Rectal Epithelial Cells Research Articles

PTGER4 signaling regulates class IIa HDAC function and SPINK4 mRNA levels in rectal epithelial cells

BackgroundThe prostaglandin receptor PTGER4 facilitates homeostasis in the gut. Previous reports indicate that goblet cells, marked by SPINK4 expression, might be affected by PTGER4 activity. Current evidence suggests that prostaglandin E2 (PGE2) produced by mesenchymal stromal cells (MSC) stimulates PTGER4 in epithelial cells during inflammatory conditions. Here, we investigate the subcellular mechanisms and mRNA levels downstream of PTGER4 activity in epithelial cells.MethodsMucosal cells, organoids, and MSC were obtained from patient biopsies harvested by endoscopy. Using independent and co-cultures, we manipulated the activity of PTGER4, the downstream enzymes, and mRNA levels, by using PGE2, in combination with chemical inhibitors, L-161982, H89, LB100, DAPT, LMK-235, or with butyrate. Immunofluorescence, single cell sequencing, RNAscope, ELISA, real time PCR, and Western blotting were used to examine these samples.ResultsSPINK4 mRNA levels were increased in organoids by co-culture with MSC or exogenous stimulation with PGE2 that could be blocked by L-161982 or LMK-235, PTGER4 or HDAC4 inhibitors, respectively. Expression of PTGER4 was co-localized with JAM-A in the basolateral surfaces in rectal epithelial cells grown as organoids. PGE2 treatment of rectal organoids decreased HDAC4, 5, and 7 phosphorylation levels that could be blocked by L-161982 treatment. Butyrate treatment, or addition of L-161982, increased the phosphorylated levels of HDAC4, 5, and 7.ConclusionsThese findings suggest a mechanism during mucosal injury whereby MSC production of PGE2 increases HDAC4, 5, and 7 activities in epithelial cells by upregulating PTGER4 signaling, ultimately increasing SPINK4 mRNA levels and extracellular release of SPINK4.

DOP59 Integration of in silico analysis and in vitro functional analysis reveals Tissue Factor Pathway Inhibitor 2 (TFPI2) as a master transcriptional regulator and potential target to impede Infliximab resistance in adult UC patients

Abstract Background Standard biological therapy for moderate-severe cases of UC consists in the use of Infliximab (IFX), an anti-tumour necrosis factor α (anti-TNF-α) targeting agent. However, approximately 40% of the patients do not respond to IFX. While transcriptional alterations have been widely associated with non-response to IFX, the precise factors that regulates this altered gene expression remains poorly understood. The aim of this study was perform a robust in silico analysis to identify master transcriptional regulators (MTRs) which regulate gene expression changes in IFX non-responsive UC patients, followed by perturbation in vitro to establish their functional role in UC pathogenesis. Methods Differentially expressed genes (DEGs) identified from four independent public datasets were applied to the GeneXplain platform to identify transcription factors (TFs) enriched for the DEG’s. This was followed by an upstream analysis to identify MTR’s which regulate expression of these genes through the TFs. We next explored the role of these MTRs in driving UC-associated phenotype, using siRNA-mediated knock-down in a novel IBD-like in vitro system comprising of normal rectal epithelial cells treated with TNFa. Results Over 200 TFs were identified in both upregulated and downregulated genes in IFX non-responders across all data sets. RELA, NANOG, FOSJUN were the top 3 TF’ enriched for upregulated genes in three out of the four datasets. Using the TF data, the upstream analysis identified CXCL8, SELE, PTGS2, FCGR3, TFPI2 as the top five MTRs. Next, we showed that of the 5 MTRs, only TFPI2 (tissue factor pathway inhibitor 2) were significantly upregulated in an IBD-like in vitro model (normal rectal epithelial cell line: CRL1831 +TNFa). siRNA-mediated knock-down of TFPI2 in this model resulted in a significant decrease in genes including TNFAIP6, TNFAIP3, FCGR3B, BIRC3 and CXCL5, all of which were predicted in silico to be upregulated by TFPI2 during the upstream analysis. Intriguingly, we showed that downregulation of TFPI2 resulted in significant amelioration of pro-inflammatory cytokines including IL1β, IFNγ, IL10, IL13, IL6 and IL8. Conclusion Our study for the first time identifies key MTRs such TFPI2 which acts as a master regulator of genes whose upregulation drives poor response to IFX. Therefore, these results warrant further in vitro/ex vivo investigation to rationalise their role of TFPI2 as a potential therapeutic target, whose perturbation in UC patient would likely result in improved response to IFX.

In silico analysis and theratyping of an ultra-rare CFTR genotype (W57G/A234D) in primary human rectal and nasal epithelial cells

Mutation targeted therapy in cystic fibrosis (CF) is still not eligible for all CF subjects, especially for cases carrying rare variants such as the CFTR genotype W57G/A234D (c.169T>G/c.701C>A). We performed in silico analysis of the effects of these variants on protein stability, which we functionally characterized using colonoids and reprogrammed nasal epithelial cells. The effect of mutations on cystic fibrosis transmembrane conductance regulator (CFTR) protein was analyzed by western blotting, forskolin-induced swelling (FIS), and Ussing chamber analysis. We detected a residual CFTR function that increases following treatment with the CFTR modulators VX661±VX445±VX770, correlates among models, and is associated with increased CFTR protein levels following treatment with CFTR correctors. Invivo treatment with VX770 reduced sweat chloride concentration to non-CF levels, increased the number of CFTR-dependent sweat droplets, and induced a 6% absolute increase in predicted FEV1% after 27weeks of treatment indicating the relevance of theratyping with patient-derived cells in CF.

P027 Effects of ivacaftor therapy confirm the results of theratyping using rectal and nasal epithelial cells of a cystic fibrosis patient carrying the ultra-rare CFTR genotype W57G (c.169T > G)/A234D (c.701C > A)

P027 Effects of ivacaftor therapy confirm the results of theratyping using rectal and nasal epithelial cells of a cystic fibrosis patient carrying the ultra-rare CFTR genotype W57G (c.169T > G)/A234D (c.701C > A)

Hypofractionated Radiation Changes The Gut Flora Into A Inflammatory Activation Pattern Which Is Related With Tumor Complete Regression In Local Advanced Rectum Cancer

Radiation therapy (RT) with or without chemotherapy is the standard regimen for local advanced rectum cancer (LARC). Some patients may achieve complete remission (CR) after (C)RT, while some others may not. The reason remains unknown. It is known that gastrointestinal microbiota plays an important role in the regulation of immune environment in the gut. However, the correlation between the treatment effect of RT, especially hypofractionated RT which has shown immunological simulating potential, and gut flora remains unclear. Here, we prospectively recruited patients with LARC and their fecal samples were collected before and after radiotherapy. A correlation analysis was done between clinical outcomes and gut flora characteristic. And cellular level experiment was performed for mechanism revealing. Twelve patients were recruited from the experimental arm of a stage III clinical trial. In brief, all patients will receive a 25Gy (5×5) regimen RT followed by 4 cycles XELOX chemotherapy. Fecal samples were collected according to the protocol of Human Microbiome Project the day before RT and 2 weeks after RT. Patients who achieved complete remission (CR) (clinical or pathological) were tagged as good response group (GRG) and those with tumor residual were tagged as poor response group (PRG). Flora samples were analyzed using several standard approaches. Rectal epithelial cells were co-cultured with a mixture of filtered fecal samples (flora samples) from GRG before irradiation (F1), GRG after irradiation (F2), PRG before irradiation (F3) and PRG after irradiation (F4). The releasement of several cytokines was analyzed with or without irradiation. Four patients finally achieved CR (3 CCR and 1 PCR) after evaluation. Eight patients underwent surgery and pathological exam found residual tumor cells. We first compared all patients’ samples before irradiation with those after irradiation. The abundance of several species was significantly different (species list 1). We then assigned all the data into GRG and PRG. Some species were significantly different between two groups, either before (species list 2) or after (species list 3) irradiation. The lacaceae family, especially the genus Coccus, appears in both List 1 and 3. Their abundance increased after irradiation regardless the clinical response, and they were significantly abundant in GRG after irradiation. When the rectum epithelial cells co-cultured with F2, the secretion of TNFα and IL-1β was higher than that of other flora samples (F2>F4>F1≈F3), and was more significant after the cells were irradiated. Meanwhile the rest cytokines (TGFβ、IL-6、IFN-γ) we tested did not show significant changes. Hypofractionated radiation could change the gut flora into an inflammatory activation pattern which may help with the recruitment of immune cells. Patients with good clinical outcome may benefit more from more pronounced changes in the abundance of a particular flora.

COVID-19 and Gastrointestinal Tract Symptoms: Recognition, Containment, and Learning From the Past

COVID-19 and Gastrointestinal Tract Symptoms: Recognition, Containment, and Learning From the Past

LncRNA XIST modulates 5-hydroxytrytophan-induced visceral hypersensitivity by epigenetic silencing of the SERT gene in mice with diarrhea-predominant IBS

Diarrhea-predominant irritable bowel syndrome (IBS-D) is a prevalent gastrointestinal disorder with a high incidence in children. The role of long non-coding RNAs (lncRNAs) in gastrointestinal diseases has been previously highlighted. Nevertheless, the underlying regulatory mechanism of lncRNA X inactivate-specific transcript (XIST) in IBS-D requires further studies. Thus, the present study was conducted with the main objective of elucidating the underlying mechanism of lncRNA XIST in visceral hypersensitivity in IBS-D. An in vivo mouse model of IBS-D was constructed via rectal perfusion of acetic acid. Next, in order to evaluate the effect of lncRNA XIST on the development of visceral hypersensitivity in IBS-D, different vector plasmids were injected into mice along with rectal mucosal epithelial cells, followed by the measurement of abdominal withdrawal reflex (AWR) score, counts of peristaltic wave, abdominal wall contraction and defecation particles. Furthermore, luciferase reporter assay, FISH, RIP and ChIP assays were conducted to determine the interactions between lncRNA XIST and SERT. Subsequently, MS-PCR was adopted to test the methylation level of SERT promoter. 5-hydroxytrytophan (HT) content in rectal tissues was detected using immunohistochemistry. The IBS-D mouse models presented with a high expression of lncRNA XIST along with low expression of SERT. LncRNA XIST was observed to recruit methylase DNMT1, DNMT3A and DNMT3B to promote SERT promoter methylation, reducing its expression. Restoration of lncRNA XIST resulted in increased AWR score, counts of peristaltic wave, abdominal wall contraction and defecation particles along with stimulated 5-HT expression and SERT methylation level, while downregulation of lncRNA XIST reversed these effects. In conclusion, the key findings from our study indicated that lncRNA XIST acts as a regulator in 5-HT-induced visceral hypersensitivity in mice with IBS-D, providing a new insight into the regulatory effect of lncRNA XIST and its epigenetic diagnostic and therapeutic properties in IBS-D.

Fluoride-induced rectal barrier damage and microflora disorder in mice.

Intestinal microflora plays a key role in maintaining the homeostasis between immune and host health. Here, we reported the fluoride-induced changes of rectal structure and microflora in mice. The morphology of rectal tissue was observed by hematoxylin and eosin staining. The rectal development parameters (the thickness of mucosa, intestinal gland and muscle layer) were evaluated. The proliferation of rectal epithelial cells was evaluated via BrdU labeling. The distribution of goblet, glycoprotein and mast cell were evaluated by specific staining. Rectal microflora was detected using 16S rRNA high-throughput sequencing. The results showed that the rectal structure was seriously damaged and the proliferation of rectal epithelial cells was significantly inhibited by fluoride. The distribution of goblet cells, glycoprotein and mast cells decreased significantly after fluoride exposure. The relative richness of microfloras was changed after fluoride treatment, such as increased Bacteroidetes and decreased Firmicutes. In summary, this study indicated that excessive fluoride damages the intestinal structure, disturbs the intestinal micro-ecology and causes intestinal microflora disorder in mice. Findings mentioned in the present study enrich a new scope for elucidating fluoride toxicity from intestinal homeostasis.

Lactoferrin translocates to the nucleus of bovine rectal epithelial cells in the presence of Escherichia coli O157:H7

Enterohemorrhagic Escherichia coli (EHEC) O157:H7 is a foodborne pathogen which causes illness in humans. Ruminants are the main reservoirs and EHEC predominantly colonizes the epithelium of the recto-anal junction of cattle. Immunosuppression by EHEC promotes re-infection of cattle. However, bovine lactoferrin (bLF) apparently can overrule the immunosuppression by inducing EHEC-specific IgA responses at the mucosal site. The IgA responses are significantly correlated with reduced EHEC shedding and the absence of colonization at the rectal mucosa following re-infection. Therefore, to examine the interaction between bLF and bovine rectal epithelial cells, we first developed a method to establish a primary cell culture of epithelial cells of the rectum of cattle. Furthermore, we used LC–MS/MS to demonstrate the presence of secreted lactoferrin in bovine milk and the absence of a “delta” isoform which is known to translocate to the nucleus of cells. Nevertheless, lactoferrin derived from bovine milk was internalized by rectal epithelial cells and translocated to the nuclei. Moreover, nuclear translocation of bLF was significantly enhanced when the epithelial cells were inoculated with EHEC, as demonstrated by confocal fluorescence microscopy and confirmed by Raman microscopy and 3D imaging.

MicroRNA‑195 suppresses rectal cancer growth and metastasis via regulation of the PI3K/AKT signaling pathway.

MicroRNAs (miRNAs) play a vital role in the progression of cancer, however, only limited data on miRNAs in rectal cancer are available. The aim of the present study was to investigate whether miR-195 could inhibit the progression of rectal cancer. The miR-195 mimic was transfected into 2 types of human rectal cancer cells (SW837 and SW1463). Cell viability and apoptosis were analyzed by Cell Counting Kit-8 (CCK-8) assay and flow cytometry, and cell migration and invasion were assessed by scratch test and Transwell assay. The results revealed that insulin-like growth factor 1 (IGF1) was predicted as a potential target of miR-195 by Targetscan7.2, and the result was verified by dual-luciferase reporter assay. The co-transfection of IGF1 was performed to confirm the underlying mechanism of tumor suppressor of miR-195 in rectal cancer. The activation of PI3K/AKT signaling was determined by western blotting. The levels of miR-195 in SW837 and SW1463 cells were revealed to be lower than in human rectal mucosa epithelial cells. After the transfection with miR-195, the cell viability was decreased, while the apoptosis was significantly increased (SW837: 5.21% vs. 20.96%; SW1463: 4.19% vs. 25.22%). Moreover, cell migration and invasion were significantly inhibited in the mimic group. miR-195 specifically targeted IGF1, however, the co-transfection of IGF1 could partially reverse the inhibitory effects of miR-195 on rectal cancer cells. It was also determined that the phosphorylation of PI3K and AKT were significantly inhibited in the mimic group. The tumor suppressive ability of miR-195 in rectal cancer cell proliferation and metastasis was mediated by blocking IGF1 expression and inhibiting the PI3K/AKT pathway.

Identification of a Conserved, Orphan G Protein-Coupled Receptor Required for Efficient Pathogen Clearance in Caenorhabditis elegans.

G protein-coupled receptors contribute to host defense across the animal kingdom, transducing many signals involved in both vertebrate and invertebrate immune responses. While it has become well established that the nematode worm Caenorhabditis elegans triggers innate immune responses following infection with numerous bacterial, fungal, and viral pathogens, the mechanisms by which C. elegans recognizes these pathogens have remained somewhat more elusive. C. elegans G protein-coupled receptors have been implicated in recognizing pathogen-associated damage and activating downstream host immune responses. Here we identify and characterize a novel G protein-coupled receptor required to regulate the C. elegans response to infection with Microbacterium nematophilum We show that this receptor, which we designate pathogen clearance-defective receptor 1 (PCDR-1), is required for efficient pathogen clearance following infection. PCDR-1 acts upstream of multiple G proteins, including the C. elegans Gαq ortholog, EGL-30, in rectal epithelial cells to promote pathogen clearance via a novel mechanism.

Assessment of the Biocompatibility and Biological Effects of Biodegradable Pure Zinc Material in the Colorectum.

Little attention has been paid to the biocompatibility and biological effects of zinc as a material. Here, we therefore investigated the biocompatibility and anti-inflammatory and collagen-promoting effects of pure zinc material in the colorectum. Our in vitro results indicated that zinc toxicity and concentration were closely related. Low concentrations of zinc ions and pure zinc material extract had only minor effects on the viability of primary rectal mucosal epithelial cells; however, cytotoxicity was observed at concentrations greater than 0.017 μg/μL and 60%, respectively. In vivo experiments demonstrated that zinc pins degraded slowly in the colorectum (their volume decreasing by approximately 7.79% over 1 month) and did not cause serious adverse reactions. Pure zinc material was found to inhibit acute inflammation through increased expression of ENA-78 and F4/80. Moreover, zinc material heightened expression of collagen and VEGF, factors conducive to wound healing, in surrounding colorectal tissues. These preliminary results suggest that zinc shows great promise as an implant material for medical applications involving colorectal surgery.

Morphological and ultrastructural characterization of the alimentary canal in larvae of Streltzoviella insularis (Staudinger) (Lepidoptera: Cossidae)

AbstractStreltzoviella insularis (Staudinger) is an important tree‐boring pest, that primarily damages Sophora japonica (Linnaeus) and Ginkgo biloba (Linnaeus), as well as other common species, at great economic cost to the urban landscape construction industry in China. In the present study, the alimentary canal morphology of S. insularis was observed using light microscopy, and its ultrastructure was investigated by scanning and transmission electron microscopy. The foregut of S. insularis can be divided into the pharynx, esophagus, crop, proventriculus, and cardiac valve. The well‐developed crop forms the longest section of the foregut. It is able to store large amounts of food and is lined with a monolayer of epithelial cells. Many sclerotized microspines occur on the surface of the anterior intima and there are dense spines on the posterior intima of the proventriculus. Epithelial cells of the midgut include columnar cells, goblet cells, and regenerative cells, but endocrine cells are absent. The hindgut consists of the pyloric valve, ileum, and rectum. There is no clear distinction between the ileum and colon. The intima surface of the pyloric valve carries many microspines, whereas the intestinal wall of the rectum is thin with well‐developed rectal pads. The rectal epithelial cells form a squamous monolayer. A cryptonephric excretory system is located in the hindgut. There are six spiral Malpighian tubules, in which a cellular layer on a basement membrane encloses a lumen. These results will provide the basis for further studies of the structure and function in S. insularis larvae.

Invasion of HEp-2 cells by Shigella spp. isolated from acute pediatric diarrhea.

Aim: Shigella infection is an important global health problem in developing countries where hygiene is poor and hence shigellosis is a main cause of diarrhoea-associated mortality and morbidity, particularly in children under the age of five. The bacterial entry into colon and rectal epithelial cells has been named ‘bacterium-directed phagocytosis’. This term highlights that the bacteria actively stimulate their own uptake into non-professional phagocytes. The aim of this study was to demonstrate the invasion of HEp-2 cells by Shigella spp. isolated from acute pediatric diarrhea in Tehran, Iran.Methods: Three-hundred and ten non-duplicative diarrheal stool samples were collected from the children admitted to Children’s Medical Center in Tehran, Iran. Samples were cultured and suspected colonies were identified by routine microbiological and biochemical tests. The invasion of the two isolated Shigella spp. to HEp-2 cells was studied.Results: Of 310 stool samples, 16 (5.2%) Shigella spp. were isolated, including seven (43.7%) S. sonnei and nine (56.3%) S. flexneri. Four (44.4%) S. sonnei and seven (42.8%) S. flexneri showed invasive phenotype to HEp-2.Conclusion: Shigella sonnei and S. flexneri are reported as the most prevalent Shigella spp. in nature which infect humans. Invasion of various cell lines gives the chance of survival to Shigella spp. This ability causes more virulent infections in the host. Despite costly and time consuming cell culture techniques, the current method described in this paper is reliable for detecting invasive behavior of Shigella spp. Results have also shown that not all the Shigella spp. are able to invade intestinal epithelial cells.

Comparative genomics of two super-shedder isolates of Escherichia coli O157:H7.

Shiga toxin-producing Escherichia coli O157:H7 (O157) are zoonotic foodborne pathogens and of major public health concern that cause considerable intestinal and extra-intestinal illnesses in humans. O157 colonize the recto-anal junction (RAJ) of asymptomatic cattle who shed the bacterium into the environment through fecal matter. A small subset of cattle, termed super-shedders (SS), excrete O157 at a rate (≥ 104 CFU/g of feces) that is several orders of magnitude greater than other colonized cattle and play a major role in the prevalence and transmission of O157. To better understand microbial factors contributing to super-shedding we have recently sequenced two SS isolates, SS17 (GenBank accession no. CP008805) and SS52 (GenBank accession no. CP010304) and shown that SS isolates display a distinctive strongly adherent phenotype on bovine rectal squamous epithelial cells. Here we present a detailed comparative genomics analysis of SS17 and SS52 with other previously characterized O157 strains (EC4115, EDL933, Sakai, TW14359). The results highlight specific polymorphisms and genomic features shared amongst SS strains, and reveal several SNPs that are shared amongst SS isolates, including in genes involved in motility, adherence, and metabolism. Finally, our analyses reveal distinctive patterns of distribution of phage-associated genes amongst the two SS and other isolates. Together, the results of our comparative genomics studies suggest that while SS17 and SS52 share genomic features with other lineage I/II isolates, they likely have distinct recent evolutionary histories. Future comparative and functional genomic studies are needed to decipher the precise molecular basis for super shedding in O157.

High-Purity Magnesium Staples Suppress Inflammatory Response in Rectal Anastomoses.

Magnesium-based materials are promising biodegradable implants, although the impact of magnesium on rectal anastomotic inflammation is poorly understood. Thus, we investigated the inflammatory effects of high-purity Mg staples in rectal anastomoses by in vivo luciferase reporter gene expression in transgenic mice, hematoxylin-eosin staining, immunohistochemistry, and Western blotting. As expected, strong IL-1β-mediated inflammation and inflammatory cell infiltration were observed 1 day after rectal anastomoses were stapled with high-purity Mg or Ti. However, inflammation and inflammatory cell infiltration decreased more robustly 4-7 days postoperation in tissues stapled with high-purity Mg. This rapid reduction in inflammation was confirmed by immunohistochemical analysis of IL-6 and TNF-α. Western blot also suggested that the reduced inflammatory response is due to suppressed TLR4/NF-κB signaling. In contrast, MCP-1, uPAR, and VEGF were abundantly expressed, in line with the notion that expression of these proteins is regulated by feedback between the VEGF and NF-κB pathways. In vitro expression of MCP-1, uPAR, and VEGF was also similarly high in primary rectal mucosal epithelial cells exposed to extracts from Mg staples, as measured by antibody array. Collectively, the results suggest that high-purity Mg staples suppress the inflammatory response during rectal anastomoses via TLR4/NF-κB and VEGF signaling.

Self-renewing Monolayer of Primary Colonic or Rectal Epithelial Cells.

Background & AimsThree-dimensional organoid culture has fundamentally changed the in vitro study of intestinal biology enabling novel assays; however, its use is limited because of an inaccessible luminal compartment and challenges to data gathering in a three-dimensional hydrogel matrix. Long-lived, self-renewing 2-dimensional (2-D) tissue cultured from primary colon cells has not been accomplished.MethodsThe surface matrix and chemical factors that sustain 2-D mouse colonic and human rectal epithelial cell monolayers with cell repertoires comparable to that in vivo were identified.ResultsThe monolayers formed organoids or colonoids when placed in standard Matrigel culture. As with the colonoids, the monolayers exhibited compartmentalization of proliferative and differentiated cells, with proliferative cells located near the peripheral edges of growing monolayers and differentiated cells predominated in the central regions. Screening of 77 dietary compounds and metabolites revealed altered proliferation or differentiation of the murine colonic epithelium. When exposed to a subset of the compound library, murine organoids exhibited similar responses to that of the monolayer but with differences that were likely attributable to the inaccessible organoid lumen. The response of the human primary epithelium to a compound subset was distinct from that of both the murine primary epithelium and human tumor cells.ConclusionsThis study demonstrates that a self-renewing 2-D murine and human monolayer derived from primary cells can serve as a physiologically relevant assay system for study of stem cell renewal and differentiation and for compound screening. The platform holds transformative potential for personalized and precision medicine and can be applied to emerging areas of disease modeling and microbiome studies.

Invivo and invitro assessment of the biocompatibility and degradation of high-purity Mg anastomotic staples.

Titanium (Ti) staples are not biodegradable, and anastomotic complications related to Ti staples are reported frequently. In the present study, the biocompatibility and degradation behavior of high-purity magnesium (HP Mg) staples with the small intestine were investigated. HP Mg staples did not affect the relative growth rate, cell cycle and apoptosis of primary rectal mucosal epithelial cells (IEC-6) invitro. At one, two and three days after immersion in intestinal juice, the weight of the 30 rinsed HP Mg staples reduced by 7.5 ± 1.6, 10.6 ± 2.2 and 13.5 ± 2.1 mg, respectively, and those in the Hanks' solution reduced by 3.9 ± 0.8, 6.1 ± 1.2 and 7.1 ± 2.4 mg. Extracts of HP Mg staples were bio-safe for IEC-6, and the corrosion rate of HP staples was faster in the small intestinal juice than in the Hanks' solution. In the invivo experiments, the small intestine of the minipigs was anastomosed by HP Mg and Ti staples. HP Mg staples neither affected important bio-chemical parameters nor induced serious inflammation or necrosis in the anastomosis tissues. The residual weight of a HP Mg staples (0.81 ± 0.13 mg) was 89.7% of the original weight (9 ± 0.09 mg) one month after surgery. The invivo corrosion rate for one HP Mg staple was determined to be∼0.007 ± 0.001 mm·month-1. The preliminary results of the biocompatibility and degradation of high-purity Mg anastomotic staples are promising, and further studies will be initiated to study in more detail.

P08.07 Chlamydia trachomatiscervical infection/rectal detection in the macaque model

<h3>Introduction</h3> Experimentally induced cervical chlamydial infection in the macaque may naturally cross-infect rectal epithelial cells which are also prone to <i>Chlamydia trachomatis</i> (CT) infection. This would be a significant finding in the field of STI preventive strategies, particularly when products intended for vaginal use are assessed for efficacy. If cross-infection does occur, it will be important to specifically assess rectal secretions for evidence of infection in vaginal product efficacy studies. <h3>Methods</h3> Twelve pigtailed macaques underwent direct cervical inoculation (1 mL <i>C. trachomatis</i> serovar E; 5E6IFU), followed by five weekly exams to detect infection in cervical and rectal secretions. Inoculant was delivered to the face of the cervix/vaginal fornix via 1 mL tuberculin syringe. Secretions were collected on dacron swabs. Chlamydial infection was detected at each site by culture and by nucleic acid amplification (NAAT: Aptima2) assays. <h3>Results</h3> Ten of twelve macaques tested positive for cervical chlamydial infection by culture and NAAT assays. The other two were transiently positive (2 weeks, 1 week) by NAAT only. All but three animals had chlamydial rRNA amplified from rectal swabs on at least one occasion. Five animals remained NAAT positive in rectal secretions for three weeks or more. One of these macaques had replicating chlamydia cultured from a rectal swab (week 3 only), followed by a spike in culture positivity from her next cervical sample (week4). <h3>Conclusion</h3> Experimental CT infection of the cervix indeed gave rise to chlamydia detection in rectal secretions in the majority of animals in this study. Culture positivity in cervical samples did not predict chlamydia detection in rectal samples. The paucity of culture positive results from rectal samples may be related to faecal contamination. Clearly it is advisable that rectal secretions be assessed for evidence of chlamydia in studies designed to assess prevention/treatment of cervical CT infections. <h3>Disclosure of interest statement</h3> This research was supported by NIH Contract Number HHSN272201000006I, Task Number HHSN27200008 and by the Office of Research Infrastructure Programs (ORIP) of the National Institutes of Health through Grant Number P51 OD010425 Washington National Primate Research Centre. No pharmaceutical grants were received in the development of this study.

Cancer-initiating cells derived from human rectal adenocarcinoma tissues carry mesenchymal phenotypes and resist drug therapies

Accumulating evidence indicates that cancer-initiating cells (CICs) are responsible for cancer initiation, relapse, and metastasis. Colorectal carcinoma (CRC) is typically classified into proximal colon, distal colon, and rectal cancer. The gradual changes in CRC molecular features within the bowel may have considerable implications in colon and rectal CICs. Unfortunately, limited information is available on CICs derived from rectal cancer, although colon CICs have been described. Here we identified rectal CICs (R-CICs) that possess differentiation potential in tumors derived from patients with rectal adenocarcinoma. The R-CICs carried both CD44 and CD54 surface markers, while R-CICs and their immediate progenies carried potential epithelial–mesenchymal transition characteristics. These R-CICs generated tumors similar to their tumor of origin when injected into immunodeficient mice, differentiated into rectal epithelial cells in vitro, and were capable of self-renewal both in vitro and in vivo. More importantly, subpopulations of R-CICs resisted both 5-fluorouracil/calcium folinate/oxaliplatin (FolFox) and cetuximab treatment, which are the most common therapeutic regimens used for patients with advanced or metastatic rectal cancer. Thus, the identification, expansion, and properties of R-CICs provide an ideal cellular model to further investigate tumor progression and determine therapeutic resistance in these patients.