AQP5 enriches for stem cells and cancer origins in the distal stomach.

LGR5 marks resident adult epithelial stem cells at the gland base in the mouse pyloric stomach1, but the identity of the equivalent human stem cell population remains unknown owing to a lack of surface markers that facilitate its prospective isolation and validation. In mouse models of intestinal cancer, LGR5+ intestinal stem cells are major sources of cancer following hyperactivation of the WNT pathway2. However, the contribution of pyloric LGR5+ stem cells to gastric cancer following dysregulation of the WNT pathway-a frequent event in gastric cancer in humans3-is unknown. Here we use comparative profiling of LGR5+ stem cell populations along the mouse gastrointestinal tract to identify, and then functionally validate, the membrane protein AQP5 as a marker that enriches for mouse and human adult pyloric stem cells. We show that stem cells within the AQP5+ compartment are a source of WNT-driven, invasive gastric cancer in vivo, using newly generated Aqp5-creERT2 mouse models. Additionally, tumour-resident AQP5+ cells can selectively initiate organoid growth in vitro, which indicates that this population contains potential cancer stem cells. In humans, AQP5 is frequently expressed in primary intestinal and diffuse subtypes of gastric cancer (and in metastases of these subtypes), and often displays altered cellular localization compared with healthy tissue. These newly identified markers and mouse models will be an invaluable resource for deciphering the early formation of gastric cancer, and for isolating and characterizing human-stomach stem cells as a prerequisite for harnessing the regenerative-medicine potential of these cells in the clinic.

ISM1 protects lung homeostasis via cell-surface GRP78-mediated alveolar macrophage apoptosis.

Alveolar macrophages (AMs) are critical for lung immune defense and homeostasis. They are orchestrators of chronic obstructive pulmonary disease (COPD), with their number significantly increased and functions altered in COPD. However, it is unclear how AM number and function are controlled in a healthy lung and if changes in AMs without environmental assault are sufficient to trigger lung inflammation and COPD. We report here that absence of isthmin 1 (ISM1) in mice (Ism1-/- ) leads to increase in both AM number and functional heterogeneity, with enduring lung inflammation, progressive emphysema, and significant lung function decline, phenotypes similar to human COPD. We reveal that ISM1 is a lung resident anti-inflammatory protein that selectively triggers the apoptosis of AMs that harbor high levels of its receptor cell-surface GRP78 (csGRP78). csGRP78 is present at a heterogeneous level in the AMs of a healthy lung, but csGRP78high AMs are expanded in Ism1-/- mice, cigarette smoke (CS)-induced COPD mice, and human COPD lung, making these cells the prime targets of ISM1-mediated apoptosis. We show that csGRP78high AMs mostly express MMP-12, hence proinflammatory. Intratracheal delivery of recombinant ISM1 (rISM1) depleted csGRP78high AMs in both Ism1-/- and CS-induced COPD mice, blocked emphysema development, and preserved lung function. Consistently, ISM1 expression in human lungs positively correlates with AM apoptosis, suggesting similar function of ISM1-csGRP78 in human lungs. Our findings reveal that AM apoptosis regulation is an important physiological mechanism for maintaining lung homeostasis and demonstrate the potential of pulmonary-delivered rISM1 to target csGRP78 as a therapeutic strategy for COPD.

CEACAM5 and TROP2 define metaplastic and dysplastic transitions in human antral gastric precancerous lesions and tumors.

BACKGROUND: Mucosal gastric atrophy and intestinal metaplasia (IM) increase the risk for the development of gastric cancer (GC) as they represent a field for development of dysplasia and intestinal-type gastric adenocarcinoma. METHODS: We have investigated the expression of two dysplasia markers, CEACAM5 and TROP2, in human antral IM and gastric tumors to assess their potential as molecular markers. RESULTS: In the normal antral mucosa, weak CEACAM5 and TROP2 expression was only observed in the foveolar epithelium, while inflamed antrum exhibited increased expression of both markers. Complete IM exhibited weak CEACAM5 expression at the apical surface, but no basolateral TROP2 expression. On the other hand, incomplete IM demonstrated high levels of both CEACAM5 and TROP2 expression. Notably, incomplete IM with dysplastic morphology (dysplastic incomplete IM) exhibited higher levels of CEACAM5 and TROP2 expression compared to incomplete IM without dysplastic features (simple incomplete IM). In addition, dysplastic incomplete IM showed diminished SOX2 and elevated CDX2 expression compared to simple incomplete IM. CEACAM5 and TROP2 positivity in incomplete IM was similar to that of gastric adenomas and GC. Significant association was found between CEACAM5 and TROP2 positivity and histology of GC. CONCLUSIONS: These findings support the concept that incomplete IM is more likely associated with GC development. Overall, our study provides evidence of the heterogeneity of gastric IM and the distinct expression profiles of CEACAM5 and TROP2 in dysplastic incomplete IM. Our findings support the potential use of CEACAM5 and TROP2 as molecular markers for identifying individuals with a higher risk of GC development in the context of incomplete IM.

Regulatory enhancer profiling of mesenchymal-type gastric cancer reveals subtype-specific epigenomic landscapes and targetable vulnerabilities.

OBJECTIVE: Gastric cancer (GC) comprises multiple molecular subtypes. Recent studies have highlighted mesenchymal-subtype GC (Mes-GC) as a clinically aggressive subtype with few treatment options. Combining multiple studies, we derived and applied a consensus Mes-GC classifier to define the Mes-GC enhancer landscape revealing disease vulnerabilities. DESIGN: Transcriptomic profiles of ~1000 primary GCs and cell lines were analysed to derive a consensus Mes-GC classifier. Clinical and genomic associations were performed across >1200 patients with GC. Genome-wide epigenomic profiles (H3K27ac, H3K4me1 and assay for transposase-accessible chromatin with sequencing (ATAC-seq)) of 49 primary GCs and GC cell lines were generated to identify Mes-GC-specific enhancer landscapes. Upstream regulators and downstream targets of Mes-GC enhancers were interrogated using chromatin immunoprecipitation followed by sequencing (ChIP-seq), RNA sequencing, CRISPR/Cas9 editing, functional assays and pharmacological inhibition. RESULTS: We identified and validated a 993-gene cancer-cell intrinsic Mes-GC classifier applicable to retrospective cohorts or prospective single samples. Multicohort analysis of Mes-GCs confirmed associations with poor patient survival, therapy resistance and few targetable genomic alterations. Analysis of enhancer profiles revealed a distinctive Mes-GC epigenomic landscape, with TEAD1 as a master regulator of Mes-GC enhancers and Mes-GCs exhibiting preferential sensitivity to TEAD1 pharmacological inhibition. Analysis of Mes-GC super-enhancers also highlighted NUAK1 kinase as a downstream target, with synergistic effects observed between NUAK1 inhibition and cisplatin treatment. CONCLUSION: Our results establish a consensus Mes-GC classifier applicable to multiple transcriptomic scenarios. Mes-GCs exhibit a distinct epigenomic landscape, and TEAD1 inhibition and combinatorial NUAK1 inhibition/cisplatin may represent potential targetable options.

A real-life comparison of hypoglycaemia symptomatology between insulin-treated type 2 diabetes participants with and without impaired awareness of hypoglycaemia.

AIMS: We aim to compare and correlate Gold and Clarke questionnaire scores with hypoglycaemic symptomatic responses between insulin-treated type 2 diabetes participants with and without IAH in a real-life study. METHODS: Insulin-treated type 2 diabetes participants attending an outpatient diabetes clinic in Singapore were asked to complete the Gold and Clarke questionnaires, record capillary blood glucose (CBG) and hypoglycaemic symptoms for 4 weeks. RESULTS: Data were collected from 153 participants (M:F = 98:55) with mean age 61.0 ± 9.4 years, duration of diabetes 19.5 ± 8.8 years and HbA1c 68 ± 17 mmol/mol (8.4 ± 1.5%). Gold and Clarke methods classified 19.6% and 26.8% of participants with IAH, respectively. Using CBG threshold of <3 mmol/L, significantly greater proportion of participants with intact awareness were experiencing autonomic symptoms than those with IAH with either method (Gold: 69% vs. 18%, p = 0.006; Clarke: 85% vs. 46%, p = 0.010). Significantly greater proportion of participants with IAH experienced no hypoglycaemia symptoms than those with intact awareness (Gold: 3.4% vs. 36%, p = 0.015; Clarke: 3.7% vs. 31%, p = 0.031). Participants with IAH had significantly higher rates of severe hypoglycaemia in the preceding year compared to those without (Gold: 17% vs. 3.3%; Clarke: 15% vs. 2.7%, p = 0.012). CONCLUSIONS: Gold and Clarke questionnaires are appropriate tools in ascertaining IAH status in insulin-treated type 2 diabetes participants. This is the first time whereby the hypoglycaemia symptomology has robustly validated the Gold and Clarke questionnaire in insulin-treated type 2 diabetes participants.

Evaluation of Care Outcomes of Patients Receiving Hyperkalemia Treatment With Insulin in Acute Care Tertiary Hospital Emergency Department.

INTRODUCTION: Treatment of hyperkalemia using intravenous insulin can result in severe hypoglycemia, but regular blood glucose monitoring is not standardized. This study aimed to (i) explore the demographics of adult patients receiving hyperkalemia treatment and (ii) identify the incidence rate of hypoglycemia and associated demographic or clinical characteristics. METHODS: A descriptive design with prospective data collection was used. This study recruited 135 patients who received hyperkalemia treatment in the emergency department. Structured blood glucose monitoring was conducted at 1, 2, 4, and 6 hours after receiving intravenous insulin. Univariate analyses of association between demographic and clinical variables and hypoglycemia outcome were performed. RESULTS: There were 31 hypoglycemic events, with 11.9%, 7.4%, 2.2%, and 1.5% occurring at the 1, 2, 4, and 6 hours after treatment. The logit regression showed no significantly increased risk of hypoglycemia in terms of the demographic and clinical variables. DISCUSSION: The variation in blood glucose response observed in this study combined with the high incidences of hypolycaemia indicated the need for frequent and longer duration of monitoring for patients who were being treated for hyperkalaemia with IDT.

Severity of gastric intestinal metaplasia predicts the risk of gastric cancer: a prospective multicentre cohort study (GCEP).

OBJECTIVE: To investigate the incidence of gastric cancer (GC) attributed to gastric intestinal metaplasia (IM), and validate the Operative Link on Gastric Intestinal Metaplasia (OLGIM) for targeted endoscopic surveillance in regions with low-intermediate incidence of GC. METHODS: A prospective, longitudinal and multicentre study was carried out in Singapore. The study participants comprised 2980 patients undergoing screening gastroscopy with standardised gastric mucosal sampling, from January 2004 and December 2010, with scheduled surveillance endoscopies at year 3 and 5. Participants were also matched against the National Registry of Diseases Office for missed diagnoses of early gastric neoplasia (EGN). RESULTS: There were 21 participants diagnosed with EGN. IM was a significant risk factor for EGN (adjusted-HR 5.36; 95% CI 1.51 to 19.0; p<0.01). The age-adjusted EGN incidence rates for patients with and without IM were 133.9 and 12.5 per 100 000 person-years. Participants with OLGIM stages III-IV were at greatest risk (adjusted-HR 20.7; 95% CI 5.04 to 85.6; p<0.01). More than half of the EGNs (n=4/7) attributed to baseline OLGIM III-IV developed within 2 years (range: 12.7-44.8 months). Serum trefoil factor 3 distinguishes (Area Under the Receiver Operating Characteristics 0.749) patients with OLGIM III-IV if they are negative for H. pylori. Participants with OLGIM II were also at significant risk of EGN (adjusted-HR 7.34; 95% CI 1.60 to 33.7; p=0.02). A significant smoking history further increases the risk of EGN among patients with OLGIM stages II-IV. CONCLUSIONS: We suggest a risk-stratified approach and recommend that high-risk patients (OLGIM III-IV) have endoscopic surveillance in 2 years, intermediate-risk patients (OLGIM II) in 5 years.

Induction of Gastric Cancer by Successive Oncogenic Activation in the Corpus.

BACKGROUND & AIMS: Metaplasia and dysplasia in the corpus are reportedly derived from de-differentiation of chief cells. However, the cellular origin of metaplasia and cancer remained uncertain. Therefore, we investigated whether pepsinogen C (PGC) transcript-expressing cells represent the cellular origin of metaplasia and cancer using a novel Pgc-specific CreERT2 recombinase mouse model. METHODS: We generated a Pgc-mCherry-IRES-CreERT2 (Pgc-CreERT2) knock-in mouse model. Pgc-CreERT2/+ and Rosa-EYFP mice were crossed to generate Pgc-CreERT2/Rosa-EYFP (Pgc-CreERT2/YFP) mice. Gastric tissues were collected, followed by lineage-tracing experiments and histologic and immunofluorescence staining. We further established Pgc-CreERT2;KrasG12D/+ mice and investigated whether PGC transcript-expressing cells are responsible for the precancerous state in gastric glands. To investigate cancer development from PGC transcript-expressing cells with activated Kras, inactivated Apc, and Trp53 signaling pathways, we crossed Pgc-CreERT2/+ mice with conditional KrasG12D, Apcflox, Trp53flox mice. RESULTS: Expectedly, mCherry mainly labeled chief cells in the Pgc-CreERT2 mice. However, mCherry was also detected throughout the neck cell and isthmal stem/progenitor regions, albeit at lower levels. In the Pgc-CreERT2;KrasG12D/+ mice, PGC transcript-expressing cells with KrasG12D/+ mutation presented pseudopyloric metaplasia. The early induction of proliferation at the isthmus may reflect the ability of isthmal progenitors to react rapidly to Pgc-driven KrasG12D/+ oncogenic mutation. Furthermore, Pgc-CreERT2;KrasG12D/+;Apcflox/flox mice presented intramucosal dysplasia/carcinoma and Pgc-CreERT2;KrasG12D/+;Apcflox/flox;Trp53flox/flox mice presented invasive and metastatic gastric carcinoma. CONCLUSIONS: The Pgc-CreERT2 knock-in mouse is an invaluable tool to study the effects of successive oncogenic activation in the mouse corpus. Time-course observations can be made regarding the responses of isthmal and chief cells to oncogenic insults. We can observe stomach-specific tumorigenesis from the beginning to metastatic development.

WBP2 promotes gastric cancer cell migration via novel targeting of LATS2 kinase in the Hippo tumor suppressor pathway.

Dysregulation of signaling pathways is responsible for many human diseases. The lack of understanding of the molecular etiology of gastric cancer (GC) poses a substantial challenge to the development of effective cancer therapy. To better understand the molecular mechanisms underlying the pathogenesis of GC, which will facilitate the identification and development of effective therapeutic approaches to improve patient outcomes, mass spectrometry-based phosphoproteomics analysis was performed to map the global molecular changes in GC. A total of 530 proteins with altered phosphorylation levels were detected across a panel of 15 normal and GC cell lines. WW domain-binding protein 2 (WBP2) was validated to be upregulated in a subset of GC cell lines. WBP2 is overexpressed in 61% cases of GC compared to non-cancer tissues and high WBP2 expression correlates with poor clinical outcomes. WBP2 was found to be required for GC cell migration but is dispensable for cell growth and proliferation. WBP2 knockdown increased p-LATS2 with a concomitant increase in p-YAP, resulting in the cytoplasmic retention of YAP and ultimately the inhibition of YAP/TEAD activity and downregulation of TEAD target genes--CTGF and CYR61. Importantly, the loss of LATS2 reversed the activation of Hippo pathway caused by WBP2 knockdown, indicating that WBP2 acts through LATS2 to exert its function on the Hippo pathway. Moreover, WBP2 interacted with LATS2 to inhibit its phosphorylation and activity. In conclusion, our study established a pivotal role for WBP2 in the promotion of GC cell migration via a novel mechanism that inactivates the Hippo pathway transducer LATS2.

Iqgap3-Ras axis drives stem cell proliferation in the stomach corpus during homoeostasis and repair.

OBJECTIVE: Tissue stem cells are central regulators of organ homoeostasis. We looked for a protein that is exclusively expressed and functionally involved in stem cell activity in rapidly proliferating isthmus stem cells in the stomach corpus. DESIGN: We uncovered the specific expression of Iqgap3 in proliferating isthmus stem cells through immunofluorescence and in situ hybridisation. We performed lineage tracing and transcriptomic analysis of Iqgap3 +isthmus stem cells with the Iqgap3-2A-tdTomato mouse model. Depletion of Iqgap3 revealed its functional importance in maintenance and proliferation of stem cells. We further studied Iqgap3 expression and the associated gene expression changes during tissue repair after tamoxifen-induced damage. Immunohistochemistry revealed elevated expression of Iqgap3 in proliferating regions of gastric tumours from patient samples. RESULTS: Iqgap3 is a highly specific marker of proliferating isthmus stem cells during homoeostasis. Iqgap3+isthmus stem cells give rise to major cell types of the corpus unit. Iqgap3 expression is essential for the maintenance of stem potential. The Ras pathway is a critical partner of Iqgap3 in promoting strong proliferation in isthmus stem cells. The robust induction of Iqgap3 expression following tissue damage indicates an active role for Iqgap3 in tissue regeneration. CONCLUSION: IQGAP3 is a major regulator of stomach epithelial tissue homoeostasis and repair. The upregulation of IQGAP3 in gastric cancer suggests that IQGAP3 plays an important role in cancer cell proliferation.

DNA damage signalling as an anti-cancer barrier in gastric intestinal metaplasia.

OBJECTIVE: Intestinal metaplasia (IM) is a premalignant stage that poses a greater risk for subsequent gastric cancer (GC). However, factors regulating IM to GC progression remain unclear. Previously, activated DNA damage response (DDR) signalling factors were shown to engage tumour-suppressive networks in premalignant lesions. Here, we interrogate the relationship of DDR signalling to mutational accumulation in IM lesions. DESIGN: IM biopsies were procured from the gastric cancer epidemiology programme, an endoscopic surveillance programme where biopsies have been subjected to (epi)genomic characterisation. IM samples were classified as genome-stable or genome-unstable based on their mutational burden/somatic copy-number alteration (CNA) profiles. Samples were probed for DDR signalling and cell proliferation, using the markers γH2AX and MCM2, respectively. The expression of the gastric stem cell marker, CD44v9, was also assessed. Tissue microarrays representing the GC progression spectrum were included. RESULTS: MCM2-positivity increased during GC progression, while γH2AX-positivity showed modest increase from normal to gastritis and IM stages, with further increase in GC. γH2AX levels correlated with the extent of chronic inflammation. Interestingly, genome-stable IM lesions had higher γH2AX levels underscoring a protective anti-cancer role for DDR signalling. In contrast, genome-unstable IM lesions with higher mutational burden/CNAs had lower γH2AX levels, elevated CD44v9 expression and modest promoter hypermethylation of DNA repair genes WRN, MLH1 and RAD52. CONCLUSIONS: Our data suggest that IM lesions with active DDR will likely experience a longer latency at the premalignant state until additional hits that override DDR signalling clonally expand and promote progression. These observations provide insights on the factors governing IM progression.

An LCM-based genomic analysis of SPEM, Gastric Cancer and Pyloric Gland Adenoma in an Asian cohort.

Spasmolytic polypeptide-expressing metaplasia (SPEM) and pyloric gland adenoma (PGA) in the stomach are metaplastic and neoplastic lesions, respectively, in which gastric body glands are replaced by pyloric glands. The aim of this study was to evaluate the genomic profile of SPEM and compare it with intestinal-type gastric cancer (GC) and PGA. Thirteen gastrectomies showing PGA with or without dysplasia, GC and SPEM were retrospectively selected. MUC5AC, MUC6, gastrin, and TFF2 IHC were performed. Lesions were subjected to laser capture microdissection followed by DNA extraction. Forty-three DNA samples were extracted from PGA without cytological dysplasia, PGA with low-grade and high-grade dysplasia and pyloric gland adenocarcinoma, GC, SPEM, and adjacent normal tissue from the body of the stomach and were subjected to exome sequencing for 49 genes that are commonly dysregulated in GC. Sanger sequencing was performed for confirmation. Twenty nonsynonymous mutations were identified in SPEM, and none of these were frameshifts or indels. PGA with or without cytological dysplasia showed a significantly higher number of mutations compared with SPEM. As cytological dysplasia increased from no dysplasia to dysplasia in PGA, the percentage of frameshift mutations, indels, and missense variations increased. Further missense or frameshift mutations were observed in the KRAS, APC, TP53, and CTNNB1 genes in the PGA group. In GC, mutations were observed in the TP53 gene (p.Arg248Gln). Missense mutations in the MUC5AC, KRAS, BRAF, and EZH2 genes were common between SPEM and GC. SPEM showed fewer genomic variations than GC and PGA, and was genomically distinct from the pyloric epithelium in PGA. Stepwise progression of PGA from PGA without dysplasia to PGA with dysplasia/adenocarcinoma was associated an increase in mutations. SPEM appears to be more genomically similar to GC than PGA.

Clustering of lifestyle risk behaviours and its determinants among school-going adolescents in a middle-income country: a cross-sectional study.

BACKGROUND: Lifestyle risk behaviours such as smoking, alcohol consumption, physical inactivity, sedentary behaviour and low fruit/vegetable intake have been identified as the major causes of chronic diseases. Such behaviours are usually instigated in adolescence and tend to persist into adulthood. Studies on the clustering of lifestyle risk behaviours among adolescents are scarce, particularly in developing countries. Therefore, the present paper aimed to determine the clustering of lifestyle risk behaviours and its determinants among school-going adolescents in Malaysia. METHODS: Data were extracted from a cross-sectional study, the Malaysian Adolescent Health Risk Behaviour (MyAHRB) study, which was conducted from May to September 2013 across 11 states in Peninsular Malaysia. A two-stage proportionate-to-size sampling method was employed to select a total of 3578 school-going adolescents aged 16-17 years from 20 selected schools in urban and rural settlements, respectively. The MyAHRB study adopted a set of self-administered questionnaires adapted from the Global School-based Student's Health Survey (GSHS) and the Youth Risk Behaviour Surveillance. RESULTS: The results from the analysis of 2991 school-going adolescents aged 16-17 years showed that 16 (in boys) and 15 (in girls) out of 32 combinations of lifestyle risk behaviours clustered. Girls (aOR 2.82, 95% CI: 2.32-3.43) were significantly more likely to have clustered risk behaviours than boys; however, no significant associated factors were observed among girls. In contrast, boys of Malay descent (aOR 0.64, 95% CI: 0.46-0.89) or boys who had at least three friends (aOR 0.65, 95% CI: 0.43-0.99) were less likely to engage in multiple risk behaviours. CONCLUSION: The present study demonstrated the clustering of multiple risk behaviours that occurred in both genders; these results suggest that multiple behaviour intervention programmes, instead of programmes based on siloed approaches, should be advocated and targeted to the high-risk sub-populations identified in the present study.

The impact of hypoglycaemia awareness status on regional brain responses to acute hypoglycaemia in men with type 1 diabetes.

AIMS/HYPOTHESIS: Impaired awareness of hypoglycaemia (IAH) in type 1 diabetes increases the risk of severe hypoglycaemia sixfold and can be resistant to intervention. We explored the impact of IAH on central responses to hypoglycaemia to investigate the mechanisms underlying barriers to therapeutic intervention. METHODS: We conducted [15O]water positron emission tomography studies of regional brain perfusion during euglycaemia (target 5 mmol/l), hypoglycaemia (achieved level, 2.4 mmol/l) and recovery (target 5 mmol/l) in 17 men with type 1 diabetes: eight with IAH, and nine with intact hypoglycaemia awareness (HA). RESULTS: Hypoglycaemia with HA was associated with increased activation in brain regions including the thalamus, insula, globus pallidus (GP), anterior cingulate cortex (ACC), orbital cortex, dorsolateral frontal (DLF) cortex, angular gyrus and amygdala; deactivation occurred in the temporal and parahippocampal regions. IAH was associated with reduced catecholamine and symptom responses to hypoglycaemia vs HA (incremental AUC: autonomic scores, 26.2 ± 35.5 vs 422.7 ± 237.1; neuroglycopenic scores, 34.8 ± 88.8 vs 478.9 ± 311.1; both p < 0.002). There were subtle differences (p < 0.005, k ≥ 50 voxels) in brain activation at hypoglycaemia, including early differences in the right central operculum, bilateral medial orbital (MO) cortex, and left posterior DLF cortex, with additional differences in the ACC, right GP and post- and pre-central gyri in established hypoglycaemia, and lack of deactivation in temporal regions in established hypoglycaemia. CONCLUSIONS/INTERPRETATION: Differences in activation in the post- and pre-central gyri may be expected in people with reduced subjective responses to hypoglycaemia. Alterations in the activity of regions involved in the drive to eat (operculum), emotional salience (MO cortex), aversion (GP) and recall (temporal) suggest differences in the perceived importance and urgency of responses to hypoglycaemia in IAH compared with HA, which may be key to the persistence of the condition.

Identification of Stem Cells in the Epithelium of the Stomach Corpus and Antrum of Mice.

BACKGROUND & AIMS: Little is known about the mechanisms of gastric carcinogenesis, partly because it has been a challenge to identify characterize gastric stem cells. Runx genes regulate development and their products are transcription factors associated with cancer development. A Runx1 enhancer element, eR1, is a marker of hematopoietic stem cells. We studied expression from eR1 in the stomach and the roles of gastric stem cells in gastric carcinogenesis in transgenic mice. METHODS: We used in situ hybridization and immunofluorescence analyses to study expression of Runx1 in gastric tissues from C57BL/6 (control) mice. We then created mice that expressed enhanced green fluorescent protein (EGFP) or CreERT2 under the control of eR1 (eR1-CreERT2;Rosa-Lox-Stop-Lox [LSL]-tdTomato, eR1-CreERT2;Rosa-LSL-EYFP mice). Gastric tissues were collected and lineage-tracing experiments were performed. Gastric organoids were cultured from eR1-CreERT2(5-2);Rosa-LSL-tdTomato mice and immunofluorescence analyses were performed. We investigated the effects of expressing oncogenic mutations in stem cells under control of eR1 using eR1-CreERT2;LSL-KrasG12D/+ mice; gastric tissues were collected and analyzed by histology and immunofluorescence. RESULTS: Most proliferation occurred in the isthmus; 86% of proliferating cells were RUNX1-positive and 76% were MUC5AC-positive. In eR1-EGFP mice, EGFP signals were detected mainly in the upper part of the gastric unit, and 83% of EGFP-positive cells were located in the isthmus/pit region. We found that eR1 marked undifferentiated stem cells in the isthmus and a smaller number of terminally differentiated chief cells at the base. eR1 also marked cells in the pyloric gland in the antrum. Lineage-tracing experiments demonstrated that stem cells in the isthmus and antrum continuously gave rise to mature cells to maintain the gastric unit. eR1-positive cells in the isthmus and pyloric gland generated organoid cultures in vitro. In eR1-CreERT2;LSL-Kras G12D/+ mice, MUC5AC-positive cells rapidly differentiated from stem cells in the isthmus, resulting in distinct metaplastic lesions similar to that observed in human gastric atrophy. CONCLUSIONS: Using lineage-tracing experiments in mice, we found that a Runx1 enhancer element, eR1, promotes its expression in the isthmus stem cells of stomach corpus as well as pyloric gland in the antrum. We were able to use eR1 to express oncogenic mutations in gastric stem cells, proving a new model for studies of gastric carcinogenesis.

Inpatient glucose management programme in the Asian healthcare setting.

AIM: To implement an inpatient glucose management (IGM) programme in the general medical wards and evaluate its clinical efficacy. METHOD: Consecutive patients admitted to selected medical wards over a 12-month period were included in the IGM programme. All patients with ≥3 capillary blood glucose (CBG) readings (>10.0 mmol/L and/or <4.0 mmol/L) over a 24-hour period were identified electronically. A multidisciplinary team of diabetes care providers would provide specialist input to these patients. The control group comprised of consecutive patients admitted over the preceding 12 months. Outcome glucose measures include mean in-hospital glucose (MHG), mean patient-day glucose (MDG), proportion of CBG readings at predefined cut-offs and length of stay (LOS). RESULTS: Both the MHG and MDG were significantly lower following intervention (10.0±2.4 mmol/L vs 11.2±2.6 mmol/L, P<.001; 10.0±2.3 mmol/L vs 11.2±2.6 mmol/L, P<.001, respectively). Prevalence of hyperglycaemic events, defined by CBG >10.0 mmol/L, was significantly lower at 36.5% versus 51.6% (P<.001). Hypoglycaemic events of CBG <4.0 mmol/L remained infrequent at <1.0% before and after IGM programme. A greater proportion of glucose readings was controlled within the target range of 4.0-10.0 mmol/L (62.6% vs 47.6%, P<.001). With the IGM programme in place, more patients received scheduled CBG monitoring, and a significant shortening of mean LOS by 3.2 days was observed (P=.02). CONCLUSIONS: The IGM programme was effective in improving inpatient glycaemic monitoring and control in the general medical wards, with a significant reduction in LOS observed. These demonstrated the programme's potential to enhance quality and efficiency of patient care.

Fiber-optic Raman spectroscopy for in vivo diagnosis of gastric dysplasia.

This study aims to assess the clinical utility of a rapid fiber-optic Raman spectroscopy technique developed for enhancing in vivo diagnosis of gastric precancer during endoscopic examination. We have developed a real-time fiber-optic Raman spectroscopy system capable of simultaneously acquiring both fingerprint (FP) (i.e., 800-1800 cm(-1)) and high-wavenumber (HW) (i.e., 2800-3600 cm(-1)) Raman spectra from gastric tissue in vivo at endoscopy. A total of 5792 high-quality in vivo FP/HW Raman spectra (normal (n = 5160); dysplasia (n = 155), and adenocarcinoma (n = 477)) were acquired in real-time from 441 tissue sites (normal (n = 396); dysplasia (n = 11), and adenocarcinoma (n = 34)) of 191 gastric patients (normal (n = 172); dysplasia (n = 6), and adenocarcinoma (n = 13)) undergoing routine endoscopic examinations. Partial least squares discriminant analysis (PLS-DA) together with leave-one-patient-out cross validation (LOPCV) were implemented to develop robust spectral diagnostic models. The FP/HW Raman spectra differ significantly between normal, dysplasia and adenocarcinoma of the stomach, which can be attributed to changes in proteins, lipids, nucleic acids, and the bound water content. PLS-DA and LOPCV show that the fiber-optic FP/HW Raman spectroscopy provides diagnostic sensitivities of 96.0%, 81.8% and 88.2%, and specificities of 86.7%, 95.3% and 95.6%, respectively, for the classification of normal, dysplastic and cancerous gastric tissue, superior to either the FP or HW Raman techniques alone. Further dichotomous PLS-DA analysis yields a sensitivity of 90.9% (10/11) and specificity of 95.9% (380/396) for the detection of gastric dysplasia using FP/HW Raman spectroscopy, substantiating its clinical advantages over white light reflectance endoscopy (sensitivity: 90.9% (10/11), and specificity: 51.0% (202/396)). This work demonstrates that the fiber-optic FP/HW Raman spectroscopy technique has great promise for enhancing in vivo diagnosis of gastric precancer during routine endoscopic examination.

Characterizing variability of in vivo Raman spectroscopic properties of different anatomical sites of normal colorectal tissue towards cancer diagnosis at colonoscopy.

This study aims to characterize the in vivo Raman spectroscopic properties of normal colorectal tissues and to assess distinctive biomolecular variations of different anatomical locations in the colorectum for cancer diagnosis. We have developed a novel 785 nm excitation fiber-optic Raman endoscope that can simultaneously acquire in vivo fingerprint (FP) spectra (800-1800 cm(-1)) and high-wavenumber (HW) Raman spectra (2800-3600 cm(-1)) from the subsurface of colorectal tissue. We applied the FP/HW Raman endoscope for in vivo tissue Raman measurements of various normal colorectal anatomical locations (i.e., ascending colon (n = 182), transverse colon (n = 249), descending colon (n = 124), sigmoid (n = 212), and rectum (n = 362)) in 50 subjects. Partial least-squares (PLS)-discriminant analysis (DA) was employed to evaluate the interanatomical variability. The normal colorectal tissue showed a subtle interanatomical variability in molecular constituents (i.e., proteins, lipids, and water content) and could be divided into three major clusterings: (1) ascending colon and transverse colon, (2) descending colon, and (3) sigmoid and rectum. The PLS-DA multiclass algorithms were able to identify different tissue sites with varying sensitivities (SE) and specificities (SP) (ascending colon: SE: 1.10%, SP: 91.02; transverse colon: SE: 14.06%, SP: 78.78; descending colon: SE: 40.32%, SP: 81.99; sigmoid: SE: 19.34%, SP: 87.90; rectum: SE: 71.55%, SP: 77.84). The interanatomical molecular variability was orders of magnitude less than neoplastic tissue transformation. Further PLS-DA modeling on in vivo FP/HW tissue Raman spectra yielded a diagnostic accuracy of 88.8% (sensitivity: 93.9% (93/99); specificity 88.3% (997/1129) for colorectal cancer detection. This work discloses that interanatomical Raman spectral variability of normal colorectal tissue is subtle compared to cancer tissue, and the simultaneous FP/HW Raman endoscopic technique has promising potential for real-time, in vivo diagnosis of colorectal cancer at the molecular level.

Comparative study of the endoscope-based bevelled and volume fiber-optic Raman probes for optical diagnosis of gastric dysplasia in vivo at endoscopy.

This study aims to compare the diagnostic performance of the two different endoscope-based fiber-optic Raman probe designs (i.e., bevelled and volume Raman probes) for real-time, in vivo detection of gastric dysplasia at endoscopy. To conduct the clinical comparison, a total of 1,050 in vivo tissue Raman spectra (normal: n = 864; dysplasia: n = 186) were acquired from 66 gastric patients (normal: n = 48; dysplasia: n = 18) by using bevelled Raman probe, while a total of 1,913 in vivo tissue Raman spectra (normal: n = 1,786; dysplasia: n = 127) were acquired from 98 gastric patients (normal: n = 87; dysplasia: n = 11) by using volume Raman probe. The bevelled Raman probe provides approximately twofold improvements in tissue Raman-to-autofluorescence intensity ratios as compared to the use of volume Raman probe. Partial least squares discriminant analysis together with leave-one patient-out cross-validation on in vivo tissue Raman spectra acquired yields a diagnostic accuracy of 93.0 % (sensitivity of 92.5 %; specificity of 93.1 %) for differentiating gastric dysplasia from normal gastric tissue by using the bevelled fiber-optic Raman probe, which is superior to the diagnostic performance (accuracy of 88.4 %; sensitivity of 85.8 %; specificity of 88.6 %) by using the volume Raman probe. This work demonstrates that the Raman spectroscopic technique coupled with bevelled fiber-optic Raman probe has great potential to enhance in vivo diagnosis of gastric precancer and early cancer at endoscopy. Graphical Abstract Comparison of in vivo gastric tissue Raman spectra acquired by using bevelled and volume fiber-optic Raman probes.

Impact of COVID-19 pandemic on neurodevelopmental outcomes of premature infants: a retrospective national cohort study.

OBJECTIVE: To compare the neurodevelopmental outcomes of preterm infants before and during the COVID-19 pandemic. DESIGN: Premature infants born in 2018 were assigned to the pre-pandemic group, while those born in 2019 were assigned to the during-pandemic group. SETTING: Nationwide cohort study. PATIENTS: Very low birthweight premature infants registered in the Taiwan Premature Infant Follow-up Network database. INTERVENTIONS: Anti-epidemic measures, including quarantine and isolation protocols, social distancing, the closure of public spaces and restrictions on travel and gatherings during COVID-19 pandemic. MAIN OUTCOME MEASURES: Outcomes were measured by Bayley Scales of Infant and Toddler Development Third Edition at corrected ages of 6, 12 and 24 months old. Generalised estimating equation (GEE) was applied to incorporate all measurements into a single model. RESULTS: Among the 1939 premature infants who were enrolled, 985 developed before the pandemic, while 954 developed during the pandemic. Premature infants whose development occurred during the pandemic exhibited better cognitive composite at the corrected age of 6 months (beta=2.358; 95% CI, 1.07 to 3.65; p<0.001), and motor composite at corrected ages of 12 months (beta=1.680; 95% CI, 0.34 to 3.02; p=0.014). GEE analysis showed that infants who had grown during the pandemic achieved higher scores in cognitive composite (beta=1.416; 95% CI, 0.36 to 2.48; p=0.009). CONCLUSION: Premature infants in Taiwan who developed during the pandemic showed better neurodevelopment compared with those born before the pandemic.