The Human Gastric Juice: A Promising Source for Gastric Cancer Biomarkers.

Gastric cancer (GC) is a major public health problem worldwide, with high mortality rates due to late diagnosis and limited treatment options. Biomarker research is essential to improve the early detection of GC. Technological advances and research methodologies have improved diagnostic tools, identifying several potential biomarkers for GC, including microRNA, DNA methylation markers, and protein-based biomarkers. Although most studies have focused on identifying biomarkers in biofluids, the low specificity of these markers has limited their use in clinical practice. This is because many cancers share similar alterations and biomarkers, so obtaining them from the site of disease origin could yield more specific results. As a result, recent research efforts have shifted towards exploring gastric juice (GJ) as an alternative source for biomarker identification. Since GJ is a waste product during a gastroscopic examination, it could provide a "liquid biopsy" enriched with disease-specific biomarkers generated directly at the damaged site. Furthermore, as it contains secretions from the stomach lining, it could reflect changes associated with the developmental stage of GC. This narrative review describes some potential biomarkers for gastric cancer screening identified in gastric juice.

POEM from A to Z: current perspectives.

The "third space endoscopy" or also called "submucosal endoscopy" is a reality we can transfer to our patients since 2010. Various modifications of the submucosal tunneling technique allow access to the submucosa or deeper layers of the gastrointestinal tract. In addition to peroral endoscopic myotomy for the treatment of achalasia, also called esophageal POEM, other variants have emerged that make it possible to treat different esophageal motility disorders, esophageal diverticula, subepithelial tumors of various locations, gastroparesis, reconnection of complete esophageal strictures or even thanks to exceptional endoscopists, pediatric disorders such as Hirschsprung's disease. Although some technical aspects are yet to be standardized, these procedures are becoming widespread worldwide and will likely become the standard treatment of these pathologies soon.

Underwater versus conventional EMR of large nonpedunculated colorectal lesions: a multicenter randomized controlled trial.

BACKGROUND AND AIMS: Underwater EMR (UEMR) is an alternative procedure to conventional EMR (CEMR) to treat large, nonpedunculated colorectal lesions (LNPCLs). In this multicenter, randomized controlled clinical trial, we aimed to compare the efficacy and safety of UEMR versus CEMR on LNPCLs. METHODS: We conducted a multicenter, randomized controlled clinical trial from February 2018 to February 2020 in 11 hospitals in Spain. A total of 298 patients (311 lesions) were randomized to the UEMR (n = 149) and CEMR (n = 162) groups. The main outcome was the lesion recurrence rate in at least 1 follow-up colonoscopy. Secondary outcomes included technical aspects, en bloc resection rate, R0 resection rates, and adverse events, among others. RESULTS: There were no differences in the overall recurrence rate (9.5% UEMR vs 11.7% CEMR; absolute risk difference, -2.2%; 95% CI, -9.4 to 4.9). However, considering polyp sizes between 20 and 30 mm, the recurrence rate was lower for UEMR (3.4% UEMR vs 13.1% CEMR; absolute risk difference, -9.7%; 95% CI, -19.4 to 0). The R0 resection showed the same tendency, with significant differences favoring UEMR only for polyps between 20 and 30 mm. Overall, UEMR was faster and easier to perform than CEMR. Importantly, the techniques were equally safe. CONCLUSIONS: UEMR is a valid alternative to CEMR for treating LNPCLs and could be considered the first option of treatment for lesions between 20 and 30 mm due to its higher en bloc and R0 resection rates. (Clinical trial registration number: NCT03567746.).

Clip closure to prevent adverse events after EMR of proximal large nonpedunculated colorectal polyps: meta-analysis of individual patient data from randomized controlled trials.

BACKGROUND AND AIMS: After EMR, prophylactic clipping is often performed to prevent clinically significant post-EMR bleeding (CSPEB) and other adverse events (AEs). Prior evidence syntheses have lacked sufficient power to assess clipping in relevant subgroups or in nonbleeding AEs. We performed a meta-analysis of individual patient data (IPD) from randomized trials assessing the efficacy of clipping to prevent AEs after EMR of proximal large nonpedunculated colorectal polyps (LNPCPs) ≥20 mm. METHODS: We searched EMBASE, MEDLINE, Cochrane Central Registry of Controlled Trials, and PubMed from inception to May 19, 2021. Two reviewers screened citations in duplicate. Corresponding authors of eligible studies were invited to contribute IPD. A random-effects 1-stage model was specified for estimating pooled effects, adjusting for patient sex and age and for lesion location and size, whereas a fixed-effects model was used for traditional meta-analyses. RESULTS: From 3145 citations, 4 trials were included, representing 1248 patients with proximal LNPCPs. The overall rate of CSPEB was 3.5% and 9.0% in clipped and unclipped patients, respectively. IPD were available for 1150 patients, in which prophylactic clipping prevented CSPEB with an odds ratio (OR) of .31 (95% confidence interval [CI], .17-.54). Clipping was not associated with perforation or abdominal pain, with ORs of .78 (95% CI, .17-3.54) and .67 (95% CI, .20-2.22), respectively. CONCLUSIONS: Prophylactic clipping is efficacious in preventing CSPEB after EMR of proximal LNPCPs. Therefore, clip closure should be considered a standard component of EMR of LNPCPs in the proximal colon.

Preventing Postendoscopic Mucosal Resection Bleeding of Large Nonpedunculated Colorectal Lesions.

The most common major adverse event of endoscopic mucosal resection (EMR) is clinically significant post-EMR bleeding (CSPEB), with an incidence of 6%-7% in large lesions. Repeat colonoscopy, blood transfusions, or other interventions are often needed. The associated direct costs are much higher than those of an uncomplicated EMR. In this review, we discuss the aspects related to CSPEB of large nonpedunculated polyps, such as risk factors, predictive models, and prophylactic measures, and we highlight evidence for preventive treatment options and explore new methods for bleeding prophylaxis. We also provide recommendations for steps that can be taken before, during, and after EMR to minimize bleeding risk. Finally, this review proposes future directions to reduce CSPEB incidence.

Outcomes of peroral endoscopic myotomy in patients with spastic esophageal motility disorders that do not fulfill Chicago Classification criteria.

INTRODUCTION: per-oral endoscopic myotomy (POEM) has become a mainstream treatment for achalasia and is a promising therapy in spastic disorders. METHODS: this is a retrospective study of prospectively collected data (case series). We present the first results of the use of POEM in patients with atypical spastic esophageal motor disorders that do not satisfy current Chicago Classification criteria. Seven consecutive patients with troublesome and persistent symptoms (12-180 months) related to atypical spastic esophageal motor dysfunction were systematically assessed before and after POEM, the extent of which was tailored by manometric findings. In five of the patients, other endoscopic or surgical procedures had failed. RESULTS: high-resolution manometry (HRM) showed a spastic esophageal body contractile segment in varying positions and lengths along the esophageal body which did not meet Chicago Classification criteria. After POEM, dysphagia and/or chest pain had either resolved or was greatly reduced. HRM 3-6 months after myotomy showed that the regions of spastic contraction targeted by myotomy had been ablated. There were no major complications. The clinical responses were fully maintained up to the most recent assessments after POEM (range 7-44 months). CONCLUSION: in our seven patients, POEM was a highly effective treatment for patients with troublesome symptoms related to atypical spastic esophageal motility disorders.

Chemically Defined Conditions Mediate an Efficient Induction of Dental Pulp Pluripotent-Like Stem Cells into Hepatocyte-Like Cells.

Liver diseases are major causes of morbidity and mortality. Dental pulp pluripotent-like stem cells (DPPSCs) are of a considerable promise in tissue engineering and regenerative medicine as a new source of tissue-specific cells; therefore, this study is aimed at demonstrating their ability to generate functional hepatocyte-like cells in vitro. Cells were differentiated on a collagen scaffold in serum-free media supplemented with growth factors and cytokines to recapitulate liver development. At day 5, the differentiated DPPSC cells expressed the endodermal markers FOXA1 and FOXA2. Then, the cells were derived into the hepatic lineage generating hepatocyte-like cells. In addition to the associated morphological changes, the cells expressed the hepatic genes HNF6 and AFP. The terminally differentiated hepatocyte-like cells expressed the liver functional proteins albumin and CYP3A4. In this study, we report an efficient serum-free protocol to differentiate DPPSCs into functional hepatocyte-like cells. Our approach promotes the use of DPPSCs as a new source of adult stem cells for prospective use in liver regenerative medicine.

Biomarkers for Gastric Cancer Screening and Early Diagnosis.

Gastric cancer is one of the most common cancers worldwide, with a bad prognosis associated with late-stage diagnosis, significantly decreasing the overall survival. This highlights the importance of early detection to improve the clinical course of these patients. Although screening programs, based on endoscopic or radiologic approaches, have been useful in countries with high incidence, they are not cost-effective in low-incidence populations as a massive screening strategy. Additionally, current biomarkers used in daily routine are not specific and sensitive enough, and most of them are obtained invasively. Thus, it is imperative to discover new noninvasive biomarkers able to diagnose early-stage gastric cancer. In this context, liquid biopsy is a promising strategy. In this review, we briefly discuss some of the potential biomarkers for gastric cancer screening and diagnosis identified in blood, saliva, urine, stool, and gastric juice.

S53P4 Bioactive Glass Inorganic Ions for Vascularized Bone Tissue Engineering by Dental Pulp Pluripotent-Like Stem Cell Cocultures.

IMPACT STATEMENT: In this study, we proposed for the first time the use of inorganic ions dissolved from BaG in a cell coculture system to induce vascularized bone formation in vitro. For that, we used dental pulp pluripotent-like stem cells from a single individual source obtained in a minimally invasive extraction manner. Moreover, we carried out all the experiments under xeno-free conditions, allowing the extrapolation of the results to the development of clinically orientated applications. Overall, these results would provide a new promising system to promote the success and survival of bone tissue engineering constructs after implantation.

The Effect of Commercially Available Endodontic Cements and Biomaterials on Osteogenic Differentiation of Dental Pulp Pluripotent-Like Stem Cells.

The aim of this study is to compare the osteogenic differentiation capacity of the dental pulp pluripotent-like stem cells (DPPSCs) using conditional media pretreated with ProRoot-MTA, Biodentine (BD) or the newly manufactured pure Portland cement Med-PZ (MZ). DPPSCs, isolated from human third molars, are the most relevant cell model to draw conclusions about the role of biomaterials on dental tissue regeneration. Cytotoxicity, alkaline phosphatase (ALP) activity, and calcium deposition analysis were evaluated at different differentiation time points. Gene expression of key osteogenic markers (RUNX2, Collagen I and Osteocalcin) was determined by qRT-PCR analysis. The osteogenic capacity of cells cultured in conditioned media prepared from MZ or MTA cements was comparable. BD conditioned media supported cell proliferation but failed to induce osteogenesis. Relative to controls and other cements, high osteogenic gene expression was observed in cultures pre-treated with the novel endodontic cement MZ. In conclusion, the in vitro behavior of a MZ- endodontic cement was evaluated, showing similar enhanced cell proliferation compared to other commercially available cements but with an enhanced osteogenic capacity with prospective potential as a novel cement for endodontic treatments.

Human dental pulp pluripotent-like stem cells promote wound healing and muscle regeneration.

BACKGROUND: Dental pulp represents an easily accessible autologous source of adult stem cells. A subset of these cells, named dental pulp pluripotent-like stem cells (DPPSC), shows high plasticity and can undergo multiple population doublings, making DPPSC an appealing tool for tissue repair or maintenance. METHODS: DPPSC were harvested from the dental pulp of third molars extracted from young patients. Growth factors released by DPPSC were analysed using antibody arrays. Cells were cultured in specific differentiation media and their endothelial, smooth and skeletal muscle differentiation potential was evaluated. The therapeutic potential of DPPSC was tested in a wound healing mouse model and in two genetic mouse models of muscular dystrophy (Scid/mdx and Sgcb-null Rag2-null γc-null). RESULTS: DPPSC secreted several growth factors involved in angiogenesis and extracellular matrix deposition and improved vascularisation in all three murine models. Moreover, DPPSC stimulated re-epithelialisation and ameliorated collagen deposition and organisation in healing wounds. In dystrophic mice, DPPSC engrafted in the skeletal muscle of both dystrophic murine models and showed integration in muscular fibres and vessels. In addition, DPPSC treatment resulted in reduced fibrosis and collagen content, larger cross-sectional area of type II fast-glycolytic fibres and infiltration of higher numbers of proangiogenic CD206+ macrophages. CONCLUSIONS: Overall, DPPSC represent a potential source of stem cells to enhance the wound healing process and slow down dystrophic muscle degeneration.

Dental pulp pluripotent-like stem cells (DPPSC), a new stem cell population with chromosomal stability and osteogenic capacity for biomaterials evaluation.

BACKGROUND: Biomaterials are widely used to regenerate or substitute bone tissue. In order to evaluate their potential use for clinical applications, these need to be tested and evaluated in vitro with cell culture models. Frequently, immortalized osteoblastic cell lines are used in these studies. However, their uncontrolled proliferation rate, phenotypic changes or aberrations in mitotic processes limits their use in long-term investigations. Recently, we described a new pluripotent-like subpopulation of dental pulp stem cells derived from the third molars (DPPSC) that shows genetic stability and shares some pluripotent characteristics with embryonic stem cells. In this study we aim to describe the use of DPPSC to test biomaterials, since we believe that the biomaterial cues will be more critical in order to enhance the differentiation of pluripotent stem cells. METHODS: The capacity of DPPSC to differentiate into osteogenic lineage was compared with human sarcoma osteogenic cell line (SAOS-2). Collagen and titanium were used to assess the cell behavior in commonly used biomaterials. The analyses were performed by flow cytometry, alkaline phosphatase and mineralization stains, RT-PCR, immunohistochemistry, scanning electron microscopy, Western blot and enzymatic activity. Moreover, the genetic stability was evaluated and compared before and after differentiation by short-comparative genomic hybridization (sCGH). RESULTS: DPPSC showed excellent differentiation into osteogenic lineages expressing bone-related markers similar to SAOS-2. When cells were cultured on biomaterials, DPPSC showed higher initial adhesion levels. Nevertheless, their osteogenic differentiation showed similar trend among both cell types. Interestingly, only DPPSC maintained a normal chromosomal dosage before and after differentiation on 2D monolayer and on biomaterials. CONCLUSIONS: Taken together, these results promote the use of DPPSC as a new pluripotent-like cell model to evaluate the biocompatibility and the differentiation capacity of biomaterials used in bone regeneration.

Improvement of osteogenesis in dental pulp pluripotent-like stem cells by oligopeptide-modified poly(ß-amino ester)s.

Controlling pluripotent stem cell differentiation via genetic manipulation is a promising technique in regenerative medicine. However, the lack of safe and efficient delivery vehicles limits this application. Recently, a new family of poly(ß-amino ester)s (pBAEs) with oligopeptide-modified termini showing high transfection efficiency of both siRNA and DNA plasmid has been developed. In this study, oligopeptide-modified pBAEs were used to simultaneously deliver anti-OCT3/4 siRNA, anti-NANOG siRNA, and RUNX2 plasmid to cells from the dental pulp with pluripotent-like characteristics (DPPSC) in order to promote their osteogenic differentiation. Results indicate that transient inhibition of the pluripotency marker OCT3/4 and the overexpression of RUNX2 at day 7 of differentiation markedly increased and accelerated the expression of osteogenic markers. Furthermore, terminally-differentiated cells exhibited higher matrix mineralization and alkaline phosphatase activity. Finally, cell viability and genetic stability assays indicate that this co-delivery system has high chromosomal stability and minimal cytotoxicity. Therefore, we conclude that such co-delivery strategy is a safe and a quick option for the improvement of DPPSC osteogenic differentiation. STATEMENT OF SIGNIFICANCE: Controlling pluripotent stem cell differentiation via genetic manipulation is a promising technique in regenerative medicine. However, the lack of safe and efficient delivery vehicles limits this application. In this study, we propose the use of a new family of oligopeptide-modified pBAEs developed in our group to control the differentiation of dental pulp pluripotential stem cells (DPPSC). In order to promote their osteogenic differentiation. The strategy proposed markedly increased and accelerated the expression of osteogenic markers, cell mineralization and alkaline phosphatase activity. Finally, cell viability and genetic stability assays indicated that this co-delivery system has high chromosomal stability and minimal cytotoxicity. These findings open a new interesting path in the usage of non-viral gene delivery systems for the control of pluripotential stem cell differentiation.

Age-dependent modifications in the mRNA levels of the rat excitatory amino acid transporters (EAATs) at 48hour reperfusion following global ischemia.

This study reports the mRNA levels of some excitatory amino acid transporters (EAATs) in response to ischemia-reperfusion (I/R) in rat hippocampus and cerebral cortex. The study was performed in 3-month-old and 18-month-old animals to analyze the possible role of age in the I/R response of these transporters. The I/R resulted in a reduced transcription of both the neuronal EAAC1 (excitatory amino acid carrier-1) and the neuronal and glial GLT-1 (glial glutamate transporter 1), while the glial GLAST1a (l-glutamate/l-aspartate transporter 1a) transcription increased following I/R. The changes observed were more striking in 3-month-old animals than in 18-month-old animals. We hypothesize that increases in the GLAST1a mRNA levels following I/R insult can be explained by increases in glial cells, while the GLT-1 response to I/R mirrors neuronal changes. GLAST1a transcription increases in 3-month-old animals support the hypothesis that this transporter would be the main mechanism for extracellular glutamate clearance after I/R. Decreases in EAAC1 and GLT-1 mRNA levels would represent either neuronal changes due to the delayed neuronal death or a putative protective down-regulation of these transporters to decrease the amount of glutamate inside the neurons, which would decrease their glutamate release. This study also reports how the treatment with the anti-inflammatory agent meloxicam attenuates the transcriptional response to I/R in 3-month-old rats and decreases the survival of the I/R-injured animals.

Age and meloxicam attenuate the ischemia/reperfusion-induced down-regulation in the NMDA receptor genes.

This study describes the effect of global brain ischemia followed by 48 h reperfusion, when delayed neuronal death can be already observed. We quantified the mRNA levels of the N-methyl-D-aspartate receptor (NMDAR) subunits and those of the astroglia (glial fibrilar acidic protein, GFAP) and microglia (CD11b) markers using real time PCR on the cerebral cortex and hippocampus of 3- and 18-month-old Sprague-Dawley rats. Data show an ischemia/reperfusion-induced decrease in the mRNA levels of the NMDAR NR1, NR2A and NR2B subunits genes, which contrasts with the increase in the CD11b and GFAP mRNA levels. These effects are attenuated in all the genes studied in 18-month-old animals, suggesting that this mechanism of response is less efficient in aged animals. Western blot assays of NR1, NR2A and NR2B show parallels with the real time PCR data, indicating that the down-regulation of these genes is controlled at the transcriptional level. We suggest that a decrease in the efficiency in the control of the NMDAR transcription could account for the higher vulnerability in aged animals, but it cannot explain by itself differences in the vulnerability to ischemia in different areas of the brain. In the assays of ischemia/reperfusion followed by a treatment with the anti-inflammatory agent meloxicam, we observed that ischemic insult was unable to elicit changes in the NMDAR transcription, thus suggesting that inflammation plays a crucial role in the transcriptional control of these genes.

Global ischemia-induced modifications in the expression of AMPA receptors and inflammation in rat brain.

Alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptors (AMPAR) and inflammatory processes have been related to ischemia-induced damage, but there are few studies addressing their response in different brain areas. Here we compare AMPAR expression after ischemia in several brain areas (hippocampus, cerebral cortex and caudate-putamen) in an attempt to correlate it with their different vulnerabilities. We found outstanding decreases in GluR1 and GluR2 mRNA levels after global ischemia and 48 h reperfusion (I/R) in all the areas studied, however, protein levels maintained in some areas such as CA3, suggesting different post-transcriptional control in different areas of the brain. To characterize the inflammatory response in these areas, we measured the mRNA levels of CD11b/CD18 membrane integrin (a reactive microglia marker), which showed an important but similar up-regulation in all brain areas studied, which was confirmed by immunohistochemistry. We conclude that the down-regulation of AMPAR gene expression following I/R does not explain differences in the vulnerability of different areas. Additionally, our data indicate that the level of inflammation is independent of the vulnerability of the different brain areas and does not explain differences in the AMPAR expression observed in the brain areas studied.

Transient global ischemia in rat brain promotes different NMDA receptor regulation depending on the brain structure studied.

The mRNA expression of the major subunits of N-methyl-d-aspartate receptors (NR1, NR2A and NR2B) following ischemia-reperfusion was studied in structures with different vulnerabilities to ischemic insult in the rat brain. The study was performed using quantitative real-time PCR on samples from 3-month-old male Sprague-Dawley rats after global transient forebrain ischemia followed by 48h of reperfusion. Expression of NMDA receptor subunits mRNAs decreased significantly in all structures studied in the injured animals as compared to the sham-operated ones. The hippocampal subfields (CA1, CA3 and dentate gyrus) as well as the caudate-putamen, both reported to be highly ischemic-vulnerable structures, showed outstandingly lower mRNA levels of NMDA receptor subunits than the cerebral cortex, which is considered a more ischemic-resistant structure. The ratios of the mRNA levels of the different subunits were analyzed as a measure of the NMDA receptor expression pattern for each structure studied. Hippocampal areas showed changes in NMDA receptor expression after the insult, with significant decreases in the NR2A with respect to the NR1 and NR2B subunits. Thus, the NR1:NR2A:NR2B (1:1:2) ratios observed in the sham-operated animals became (2:1:4) in insulted animals. This modified expression pattern was similar in CA1, CA3 and the dentate gyrus, in spite of the different vulnerabilities reported for these hippocampal areas. In contrast, no significant differences in the expression pattern were observed in the caudate-putamen or cerebral cortex on comparing the sham-operated animals with the ischemia-reperfused rats. Our results support the notion that the regulation of NMDA receptor gene expression is dependent on the brain structure rather than on the higher or lower vulnerability of the area studied.

Muscarinic receptor changes in the gerbil thalamus during aging.

Here we studied muscarinic receptors in the gerbil thalamus at 8 different ages - from 6 to 36 months - using receptor and functional autoradiography. The pharmacological profile inhibiting [(3)H]N-methyl scopolamine ([(3)H]NMS) binding with 50 and 200 nM pirenzepine, 30 nM pFHHSiD and 100 nM AF-DX 116 revealed the predominance of the M(2) muscarinic subtype in the thalamic nuclei studied, mainly in the anteroventral, anteromedial and paraventricular thalamic nuclei. These data correlated with the highest [(35)S]guanylyl-5'-O-(gamma-thio)-triphosphate ([(35)S]GTP gamma S) binding induced in these nuclei by the muscarinic agonist oxotremorine in functional autoradiographic assays. Significant aging-dependent increases in the functional response in these three nuclei were observed, but only the anteroventral and anteromedial thalamic nuclei showed aging-dependent increases in [(3)H]NMS binding. Since these nuclei exert relevant functions, in which cholinergic pathways are involved and acetylcholine release is reported to decrease during aging, we suggest that the anteroventral and anteromedial thalamic nuclei would play critical roles in the cholinergic transmission that require compensatory mechanisms during the aging process and that are not observed in other thalamic nuclei.