Epithelial Disintegration Research Articles

EVALUATION OF THE EFFECTIVENESSE OF ORTOGNATHIC SURGERY IN THE TREATMENT OF FACIAL ASYMMETRY

The beginning of facial development occurs between the fourth and eighth week of gestation, during this period the cells of the neural crest migrate, forming the four pharyngeal arches that will provide the emergence of most of the structures of the head and neck. During this process, failures may occur such as: epithelial disintegration, absence of fusion, genetic mutations, prognatism or retrognatism, among other factors that cause the appearance of asymmetries in the bones of the face and soft tissues. Aiming at this perspective, the buccal-maxillofacial surgical procedures such as: the treatment of cleft lip and palate, the possibility of bone or alloplastic grafting, in regions of deficit or that have suffered some type of facial trauma, and orthognathic surgery, a theme that will be addressed in this article, have emerged as a new possibility of definitive and resolutive treatment, generating aesthetic harmony and quality of life to the treated patients.

Effects of Salinity Stress on Histological Changes, Glucose Metabolism Index and Transcriptomic Profile in Freshwater Shrimp, Macrobrachium nipponense.

Salinity is an important factor in the aquatic environment and affects the ion homeostasis and physiological activities of crustaceans. Macrobrachium nipponense is a shrimp that mainly lives in fresh and low-salt waters and plays a huge economic role in China's shrimp market. Currently, there are only a few studies on the effects of salinity on M. nipponense. Therefore, it is of particular importance to study the molecular responses of M. nipponense to salinity fluctuations. In this study, M. nipponense was set at salinities of 0, 8, 14 and 22‱ for 6 weeks. The gills from the control (0‱) and isotonic groups (14‱) were used for RNA extraction and transcriptome analysis. In total, 593 differentially expressed genes (DEGs) were identified, of which 282 were up-regulated and 311 were down-regulated. The most abundant gill transcripts responding to different salinity levels based on GO classification were organelle membrane (cellular component), creatine transmembrane transporter activity (molecular function) and creatine transmembrane transport (biological function). KEGG analysis showed that the most enriched and significantly affected pathways included AMPK signaling, lysosome and cytochrome P450. In addition, 15 DEGs were selected for qRT-PCR verification, which were mainly related to ion homeostasis, glucose metabolism and lipid metabolism. The results showed that the expression patterns of these genes were similar to the high-throughput data. Compared with the control group, high salinity caused obvious injury to gill tissue, mainly manifested as contraction and relaxation of gill filament, cavity vacuolation and severe epithelial disintegration. Glucose-metabolism-related enzyme activities (e.g., pyruvate kinase, hexokinase, 6-phosphate fructose kinase) and related-gene expression (e.g., hexokinase, pyruvate kinase, 6-phosphate fructose kinase) in the gills were significantly higher at a salinity of 14‱. This study showed that salinity stress activated ion transport channels and promoted an up-regulated level of glucose metabolism. High salinity levels caused damage to the gill tissue of M. nipponense. Overall, these results improved our understanding of the salt tolerance mechanism of M. nipponense.

Restoration of the gut barrier integrity and restructuring of the gut microbiome in aging by angiotensin-(1-7).

Compromised barrier function of colon epithelium with aging is largely due to gut microbial dysbiosis. Recent studies implicate an important role for angiotensin converting enzymes, ACE and ACE2, angiotensins, and the receptors, AT1 receptor (AT1R) and Mas receptor (MasR), in the regulation of colon functions. The present study tested the hypothesis that leaky gut in aging is associated with an imbalance in ACE2/ACE and that the treatment with angiotenisn-(1-7) (Ang-(1-7)) will restore gut barrier integrity and microbiome. Studies were carried out in Young (3-4 months) and old (20-24 months) male mice. Ang-(1-7) was administered by using osmotic pumps. Outcome measures included expressions of ACE, ACE2, AT1R, and MasR, intestinal permeability by using FITC-dextran, and immunohistochemistry of claudin 1 and occludin, and intestinal stem cells (ISCs). ACE2 protein and activity were decreased in Old group while that of ACE were unchanged. Increased intestinal permeability and plasma levels of zonulin-1 in the Old group were normalized by Ang-(1-7). Epithelial disintegrity, reduced number of goblet cells and ISCs in the old group were restored by Ang-(1-7). Expression of claudin 1 and occludin in the aging colon was increased by Ang-(1-7). Infiltration of CD11b+ or F4/80+ inflammatory cells in the old colons were decreased by Ang-(1-7). Gut microbial dysbiosis in aging was evident by decreased richness and altered beta diversity that were reversed by Ang-(1-7) with increased abundance of Lactobacillus or Lachnospiraceae. The present study shows that Ang-(1-7) restores gut barrier integrity and reduces inflammation in the aging colon by restoring the layer of ISCs and by restructuring the gut microbiome.

The perspectives of a novel operative procedure for free gingival graft

Graft shrinkage, epithelial disintegration, and even necrosis are the well-known clinical complications during initial healing stage of free gingival graft (FGG) procedure. This article presented a novel operative prodecure for FGG in a dental implant with insufficient keratinized tissue over a 3-year follow-up period. Briefly, donor site for FGG harvested from maxillary tuberosity would minimize the volume of graft shrinkage. The new periosteum suture technique could secure FGG firmly adapted on the recipient site. With 1 mm gap between FGG and muco-gingival junction may improve plasmatic circulation and revascularization. The clinical findings of case report demonstrated that this novel operative procedure may provide a viable therapeutic alternative for FGG.

Post-Cardiac Arrest Syndrome Is Not Associated With an Early Bacterial Translocation.

The aim of this study was to investigate bacterial translocation and its possible role in the development of post-resuscitation inflammatory response following Cardio-Pulmonary Resuscitation (CPR) after cardiac arrest. Munich female swine were employed for a model of cardiac arrest via application of electrical current. After 7 min, CPR was initiated, and animals were either successfully return to spontaneous circulation (ROSC) within 40 min or not (no-ROSC). At the end of experimental period and prior to sacrifice, samples from the intestine, mesenteric lymph nodes (MLN), liver and portal vein blood were obtained. Evaluation of inflammation and gut permeability was performed; MLN, liver and portal vein samples were analyzed for 16 s rRNA detection and cytokine mRNA expression. A decreased expression of the tight junction protein Occludin, with higher levels of inflammation, greater epithelial disintegration, ulceration, loss of crypts and villi height were found in the intestines of the ROSC swine in comparison to no-ROSC. The macrophage surface antigen CD-14 staining was relatively more intense in the ROSC than in no-ROSC. Higher levels of TNF-α mRNA expression were present in the liver of the ROSC group. Finally, despite the inflammatory response and the gut mucosal alterations in ROSC group, no bacterial translocation was detected in liver, MLN and portal vein. We show that resuscitation from cardiac arrest induces inflammatory response and intestinal permeability in swine 4h after resuscitation, but not a bacterial translocation. Bacterial translocation is not an early phase phenomenon but probably part of the pathophysiologic sequelae.

The outcome of potassium dichromate exposure on histological structure of male rat sexual organs in suckling period

The aim of this study was the evaluation of integrity biomarkers of reproductive toxicity: histoarchitecture of genital organs (testis and epididymis) and sexual accessory glands (prostate, seminal vesicles and bulbo-urethral glands) at sexual maturity after exposure to potassium dichromate (Cr VI) in suckling period. Consequent to the exposure of male rat pups to hexavalent chromium during suckling period structural changes appeared in genital organs and sexual accessory glands, such as: basal membrane and epithelial disintegration, interstitial edema, wavy basal membrane due to a reduction in tubule diameter, Leydig cell necrosis; epithelial smoothing, epithelial necrosis; epithelial cells desquamation and falling in lumen. Exposure to potassium dichromate (Cr VI) during suckling period produced severe congestive and degenerative lesions in genital organs and sexual accessory glands.

An experimental model of chemically-induced ulceration of the buccal mucosa of Mus musculus

Background: Ulceration caused by chemical agents used in dental practice for in-office or home-used is a common event, resulting in discomfort and pain. Treatments for such conditions are still being developed, requiring extensive experiments both in vitro and in vivo studies. At present, a standardized experimental mouse model for mucosal ulceration caused by a chemical inducer to study the pathogenesis of ulceration and to develop medications for treatment of ulceration is still not available. The aim of this study was to create a chemically induced model of ulceration of the buccal mucosa of mice. Methods: An in vivo study model of ulceration using a total of 9 mice (Swiss Webster) was performed. All mice received 70% acetic acid application on the left buccal mucosa, while the right buccal mucosa received only saline. Clinical and histological observations of ulcer formation and healing were performed, including the presence of redness and swelling, ulcer diameter, bodyweight as well as epithelial disintegration, dilation of blood vessels, and infiltration of inflammatory cells. Results: Buccal mucosa application of 70% acetic acid generated ulcers on day 2, reached its peak on day 3 and recovered by day 14. The histological features of inflammation were also seen in the ulcer model, and the degree of inflammation was consistent with the day of ulcers. Conclusion: Chemical trauma by the administration of 70% acetic acid effectively induce ulceration on buccal mucosa in mice, and this method can be considered as a novel, reproducible, and clinically relevant model to study pathogenesis and therapeutic approach for treating oral mucosal ulceration.

Attenuation by l-thyroxine of oxidant-induced gut epithelial damage.

Objective(s):Severe injuries are often associated with tissue hypothyroidism, elevated damaging mediators in circulation, and broken gut epithelial barrier. However, the relationships between the hypothyroid state and gut epithelial damage are largely unknown. Therefore, in this study, we investigated the effects of L-thyroxine (T4) on in vitro models of intact and compromised gut epithelium. Materials and Methods:Gut epithelium equivalent was generated by cultivation of IEC-18 rat intestinal epithelial cells into transwell inserts. Confluent cultures were then compromised by scratching or H2O2 and traumatized rat sera (TUR sera) treatments. Macromolecules permeation and transepithelial electrical resistance (TEER) were evaluated by conventional methods. Morphology and scratch wound closure were assessed microscopically. Cell viability/proliferation was assessed by MTT assay. Results:Both H2O2 and TUR sera induced marked yet different types of epithelial disintegration. While H2O2 significantly increased and decreased probe permeation and TEER, respectively, TUR sera was ineffective. Cultures treated with normal rat sera (sham sera) exhibited morphology, probe permeation, and TEER comparable to those of control cultures. Presence of T4 attenuated the H2O2-induced but not TUR sera-induced damages. T4 treatment accelerated, albeit marginally, wound closure but had virtually no effects on cell viability/proliferation. Conclusion:These data suggest that different mechanisms are involved in oxidant- and trauma-induced gut epithelial barrier breakdown. Besides, they show that T4 markedly attenuates oxidant-induced gut epithelial damage. Accordingly, one may also conclude that tissue hypothyroidism does not contribute to trauma-induced gut barrier breakdown.

Abstract 3786: A study of HPV induced anal cancer progression in a raft tissue culture system

Abstract Introduction: Anal cancer is one of few malignancies for which the incidence continues to rise in the US. It is highly associated with high risk (HR) HPV infection with HPV 16 and 18 accounting for the vast majority of cases. Understanding the molecular mechanisms underlying anal cancer progression would be valuable for the purposes of developing novel clinical management strategies. However, there remains a paucity of experimental models allowing for the study of HPV-associated anal cancer development. Herein, we describe an in vitro raft tissue culture system model to study anal squamous neoplastic progression via anal intraepithelial neoplasa (AIN) induced by HPV infection. Methods: Primary human anal keratinocyte (HAK) cells were transfected with either HPV16 or 18 DNA and grown in a raft tissue culture system. The progression of dysplastic changes was sequentially examined at ~5 passage intervals by histologic review including hematoxylin-eosin (H&E) staining, immunohistochemistry (p16, Ki67, cytokeratins CK5/6, CK8/18 and CK10) and in situ hybridization (ISH) with an HPV16/18 RNA probe cocktail. The degree of dysplasia was determined by the degree of disorganization of the epithelial architecture, presence of mitotic/dysplastic cells, p16 positivity, diffuse Ki67 staining, increased number of basal-parabasal type cells (CK5/6 and CK10 markers) and lack of epithelial differentiation. Results: Advanced dysplastic changes (corresponding to AIN2-3 lesions) were already apparent at the earliest of initial passages of transfected HAK cells (passage 8 for HPV16 and 7 for HPV 18). Later passages suggested development of AIN3/squamous cell carcinoma in situ in both lines with HPV16 HAK cells showing progressively increased labeling of CK8/18 and HPV18 HAK cells demonstrating increasing cell keratinization and epithelial disintegration. In both cell lines, HPV was present in both episomal and integrated forms throughout all stages of progression as suggested by ISH signal patterns. Conclusions: A raft tissue culture model system has been developed to study HPV-induced dysplastic progression in human anal keratinocytes. Importantly, this system is of great interest as an in vitro tool to characterize the sequential molecular alterations associated with the development of anal cancer which in turn, may have implications for novel screening, prevention and treatment strategies. Citation Format: Irina V. Getun, Abidemi Ajidahun, Janice Milici, Sreejata Chatterjee, Jorge Solares, Abul Elahi, Leah E. Hendrick, Evan S. Glazer, Louisa Balazs, Craig Meyers, David Shibata. A study of HPV induced anal cancer progression in a raft tissue culture system [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3786.

Axin2 overexpression promotes the early epithelial disintegration and fusion of facial prominences during avian lip development.

The epithelial disintegration and the mesenchymal bridging are critical steps in the fusion of facial prominences during the upper lip development. These processes of epithelial-mesenchymal transition and programmed cell death are mainly influenced by Wnt signals. Axis inhibition protein2 (Axin2), a major component of the Wnt pathway, has been reported to be involved in lip development and cleft pathogenesis. We wanted to study the involvement of Axin2 in the lip development, especially during the epithelial disintegration of facial prominences. Our results show that Axin2 was expressed mainly in the epithelium of facial prominences and decreased when the prominences were about to contact each other between Hamburger-Hamilton stages 27 and 28 of chicken embryos. The epithelial integrity was destructed or kept intact by the local gain or loss of Axin2 expression, resulting in morphological changes in the facial processes and their skeletal derivatives including the maxilla, nasal, premaxilla bone, and their junctions without cleft formation. These changes were related to expression changes in nuclear β-catenin, pGSK3β, Slug, Smad3, E-cadherin, and p63. All these data indicate that Axin2 participates in the regulation of epithelial integrity and fusion by promoting epithelial disassociation, basement membrane breakdown, and seam loss during the fusion of facial prominences in lip development.

Ubc9 overexpression and SUMO1 deficiency blunt inflammation after intestinal ischemia/reperfusion

The intestinal epithelium constitutes a crucial defense to the potentially life-threatening effects of gut microbiota. However, due to a complex underlying vasculature, hypoperfusion and resultant tissue ischemia pose a particular risk to function and integrity of the epithelium. The small ubiquitin-like modifier (SUMO) conjugation pathway critically regulates adaptive responses to metabolic stress and is of particular significance in the gut, as inducible knockout of the SUMO-conjugating enzyme Ubc9 results in rapid intestinal epithelial disintegration. Here we analyzed the pattern of individual SUMO isoforms in intestinal epithelium and investigated their roles in intestinal ischemia/reperfusion (I/R) damage. Immunostaining revealed that epithelial SUMO2/3 expression was almost exclusively limited to crypt epithelial nuclei in unchallenged mice. However, intestinal I/R or overexpression of Ubc9 caused a remarkable enhancement of epithelial SUMO2/3 staining along the crypt–villus axis. Unexpectedly, a similar pattern was found in SUMO1 knockout mice. Ubc9 transgenic mice, but also SUMO1 knockout mice were protected from I/R injury as evidenced by better preserved barrier function and blunted inflammatory responses. PCR array analysis of microdissected villus-tip epithelia revealed a specific epithelial contribution to reduced inflammatory responses in Ubc9 transgenic mice, as key chemotactic signaling molecules such as IL17A were significantly downregulated. Together, our data indicate a critical role particularly of the SUMO2/3 isoforms in modulating responses to I/R and provide the first evidence that SUMO1 deletion activates a compensatory process that protects from ischemic damage.

Chemokine Signaling during Midline Epithelial Seam Disintegration Facilitates Palatal Fusion.

Disintegration of the midline epithelial seam (MES) is crucial for palatal fusion, and failure results in cleft palate. Palatal fusion and wound repair share many common signaling pathways related to epithelial-mesenchymal cross-talk. We postulate that chemokine CXCL11, its receptor CXCR3, and the cytoprotective enzyme heme oxygenase (HO), which are crucial during wound repair, also play a decisive role in MES disintegration. Fetal growth restriction and craniofacial abnormalities were present in HO-2 knockout (KO) mice without effects on palatal fusion. CXCL11 and CXCR3 were highly expressed in the disintegrating MES in both wild-type and HO-2 KO animals. Multiple apoptotic DNA fragments were present within the disintegrating MES and phagocytized by recruited CXCR3-positive wt and HO-2 KO macrophages. Macrophages located near the MES were HO-1-positive, and more HO-1-positive cells were present in HO-2 KO mice compared to wild-type. This study of embryonic and palatal development provided evidence that supports the hypothesis that the MES itself plays a prominent role in palatal fusion by orchestrating epithelial apoptosis and macrophage recruitment via CXCL11-CXCR3 signaling.

Recurrent Fusion Genes in Gastric Cancer: CLDN18-ARHGAP26 Induces Loss of Epithelial Integrity

Genome rearrangements, a hallmark of cancer, can result in gene fusions with oncogenic properties. Using DNA paired-end-tag (DNA-PET) whole-genome sequencing, we analyzed 15 gastric cancers (GCs) from Southeast Asians. Rearrangements were enriched in open chromatin and shaped by chromatin structure. We identified seven rearrangement hot spots and 136 gene fusions. In three out of 100 GC cases, we found recurrent fusions between CLDN18, a tight junction gene, and ARHGAP26, a gene encoding a RHOA inhibitor. Epithelial cell lines expressing CLDN18-ARHGAP26 displayed a dramatic loss of epithelial phenotype and long protrusions indicative of epithelial-mesenchymal transition (EMT). Fusion-positive cell lines showed impaired barrier properties, reduced cell-cell and cell-extracellular matrix adhesion, retarded wound healing, and inhibition of RHOA. Gain of invasion was seen in cancer cell lines expressing the fusion. Thus, CLDN18-ARHGAP26 mediates epithelial disintegration, possibly leading to stomach H(+) leakage, and the fusion might contribute to invasiveness once a cell is transformed.

Grainyhead-like 2 (GRHL2) distribution reveals novel pathophysiological differences between human idiopathic pulmonary fibrosis and mouse models of pulmonary fibrosis

Chronic injury of alveolar lung epithelium leads to epithelial disintegrity in idiopathic pulmonary fibrosis (IPF). We had reported earlier that Grhl2, a transcriptional factor, maintains alveolar epithelial cell integrity by directly regulating components of adherens and tight junctions and thus hypothesized an important role of GRHL2 in pathogenesis of IPF. Comparison of GRHL2 distribution at different stages of human lung development showed its abundance in developing lung epithelium and in adult lung epithelium. However, GRHL2 is detected in normal human lung mesenchyme only at early fetal stage (week 9). Similar mesenchymal reexpression of GRHL2 was also observed in IPF. Immunofluorescence analysis in serial sections from three IPF patients revealed at least two subsets of alveolar epithelial cells (AEC), based on differential GRHL2 expression and the converse fluorescence intensities for epithelial vs. mesenchymal markers. Grhl2 was not detected in mesenchyme in intraperitoneal bleomycin-induced injury as well as in spontaneously occurring fibrosis in double-mutant HPS1 and HPS2 mice, whereas in contrast in a radiation-induced fibrosis model, with forced Forkhead box M1 (Foxm1) expression, an overlap of Grhl2 with a mesenchymal marker was observed in fibrotic regions. Grhl2's role in alveolar epithelial cell plasticity was confirmed by altered Grhl2 gene expression analysis in IPF and further validated by in vitro manipulation of its expression in alveolar epithelial cell lines. Our findings reveal important pathophysiological differences between human IPF and specific mouse models of fibrosis and support a crucial role of GRHL2 in epithelial activation in lung fibrosis and perhaps also in epithelial plasticity.

Tetraspanin CD151 Protects against Pulmonary Fibrosis by Maintaining Epithelial Integrity

Idiopathic pulmonary fibrosis (IPF) is a chronic pulmonary disorder of unknown etiology with few treatment options. Although tetraspanins are involved in various diseases, their roles in fibrosis have not been determined. To investigate the role of tetraspanin CD151 in pulmonary fibrosis. CD151 knockout (KO) mice were studied by histological, biochemical, and physiological analyses and compared with wild-type mice and CD9 KO mice. Further mechanistic analyses were performed in vitro, in vivo, and on samples from patients with IPF. A microarray study identified an enrichment of genes involved in connective tissue disorders in the lungs of CD151 KO mice, but not in CD9 KO mice. Consistent with this, CD151 KO mice spontaneously exhibited age-related pulmonary fibrosis. Deletion of CD151 did not affect pulmonary fibroblast functions but instead degraded epithelial integrity via attenuated adhesion strength on the basement membrane; CD151-deleted alveolar epithelial cells exhibited increased α-SMA expression with activation of p-Smad2, leading to fibrotic changes in the lungs. This loss of epithelial integrity in CD151 KO lungs was further exacerbated by intratracheal bleomycin exposure, resulting in severe fibrosis with increased mortality. We also observed decreased numbers of CD151-positive alveolar epithelial cells in patients with IPF. CD151 is essential for normal function of alveolar epithelial cells; loss of CD151 causes pulmonary fibrosis as a result of epithelial disintegrity. Given that CD151 may protect against fibrosis, this protein represents a novel target for the treatment of fibrotic diseases.

The Alimentary Canal of the African Lungfish Protopterus annectens During Aestivation and After Arousal

We describe the structural modifications that occur in the alimentary canal of the African lungfish Protopterus annectens during aestivation and after arousal. With fasting, all gut segments undergo structural modifications. The epithelium covering the intestinal vestibule undergoes bursts of activation at 4 months of aestivation, adopting a more quiescent appearance at 6 months. The ridge area of the spiral intestine shows, at 4 months of aestivation, epithelial disintegration, cell desquamation, cell death, and loss of the freshwater phenotype. Surprisingly, the epithelium adopts a stratified appearance at 6 months of aestivation. Except for epithelial disintegration, the smooth portion of the spiral intestine follows a similar pattern of modifications than the ridge area. The entire epithelium of spiral intestine appears to be renewed during aestivation. The presence of intraepithelial mast cells suggests that inflammation is part of the cellular response to aestivation. After arousal, cell phenotypes are restored in about 6 days, but full structural recovery is not attained during the experimental period (15 days post-aestivation). Several aspects of the cellular response to fasting are shared by a wide range of animal groups. This commonality agrees with the presence of a character that allows to adjust the structural and functional properties of the gut to food availability and food quality, and to the characteristics of the fasting episodes.

E-cadherin regulates MAL-SRF-mediated transcription in epithelial cells

Epithelial junctions are dynamically and functionally linked to the actin cytoskeleton, and their disassembly is a key event during physiological and pathological processes. We recently showed that epithelial disintegration facilitates transcriptional activation via Rac, G-actin, MAL (also known as MRTF) and serum response factor (SRF). Here, we investigate which specific component of the epithelial junction is essential for this MAL-SRF-mediated transcription. The Ca(2+)-dependent dissociation of polarised epithelial cells depleted of ZO proteins - which form adherens junctions (AJs) but completely lack tight junctions (TJs) - fully activated SRF. By contrast, AGS gastric adenocarcinoma epithelial cells, which form TJs but are deficient in E-cadherin, and therefore also in AJs, failed to activate SRF. The introduction of wild-type E-cadherin in AGS cells restored AJ formation and MAL-SRF inducibility. To gain further insight into the membrane-proximal signalling, AGS cells were stably transfected with E-cadherin-alpha-catenin fusions. Despite restored formation of cell-cell contacts containing the nectin-afadin complex and p120-catenin, these cells did not activate SRF upon junction dissociation, suggesting that signal transmission depends on the C-terminal tail of E-cadherin. We conclude that the dissociation of intercellular E-cadherin interactions from AJs, and signals originating from the C-terminal region covering the beta-catenin-binding site of E-cadherin, are essential for transcriptional activation via Rac, MAL and SRF, whereas TJs are not involved.

Epithelial cell-cell contacts regulate SRF-mediated transcription via Rac-actin-MAL signalling

Epithelial cell-cell junctions are specialised structures connecting individual cells in epithelial tissues. They are dynamically and functionally linked to the actin cytoskeleton. Disassembly of these junctions is a key event during physiological and pathological processes, but how this influences gene expression is largely uncharacterised. Here, we investigate whether junction disassembly regulates transcription by serum response factor (SRF) and its coactivator MAL/MRTF. Ca2+-dependent dissociation of epithelial integrity was found to correlate strictly with SRF-mediated transcription. In cells lacking E-cadherin expression, no SRF activation was observed. Direct evidence is provided that signalling occurs via monomeric actin and MAL. Dissociation of epithelial junctions is accompanied by induction of RhoA and Rac1. However, using clostridial cytotoxins, we demonstrate that Rac, but not RhoA, is required for SRF and target gene induction in epithelial cells, in contrast to serum-stimulated fibroblasts. Actomyosin contractility is a prerequisite for signalling but failed to induce SRF activation, excluding a sufficient role of the Rho-ROCK-actomyosin pathway. We conclude that E-cadherin-dependent cell-cell junctions facilitate transcriptional activation via Rac, G-actin, MAL and SRF upon epithelial disintegration.

Five-Year Outcome of LASIK for Myopia

To investigate the efficacy and safety of LASIK over a 5-year postoperative period. Observational case series. We examined 779 eyes in 402 patients with myopia or myopic astigmatism who underwent LASIK to correct their refractive errors and received regular postoperative assessments for 5 years. Postoperative examinations were performed 1 day; 1 week; 1, 3, and 6 months; and 1, 2, 3, 4, and 5 years after LASIK surgery. We evaluated changes in uncorrected visual acuity (UCVA) (logarithm of the minimum angle of resolution [logMAR]), manifest refraction, best-corrected visual acuity (BCVA) (logMAR), intraocular pressure, corneal thickness, corneal endothelial cell counts, and complications. Preoperative UCVA of 1.27 improved to -0.03 at 1 day after surgery and -0.08 at 1 month and revealed minimal but significant decreases thereafter. Postoperative manifest refraction was also improved by surgery, showing minimal but significant regression after 1 year. Final BCVA loss was seen in 10 eyes of 7 patients; in 7 cases, there were obvious reasons such as the progression of cataracts in 3 eyes, epithelial disintegrity due to dry eye in 2 eyes, irregular astigmatism due to flap striae in 1 eye, and age-related macular dystrophy in 1 eye. Intraocular pressure and corneal thickness decreased by 4.0 mmHg and 76.9 microm, respectively, due to surgery, but remained stable throughout the follow-up period. Corneal endothelial cell counts (2689.0+/-232.9 cells/mm(2) before surgery) showed a statistically significant decrease at 5 years after surgery (2658.0+/-183.1 cells/mm(2); 1.2% loss for 5 years), likely within the range due to physiological age-related loss. No serious, vision-threatening, irreversible complication such as keratectasia or progressive endothelial cell loss was observed. LASIK surgery is an effective and safe procedure for correcting myopia/myopic astigmatism as long as inclusion and exclusion criteria are strictly respected. However, minimal regression occurred during the 5-year investigative period.

Recombinant human interleukin 11 and bacterial infection in patients with haemological malignant disease undergoing chemotherapy: a double-blind placebo-controlled randomised trial

Bacteraemia in patients with haematological malignant disease causes substantial morbidity. Recombinant human interleukin 11 (rhIL-11) prevents gastrointestinal epithelial disintegrity and has immunomodulatory actions. Our aim was to ascertain whether or not treatment with rhIL-11 can prevent gut-associated infections. We did a double-blind placebo-controlled randomised trial, to which we enrolled 40 patients with haematological malignant disease who were undergoing chemotherapy. Patients received either rhIL-11 50 microg/kg (n=20) or placebo (n=20) daily by subcutaneous injection from the day before the start of chemotherapy until resolution of neutropenia or for 21 days, whichever was longer. Our primary outcome measure was a reduction in bacteraemia. Analysis was by intention to treat. Significantly fewer patients who received rhIL-11 rather than placebo developed bacteraemia, particularly of gastrointestinal origin: the proportion of patients with at least one positive blood culture was 0.65 and 0.25, respectively (p=0.02). The numbers of patients (placebo vs rhIL-11) for each number of distinct isolates were: no organism isolated seven versus 15, one organism nine versus four, two organisms two versus one, three organisms one versus none, and four organisms one versus none (p=0.01), suggesting a lower bacterial load in the rhIL-11 than in the placebo group. Time to first bacteraemic event was longer in patients who received rhIL-11 (p=0.03) than in those who received placebo. rhIL-11 reduces the frequency and load of bacteraemia in patients with haematological malignant disease undergoing chemotherapy, possibly by gastrointestinal cytoprotective or immunological mechanisms [corrected].