide significant benefits to young people with cystic fibrosis (CF). However, the development of 2D airway epithelial cell models for individual response tests in CF children remains a central task. The objective of this study was to determine the utility of EpiXTM technology for expansion of nasal epithelial cells for use in electrophysiological CFTR function measurements. An initial harvest of as few as 20,000 cells was sufficient to expand up to 50 million cells that were used to generate air-liquid interface (ALI) cultures for ion transport studies with the Ussing assay. CFTR function was assessed by measuring responses to forskolin and the CFTR potentiator VX-770 (ivacaftor) in ALI cultures generated from passage 3 and 4 cells. Short-circuit current (Isc) measurements of blocked CFTR currents (ΔICFTRinh) discriminated CFTR function between healthy control (wild type, WT) and patients with intermediate (F508del/R117H-7T: 56% WT) and severe (F508del/F508del: 12% WT) CF disease. For the mixed genotypes, CFTR activity for F508del/c.850dupA was 12% WT, R334W/406-1G>A was 24% WT, and CFTRdele2,3(21 kb)/CFTRdele2,3(21 kb) was 9% WT. The CFTR correctors VX-809 (lumacaftor) and VX-661 (tezacaftor) significantly increased CFTR currents for F508del/R117H to 73 and 67% WT, respectively. Cultures with the large deletion mutation CFTRdele2,3(21 kb) unexpectedly responded to VX-661 treatment (20% WT). Amiloride-sensitive sodium currents were robust and ranged between 20-80 μA/cm2 depending on the subject. In addition to characterizing the electrophysiological profile of mutant CFTR activity in cultures for five genotypes, our study exemplifies the promising paradigm of bed-to-bench side cooperation and personalized medicine