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The importance of lubrication between oral surfaces provided by the salivary film is most acutely apparent when it is disrupted, a prevalent consequence of salivary gland hypofunction experienced with aging, a symptom of certain diseases, or a side effect of some medical interventions. Sufferers report difficulty with speech and oral food processing and collectively is detrimental to quality of life. Polyethylene glycol (PEG) is widely employed as a successful biocompatible boundary lubricant in engineering and biomedical applications. It is hypothesized that the immobilization of PEG to biological materials such as oral epithelial cells and tissue can mimic the salivary film and provide durable relief from the symptoms of mucosal dryness. To do so, PEG is functionalized with a sugar binding lectin (wheat germ agglutinin) to enhance epithelial adhesion through lectin-sugar interactions. Retention and lubricity are characterized on an ex vivo oral tissue tribology rig. WGA-PEG coats and retains on mucin films, oral epithelial cells, and porcine tongue tissue, and offers sustained reduction in coefficient of friction (COF). WGA-PEG could be developed into a useful topical treatment for reducing oral friction and the perception of dry mouth., QC 20220615

Background and Aim: Diagnosis and management of inflammatory bowel disease (IBD) requires a clinical approach, combining information from histology and other modalities for clinical decision making. However, histology is costly due to specimen processing, and intra- and interobservational variation and the significance of subtle findings remain unclear when translating into clinical practice. Biospectroscopy involves combining microscopy with spectroscopy and provides chemical and imaging identification for complex biological specimens, which, as an advantage, do not require any chemical processing techniques or staining. Briefly, infrared (IR) or laser energies vibrate biochemical bonds in biospecimens (fluids or tissue), which are then recorded as absorbance intensities in large hyperspectral datasets. Regression and classification algorithms used in multivariate analysis for machine learning are then applied for modeling and predictive capacity. To date, there has been little published research using this technology for assessing IBD. We aimed to undertake an explorative analysis of intestinal inflammation using Fourier-transform infrared microspectroscopy, develop a model for assessing severity of colitis, and identify potential spectral biomarkers for colitis. This study was a proof-of-concept study using a gold-standard murine colitis model to inform further research in human work. Method(s): We used formalin-fixed paraffin-embedded intestinal sections from C57/BL6 mice (30 mice, three sections per mouse) with chronic colitis induced by 3% oral dextran sodium sulfate (n = 27), of which a subset (n = 14) were treated with intravenous human amnion epithelial cells and showed a reduction in histological severity of inflammation. Tissue sections were placed on a reflective slide, and an adjacent 5 mum section was stained with hematoxylin and eosin for comparison. Background scans and regions of interest from the intestinal mucosa were imaged using the Agilent Cary

Aim: To assess whether ASK1 blockade inhibits the progression of experimental diabetic kidney disease by reducing stress-induced premature senescence (SIPS) and its consequences. Background(s): The development of SIPS in kidney tubular epithelial cells (TEC) results in the release of stress-associated secretory proteins (SASPs), including proinflammatory and profibrotic factors which promote progression of kidney diseases. Activation of p38 MAPK is a key mechanism promoting SIPS. Therefore, we hypothesized that blockade of ASK1-p38 MAPK signalling would inhibit kidney diseases by reducing SIPS and SASPs. Method(s): Development of SIPS and production of SASPs were assessed in archived tissue from a model of progressive fibrotic kidney disease- streptozotocin (STZ)-induced diabetic kidney disease (DKD) in hypertensive endothelial nitric oxide synthase (Nos3) deficient mice. Mice with DKD were treated with an ASK1 inhibitor (GS-444217) as either an early intervention (2-8 weeks after STZ) or a late intervention (8-15 weeks after STZ). Result(s): STZ-induced diabetes resulted in the development of senescent TEC in Nos3-/- mice. Treatment with GS-444217 blocked activation of the p38 MAPK pathway without affecting hyperglycaemia. This was associated with reduced levels of cellular senescence (Cdkn2A) and SASPs (CCL2, TNF-alpha, TGF-beta1), which correlated with diminished levels of kidney fibrosis and renal function impairment. Therapeutic blockade of ASK1 was effective in reducing SIPS as both an early intervention (Cdkn2A #51%; P = 0.0005) and as a late intervention (Cdkn2A #45%; P = 0.02) in the DKD model. Conclusion(s): Our data show that ASK1 inhibition reduces SIPS and SASPs independent of the underlying diabetic state. This may contribute to the overall benefit seen with ASK1 blockade in DKD. Further studies are warranted to explore this mechanism.

Purpose of Review: In this review, we will explore the clinical and commercial focus on amniotic membranes (AMs) and their use in tissue engineering (TE). We will showcase the therapeutic potential of AM-isolated epithelial cells (hAECs) and the prospective use of their secreted factors, extracellular vesicles (EVs), in cell and non-cell therapies. Recent Findings: The potential of the hAECs as a therapeutic source has been investigated in various preclinical models with some progressing into phase I clinical trials to evaluate their safety. Additionally, multiple animal studies showcase the therapeutic potential of EVs as non-cellular treatments. Summary: The amniotic membrane (AM) has been used as a form of regenerative medicine in wound healing for burns and ulcerated surfaces and in ophthalmology for over a century. In the last few decades, research has looked to the use of the various stem cells that can be isolated from the AM. The use of AM-isolated hAECs has proven rather promising with phase I clinical trials currently underway across life-threatening diseases in both pediatric and adult populations. However, due to limitations of using cell-based therapies (e.g., cost of production, delivery restricted to major hospitals, etc.), attention has turned to investigating EVs secreted by the cells.Copyright © 2021, The Author(s), under exclusive licence to Springer Nature Switzerland AG part of Springer Nature.

Human amnion epithelial cells (hAECs) exert potent antifibrotic and anti-inflammatory effects when transplanted into preclinical models of tissue fibrosis. These effects are mediated in part via the secretion of soluble factors by hAECs which modulate signaling pathways and affect cell types involved in inflammation and fibrosis. Based on these reports, we hypothesized that these soluble factors may also support liver regeneration during chronic liver injury. To test this, we characterized the effect of both hAECs and hAEC-conditioned medium (CM) on liver repair in a mouse model of carbon tetrachloride (CCl4)-induced fibrosis. Liver repair was assessed by liver fibrosis, hepatocyte proliferation, and the liver progenitor cell (LPC) response. We found that the administration of hAECs or hAEC-CM reduced liver injury and fibrosis, sustained hepatocyte proliferation, and reduced LPC numbers during chronic liver injury. Additionally, we undertook in vitro studies to document both the cell-cell and paracrine-mediated effects of hAECs on LPCs by investigating the effects of co-culturing the LPCs and hAECs and hAEC-CM on LPCs. We found little change in LPCs co-cultured with hAECs. In contrast, hAEC-CM enhances LPC proliferation and differentiation. These findings suggest that paracrine factors secreted by hAECs enhance liver repair by reducing fibrosis while promoting regeneration during chronic liver injury.Copyright © The Author(s) 2020.

Aim: To investigate the role of the c-Jun amino terminal kinase (JNK) signalling pathway in mouse models of Aristolochic acid (AA)-induced acute and chronic kidney injury. Background(s): AA is a toxin that damages tubular epithelial cells of the kidney and is the cause of Chinese Herb Nephropathy and Balkan Nephropathy. Exposure of cultured tubular epithelial cells to AA induces a pro-fibrotic response via the JNK pathway. Method(s): In the acute model, groups (n = 7-8) of mice received a single injection of 5 mg/kg AA and were treated with vehicle, JNK inhibitor (CC-930) at 75 mg/kg/BID or untreated from day 0 until being killed on day 3. In the chronic model, mice (n = 10) received 2 mg/kg AA every second day from day 0 to day 22, and were treated with vehicle or CC-930 for the entire period. Result(s): In the acute model, CC-930 treatment substantially inhibited JNK signalling and gave significant protection from AA-induced renal function impairment (normal 8.3 +/- 1.5; untreated 33.4 +/- 4.1; vehicle 35.0 +/- 5.8; CC- 930 23.3 +/- 2.1 mumol/L serum creatinine: P < 0.001) and tubular cell damage (KIM-1 and NGAL mRNA levels; P < 0.001). This protection was associated with reduced macrophage infiltration and reduced expression of proinflammatory molecules (TNF, IL-36A). In the chronic model, CC-930 treatment significantly inhibited JNK signalling, but did not affect AA-induced renal function impairment, tubular cell damage or renal fibrosis, despite a significant reduction in the macrophage pro-inflammatory response. Conclusion(s): Blockade of JNK signalling with CC-930 suppressed the acute effects of AA on tubular cell damage and renal function impairment, but was unable to protect the kidney against the effects of chronic AA exposure.

Chronic kidney disease (CKD) remains a major epidemiological, clinical, and biomedical challenge. During CKD, renal tubular epithelial cells (TECs) present a persistent inflammatory and profibrotic response. Fatty acid oxidation (FAO), the main source of energy for TECs, is reduced in kidney fibrosis and contributes to its pathogenesis. To determine whether gain of function in FAO (FAO-GOF) could protect from fibrosis, we generated a conditional transgenic mouse model with overexpression of the fatty acid shuttling enzyme carnitine palmitoyl-transferase 1A (CPT1A) in TECs. Cpt1a-knockin (CPT1A-KI) mice subjected to 3 models of renal fibrosis (unilateral ureteral obstruction, folic acid nephropathy [FAN], and adenine-induced nephrotoxicity) exhibited decreased expression of fibrotic markers, a blunted proinflammatory response, and reduced epithelial cell damage and macrophage influx. Protection from fibrosis was also observed when Cpt1a overexpression was induced after FAN. FAO-GOF restored oxidative metabolism and mitochondrial number and enhanced bioenergetics, increasing palmitate oxidation and ATP levels, changes that were also recapitulated in TECs exposed to profibrotic stimuli. Studies in patients showed decreased CPT1 levels and increased accumulation of shortand middle chain acylcarnitines, reflecting impaired FAO in human CKD. We propose that strategies based on FAO-GOF may constitute powerful alternatives to combat fibrosis inherent to CKD.

We investigated SARS-CoV-2 potential tropism by surveying expression of viral entry-associated genes in single-cell RNA-sequencing data from multiple tissues from healthy human donors. We co-detected these transcripts in specific respiratory, corneal and intestinal epithelial cells, potentially explaining the high efficiency of SARS-CoV-2 transmission. These genes are co-expressed in nasal epithelial cells with genes involved in innate immunity, highlighting the cells’ potential role in initial viral infection, spread and clearance. The study offers a useful resource for further lines of inquiry with valuable clinical samples from COVID-19 patients and we provide our data in a comprehensive, open and user-friendly fashion at www.covid19cellatlas.org., QC 20200625

Background & Aim: Background: Chronic liver disease is characterised by progressive hepatocyte injury, inflammation and accumulation of extracellular matrix. Liver progenitor cells (LPC) are bipotent cells induced during liver injury that can regenerate both hepatocytes and cholangiocytes. In addition, macrophages play a pivotal role in fibrogenesis and fibrosis resolution. Classically, macrophages are classified into M1 (pro-inflammatory macrophages) and M2 (pro-resolution macrophages). Recently, Ly6C expression has been used to identify distinct macrophage populations, Ly6Chi (inflammatory) and Ly6Clow (resolution). We are developing a therapy for chronic liver fibrosis using human amnion epithelial cells (hAECs). The choline-deficient ethionine-supplemented (CDE) diet is a non-invasive and quick method to induce liver injury and LPC proliferation. Here, we aimed to investigate the effect of hAECs on macrophages and LPC during liver injury induced by a CDE diet. Aim(s): Methods, Results & Conclusion Methods: C57/B16J mice were commenced on a CDE diet, then 2 or 4 million hAECs were infused intraperitoneally 24h later and the CDE diet continued. Outcomes were assessed at days 1, 3, 5, 14, 21 and 42 following hAEC administrations. Sirius red staining and immunohistochemistry were used to determine fibrosis and identify LPC in liver sections. Macrophages and LPC were further identified and characterised using flow cytometery sorting and PCR. Result(s): In CDE treated mice, hAEC significantly reduced liver fibrosis compared to untreated mice. hAEC decreased the number of proinflammatory (Ly6Chi) monocyte-derived macrophages (MDM) and increased the number of anti-inflammatory and tissue remodelling (Ly6Clow) MDM. hAEC also enhanced the expression of matrix-metalloproteinase (MMP)12 and MMP13 in Ly6CLow sorted cells. LPC numbers were reduced significantly in hAEC treated mice. Conclusion(s): Our data confirm the potential efficacy of hAECs as an effective therapy in liv

Activation of cyclic GMP-AMP (cGAMP) synthase (cGAS) plays a critical role in antiviral responses to many DNA viruses. Sensing of cytosolic DNA by cGAS results in synthesis of the endogenous second messenger cGAMP that activates stimulator of interferon genes (STING) in infected cells. Critically, cGAMP can also propagate antiviral responses to uninfected cells through intercellular transfer, although the modalities of this transfer between epithelial and immune cells remain poorly defined. We demonstrate here that cGAMP-producing epithelial cells can transactivate STING in cocultured macrophages through direct cGAMP transfer. cGAMP transfer was reliant upon connexin expression by epithelial cells and pharmacological inhibition of connexins blunted STING-dependent transactivation of the macrophage compartment. Macrophage transactivation by cGAMP contributed to a positive-feedback loop amplifying antiviral responses, significantly protecting uninfected epithelial cells against viral infection. Collectively, our findings constitute the first direct evidence of a connexin-dependent cGAMP transfer to macrophages by epithelial cells, to amplify antiviral responses. IMPORTANCE Recent studies suggest that extracellular cGAMP can be taken up by macrophages to engage STING through several mechanisms. Our work demonstrates that connexin-dependent communication between epithelial cells and macrophages plays a significant role in the amplification of antiviral responses mediated by cGAMP and suggests that pharmacological strategies aimed at modulating connexins may have therapeutic applications to control antiviral responses in humans.Copyright © 2020 Pepin et al.

Background: Regenerative cell therapies for various neonatal morbidities are increasingly entering early phase clinical trials. However, infusion protocols for delivering cells have been largely adopted from adult protocols and have not been evaluated in infants. We aimed to explore the effects of the different cell infusion parameters on cell delivery to define the optimum protocol for the administration of a leading cell therapy, human amnion epithelial cells (HAECS), to extremely preterm infants. Method(s): Standard cell infusion protocols were modelled, varying the infusion parameters including: albumin concentration (2% vs. 4%), syringe orientation (horizontal vs. vertical), IVline volume (0.2-2.2 ml), and flow rate (3-15 ml/HR). Result(s): The standard (previously published) cell infusion protocol delivered a mean (SD) of 20(9)% of intended HAEC dose. Increasing albumin concentration to 4%, positioning the syringe vertically, and decreasing IV-line volume to 0.6 ml increased cell delivery to a mean (SD) of 98(6)% of intended cell dose. Flow rate had little effect on dose delivery when other conditions were optimised. Conclusion(s): It is essential that cell infusion protocols are optimised for small volume delivery in preterm infants. We describe the refinement and validation of a cell infusion protocol that offers reliable delivery of intended cell doses, suitable for extremely preterm neonates. Previously published work should be viewed with caution given the possibility that only a fraction of intended cell dose was actually delivered.

OBJECTIVE: Endometriosis is a common and painful condition characterised by the formation of endometrial lesions within the peritoneal cavity. Previous studies have suggested a role for hedgehog signalling in the pathogenesis of endometriosis. We investigated the role of hedgehog signalling in the establishment of endometriosis lesions using 5E1, a hedgehog ligand neutralising antibody, and a mouse model of endometriosis. To mimic the initiation of endometriosis by retrograde menstruation, which is believed to occur in humans, donor mice underwent an artificial menstruation protocol. Fragments of menstrual endometrium were injected into the peritoneal cavity of estrogen primed recipients. Recipients received twice weekly injections of 5E1 or an isotype matched control antibody for three weeks. Lesions were collected and analysed for markers of epithelium, proliferation and apoptosis by immunofluorescence microscopy. RESULT(S): Treatment with 5E1 reduced the number of lesions found on the mesentery. No significant changes were found in the size of lesions, abundance of endometrial epithelial cells, proliferation or apoptosis.

INTRODUCTION: Bronchopulmonary dysplasia (BPD), an important sequela of preterm birth, is associated with long-term abnormalities of lung function and adverse neurodevelopmental outcomes. Inflammation, inhibition of secondary septation and vascular maldevelopment play key roles in the pathogenesis of BPD. Human amnion epithelial cells (hAECs), stem-like cells, derived from placental tissues are able to modulate the inflammatory milieu and, in preclinical studies of BPD-like injury, restore lung architecture and function. Allogeneic hAECs may present a new preventative and reparative therapy for BPD. METHODS AND ANALYSIS: In this two centre, phase Icell dose escalation study we will evaluate the safety of intravenous hAEC infusions in preterm infants at high risk of severe BPD. Twenty-four infants born at less than 29 weeks' gestation will each receive intravenous hAECs beginning day 14 of life. We will escalate the dose of cells contained in a single intravenous hAEC infusion in increments from 2million cells/kg to 10million cells/kg. Further dose escalation will be achieved with repeat infusions given at 5day intervals to a maximum total dose of 30million cells/kg (three infusions). Safety is the primary outcome. Infants will be followed-up until 2 years corrected age. Additional outcome measures include a description of infants' cytokine profile following hAEC infusion, respiratory outcomes including BPD and pulmonary hypertension and other neonatal morbidities including neurodevelopmental assessment at 2 years. ETHICS AND DISSEMINATION: This study was approved on the June12th, 2018 by the Human Research Ethics Committee of Monash Health and Monash University. Recruitment commenced in August 2018 and is expected to take 18 months. Accordingly, follow-up will be completed mid-2022. The findings of this study will be disseminated via peer-reviewed journals and at conferences. PROTOCOL VERSION: 5, 21 May 2018. TRIAL REGISTRATION NUMBER: ACTRN12618000920291; Pre-resul

Background & Aim: Background: Advances in neonatal intensive care have been associated with improved survival of preterm infants. But along with survival, the morbidity bronchopulmonary dysplasia (BPD), the chronic lung disease of prematurity, is also increasing. BPD is associated with adverse respiratory outcomes into adulthood and greater risk of adverse neurodevelopment. Infants born <= 28 weeks' gestation, during the canalicular phase of lung development prior to alveolar and distal capillary development, are at greatest risk of BPD. Disruption of lung maturation by the pro-inflammatory environment created by lifesaving neonatal intensive care interventions results in the BPD we see today - a disease characterised by arrested development with simplified alveoli and abnormal angiogenesis. As the role of inflammation in BPD comes into focus, cell therapy presents a promising intervention for its prevention. Human amnion epithelial cells (hAECs), stem-like cells derived from placental tissues, have multi-lineage potential, are immune privileged and, in small animal models are non-teratogenic. Preclinical work demonstrates the cells capacity to modulate the inflammatory environment to both prevent injury and aid repair of established injury in diverse models of lung disease. A first-in-human study recently demonstrated the safety of low dose hAECs in preterm infants with established BPD. However, hAECs are likely to have greater therapeutic potential when used to prevent lung injury thus, the next step in translation is to evaluate the safety of hAECs in less mature infants at high risk of developing BPD. Aim(s): To evaluate the safety of escalating doses of intravenous hAECs in infants born <= 28 weeks' gestation at high risk of BPD. Methods, Results & Conclusion Method: In this phase 1 study, 24 infants born at <=28 weeks' gestation at high risk of BPD will each receive intravenous hAECs from Day 14 of life. The infusion dose will escalate from 2 million cells/kg

Increased intragenic DNA methylation of the Fragile X Related Epigenetic Element 2 (FREE2) in blood has been correlated with lower intellectual functioning in females with fragile X syndrome (FXS). This study explored these relationships in a paediatric cohort of males with FXS using Buccal Epithelial Cells (BEC). BEC were collected from 25 males with FXS, aged 3 to 17 years and 19 age-matched male controls without FXS. Methylation of 9 CpG sites within the FREE2 region was examined using the EpiTYPER approach. Full Scale IQ (FSIQ) scores of males with FXS were corrected for floor effect using the Whitaker and Gordon (WG) extrapolation method. Compared to controls, children with FXS had significant higher methylation levels for all CpG sites examined (p<3.3x10-7), and within the FXS group, lower FSIQ (WG corrected) was associated with higher levels of DNA methylation, with the strongest relationship found for CpG sites within FMR1 intron 1 (p<5.6x10-5). Applying the WG method to the FXS cohort unmasked significant epi-genotype-phenotype relationships. These results extend previous evidence in blood to BEC and demonstrate FREE2 DNA methylation to be a sensitive epigenetic biomarker significantly associated with the variability in intellectual functioning in FXS.

Oncogenic KRAS mutations are major drivers of lung adenocarcinoma (LAC), yet the direct therapeutic targeting of KRAS has been problematic. Here, we reveal an obligate requirement by oncogenic KRAS for the ADAM17 protease in LAC. In genetically engineered and xenograft (human cell line and patient-derived) KrasG12D-driven LAC models, the specific blockade of ADAM17, including with a non-toxic prodomain inhibitor, suppressed tumor burden by reducing cellular proliferation. The pro-tumorigenic activity of ADAM17 was dependent upon its threonine phosphorylation by p38 MAPK, along with the preferential shedding of the ADAM17 substrate, IL-6R, to release soluble IL-6R that drives IL-6 trans-signaling via the ERK1/2 MAPK pathway. The requirement for ADAM17 in KrasG12D-driven LAC was independent of bone marrow-derived immune cells. Furthermore, in KRAS mutant human LAC, there was a significant positive correlation between augmented phospho-ADAM17 levels, observed primarily in epithelial rather than immune cells, and activation of ERK and p38 MAPK pathways. Collectively, these findings identify ADAM17 as a druggable target for oncogenic KRAS-driven LAC and provide the rationale to employ ADAM17-based therapeutic strategies for targeting KRAS mutant cancers.Copyright © 2019 The Authors. Published under the terms of the CC BY 4.0 license

Aim: Albumin can be covalently modified at surface lysine residues and thus the circulation contains a mixture of native albumin (i.e. not modified) and albumin with varying degrees of modification. Uptake and lysosomal degradation of glomerular filtered albumin by proximal tubular cells via the megalin scavenger receptor is considered an important mechanism to limit albumin loss in the urine. However, whether this is a general mechanism of tubular uptake of albumin or if this is restricted to modified albumin is unknown. To address this question, we investigated the uptake of modified versus native albumin by proximal tubular cells. Method(s): A well-characterized proximal tubular cell model of albumin uptake was used to compare the uptake of modified albumin (covalent labelling of lysine residues with fluorescent probes) to that of native recombinant human albumin (rHA) labelled with 14C during protein synthesis (14C-rHA). Result(s): Opossum kidney (OK) cells showed significant uptake of fluorescence-labelled albumin via an endocytosis mechanism. This uptake was inhibited by an equimolar ratio of different types of covalently modified albumin; however, purified bovine serum albumin and rHA failed to compete with the uptake of fluorescence-labelled albumin. In contrast, OK cells failed to endocytose native 14C-rHA despite efficiently endocytosing covalently modified rHA. Conclusion(s): Our studies show that OK cells preferentially endocytose covalently-modified albumin compared to native albumin. This apparent selectivity of the megalin scavenger receptor complex suggests a specific role for this pathway in the removal of modified albumin from the circulation.Copyright © 2017 Asian Pacific Society of Nephrology

Glomerular crescent formation is a hallmark of rapidly progressive forms of glomerulonephritis. Thrombosis and macrophage infiltration are features of crescent formation in human and experimental kidney disease. Protease-activated receptor-2 (PAR-2) is a G-protein coupled receptor that links coagulation and inflammation. This study investigated whether pharmacological inhibition of PAR-2 can suppress glomerular crescent formation in rat nephrotoxic serum nephritis (NTN). Disease was induced in Wistar Kyoto rats by immunisation with sheep IgG followed by administration of sheep nephrotoxic serum. Rats (n = 8/group) received the PAR-2 antagonist (GB88, 10 mg/kg/p.o.), vehicle or no treatment starting 3 days before nephrotoxic serum injection and continuing until day 14. Vehicle and untreated rats developed thrombosis and macrophage infiltration in the glomerular tuft and Bowman's space in conjunction with prominent crescent formation. Activation of JNK signalling and proliferation in parietal epithelial cells was associated with crescent formation. GB88 treatment significantly reduced crescent formation with a substantial reduction in glomerular thrombosis, reduced macrophage infiltration in Bowman's space, and reduced activation of parietal epithelial cells. However, GB88 did not protect against the development of proteinuria, renal function impairment, inflammation or tubular cell damage in the NTN model. In conclusion, PAR-2 plays a specific role in glomerular crescent formation by promoting glomerular thrombosis, macrophage accumulation in Bowman's space and activation of parietal epithelial cells.Copyright © 2019 John Wiley & Sons Australia, Ltd

Background and Aim: Inflammatory bowel disease (IBD) requires longterm therapy to control symptoms and prevent complications. Despite therapeutic advances, some patients experience side effects or are refractory to conventional treatment. Current therapies are unable to reduce or reverse established intestinal fibrosis. Isolated from placentas, human amnion epithelial cells (hAECs) are ethically sourced pluripotent stem cells that possess anti-inflammatory and antifibrotic properties. With no tumorogenicity and an excellent safety profile, hAECs are a promising novel therapy for IBD. We examined the efficacy of hAECs and their conditioned media (hAEC-CM) in a dextran sulfate sodium (DSS)-induced murine model of chronic colitis. Method(s): C57BL/6J mice received a cycle of 2% DSS (w/v) in normal water for 1 week and then normal water for 2 weeks. This 3-week regimen was repeated twice until cull at week 10. Treatment began at week 6, with one group (Chr-hAEC, n = 14) receiving weekly tail vein injections of 2 x 106 hAECs, and another group (Chr-CM, n = 15) receiving 250-muL hAEC-CM three times weekly. Normal mice (n = 7) and DSS control mice (Chr-DSS, n = 13) received no therapy. Mice were monitored daily. Colitis severity was scored using HE-stained colon sections. Interleukin (IL)-1beta, tumor necrosis factor (TNF)-alpha, IL-6, transforming growth factor (TGF)-beta, interferon (IFN)-gamma, IL-4, and IL- 10 were measured by ELISA. Fibrosis extent was measured via computer- assisted morphometry of picro-sirius-stained colon sections. Fibroblasts and macrophages were identified by immunohistochemistry (alpha-SMA, F4/80). Result(s): Bodyweight significantly improved with hAECs (10%, P < 0.05) and hAEC-CM (12.5%, P < 0.001) compared with untreated mice on DSS (4.2%). hAECs and hAEC-CM also prevented colon shortening (6.8 cm, P < 0.001; and 7.3 cm, P < 0.0001, respectively) compared with controls (5.9 cm). Splenomegaly was greater in Chr-DSS mice (212 mg) than mice treat

Recent developments in optical tissue clearing have been difficult to apply for the morphometric analysis of organs with high cellular content and small functional structures, such as the kidney. Here, we establish combinations of genetic and immuno-labelling for single cell identification, tissue clearing and subsequent de-clarification for histoimmunopathology and transmission electron microscopy. Using advanced light microscopy and computational analyses, we investigated a murine model of crescentic nephritis, an inflammatory kidney disease typified by immune-mediated damage to glomeruli leading to the formation of hypercellular lesions and the rapid loss of kidney function induced by nephrotoxic serum. Results show a graded susceptibility of the glomeruli, significant podocyte loss and capillary injury. These effects are associated with activation of parietal epithelial cells and formation of glomerular lesions that may evolve and obstruct the kidney tubule, thereby explaining the loss of kidney function. Thus, our work provides new high-throughput endpoints for the analysis of complex tissues with single-cell resolution.Copyright © 2019 International Society of Nephrology

Beside the classical flat parietal epithelial cells (PECs), we investigated proximal tubular epithelial-like cells, a neglected subgroup of PECs. These cells, termed cuboidal PECs, make up the most proximal part of the proximal tubule and may also line parts of Bowman's capsule. Additionally, a third intermediate PEC subgroup was identified at the junction between the flat and cuboidal PEC subgroups at the tubular orifice. The transgenic mouse line PEC-rtTA labeled all three PEC subgroups. Here we show that the inducible Pax8-rtTA mouse line specifically labeled only cuboidal and intermediate PECs, but not flat PECs. In aging Pax8-rtTA mice, cell fate mapping showed no evidence for significant transdifferentiation from flat PECs to cuboidal or intermediate PECs or vice versa. In murine glomerular disease models of crescentic glomerulonephritis, and focal segmental glomerulosclerosis (FSGS), intermediate PECs became more numerous. These intermediate PECs preferentially expressed activation markers CD44 and Ki-67, suggesting that this subgroup of PECs was activated more easily than the classical flat PECs. In mice with FSGS, cuboidal and intermediate PECs formed sclerotic lesions. In patients with FSGS, cells forming the tip lesions expressed markers of intermediate PECs. These novel PEC subgroups form sclerotic lesions and were more prone to cellular activation compared to the classical flat PECs in disease. Thus, colonization of Bowman's capsule by cuboidal PECs may predispose to lesion formation and chronic kidney disease. We propose that tip lesions originate from this novel subgroup of PECs in patients with FSGS.Copyright © 2019 International Society of Nephrology

Background and Aim: Non-alcoholic steatohepatitis (NASH) can lead to cirrhosis and hepatocellular carcinoma. Currently, lifestyle modification is the only effective treatment. We have shown that human amnion epithelial cells (hAECs) reduce inflammation and fibrosis in toxin-induced liver injury models. We examined the effect of these cells and the soluble factors released by the cells into culture medium (hAEC conditioned medium [hAEC-CM]) in a diet-induced murine NASH model. Method(s): C57BL/6J male mice received a Western "fast food diet" for 42 weeks. Group 1 received an intraperitoneal injection of 2 x 106 hAECs at week 34, group 2 received an additional hAEC dose at week 38, and group 3 received thrice weekly hAEC-CM injections intraperitoneal for 8 weeks from week 34. Liver fibrosis area, inflammation, and fibrosis regulators were measured by immunohistochemistry, qPCR, and gelatin zymography. Metabolic parameters were also assessed. Result(s): Fast food diet-fed mice demonstrated peri-cellular hepatic fibrosis, inflammation, and steatosis typical of NASH. Liver fibrosis area was reduced by 40% in hAEC-treated and hAEC-CM-treated mice. hAEC treatment significantly reduced pSMAD 2/3 signaling and the number of activated hepatic stellate cells and liver macrophages. Matrix metalloproteinase 2 and 9 gene and protein expression were variably affected. hAEC treatment did not alter the NASH activity score or metabolic parameters such as bodyweight, total cholesterol, or glucose tolerance. Conclusion(s): Human amnion epithelial cell and hAEC-CM significantly reduced hepatic inflammation and fibrosis in a diet-induced non-alcoholic fatty liver disease model. Although hAEC and hAEC-CM did not affect the metabolic components of NASH, their therapeutic potential is promising and warrants further investigation.Copyright © 2019 Journal of Gastroenterology and Hepatology Foundation and John Wiley & Sons Australia, Ltd

Aim: Protease-activated receptor 2 (PAR2) has been implicated in the development of renal inflammation and fibrosis. In particular, activation of PAR2 in cultured tubular epithelial cells induces extracellular signal-regulated kinase signalling and secretion of fibronectin, C-C Motif Chemokine Ligand 2 (CCL2) and transforming growth factor-beta1 (TGF-beta1), suggesting a role in tubulointerstitial inflammation and fibrosis. We tested this hypothesis in unilateral ureteric obstruction (UUO) in which ongoing tubular epithelial cell damage drives tubulointerstitial inflammation and fibrosis. Method(s): Unilateral ureteric obstruction surgery was performed in groups (n = 9/10) of Par2-/- and wild type (WT) littermate mice which were killed 7 days later. Non-experimental mice were controls. Result(s): Wild type mice exhibited a 5-fold increase in Par2 messenger RNA (mRNA) levels in the UUO kidney. In situ hybridization localized Par2 mRNA expression to tubular epithelial cells in normal kidney, with a marked increase in Par2 mRNA expression by tubular cells, including damaged tubular cells, in WT UUO kidney. Tubular damage (tubular dilation, increased KIM-1 and decreased alpha-Klotho expression) and tubular signalling (extracellular signal-regulated kinase phosphorylation) seen in WT UUO were not altered in Par2-/- UUO. In addition, macrophage infiltration, up-regulation of M1 (NOS2) and M2 (CD206) macrophage markers, and up-regulation of pro-inflammatory molecules (tumour necrosis factor, CCL2, interleukin-36alpha) in WT UUO kidney were unchanged in Par2-/- UUO. Finally, the accumulation of alpha-SMA+ myofibroblasts, deposition of collagen IV and expression of pro-fibrotic factors (CTGF, TGF-beta1) were not different between WT and Par2-/- UUO mice. Conclusion(s): Protease-activated receptor 2 expression is substantially up-regulated in tubular epithelial cells in the obstructed kidney, but this does not contribute to the development of tubular damage, renal inflammatio

Chronic kidney disease can be understood as a pathological reduction in the number of functional glomeruli. It is a frequent medical problem and one of the major independent risk factors for cardiovascular morbidity and mortality. In humans, glomeruli/nephrons are generated during the prenatal period (glomerular endowment), which may be impaired by multiple conditions. After birth, glomeruli are progressively lost - mostly due to glomerular scarring (focal segmental glomerulosclerosis; FSGS). Multiple independent studies have shown that significant loss of glomerular visceral epithelial cells (podocytes) is sufficient to induce FSGS. It is generally believed that podocytes cannot renew themselves and it has been generally assumed that their number is determined at birth (podocyte endowment). However, there are several lines of experimental evidence showing that podocytes can be replenished in the postnatal period. First, a limited reserve of podocytes has been reported on Bowman's capsule, which may be associated with body growth and increases in glomerular size between childhood and adulthood. Second, two intrinsic progenitor cell niches have been proposed to replenish podocytes throughout adult life and in association with glomerular injury and podocyte loss: parietal epithelial cells and/or cells of the renin lineage. While there is increasing evidence supporting postnatal podocyte gain, controversy remains about the involved signalling pathways and the efficiency of these sources to prevent nephron loss.Copyright © 2018 Elsevier Ltd

Objective: Aspergillus fumigatus is the main causative agent of aspergillosis. Most infections occur in immunocompromised individuals, indicating an efficient clearance of conidia by the pulmonary defence system in immunocompetent individuals. Infections by other aspergilli like Aspergillus niger can occur, but to lesser extent. Previous studies showed that A. fumigatus and A. niger behave differently in the presence of type II alveolar A549 epithelial cells. A. fumigatus is more efficiently internalized by the A549 cells and shows a delay in germination, when compared to A. niger. The hyphae of A. fumigatus, that escaped the epithelial cells grow parallel to the epithelium, while the A. niger hyphae grow away from the epithelial cell layer. This study focusses on the gene expression of A. fumigatus and A. niger after co-cultivation with A549 cells. Our hypothesis is that the difference in lifestyle between the two aspergilli is also observed in the gene expression profiles. Methods: RNA of the co-cultivation of the A549 cells with A. fumigatus or A. niger was isolated and sequenced. The obtained RNA sequences were analysed with custom R and python scripts to obtain the differentially expressed genes and GO terms. Results: The obtained RNA sequences show big differences in the global gene expression of A. fumigatus and A. niger upon contact with A549 cells. A total of 545 and 473 genes for respectively A. fumigatus and A. niger were differently expressed when compared to growth in absence of A549 cells. Of these genes only 53 (∼10%) were shared between both species. The different response was also illustrated by the fact that only 4 GO terms were shared between the differentially expressed genes of both gene sets. Genes described in hypoxia regulation and heat shock were found up-regulated in A. fumigatus and their homologs in A. niger. The A. fumigatus thioredoxin reductase and allergen genes were found up-regulated in this fungus, but homologous genes were down-re

Background: Bronchopulmonary dysplasia (BPD) is a relatively common morbidity in preterm infants, especially in those born before 28 weeks of gestation. Current management of BPD, which has a significant impact on the patient, family and community, is largely supportive and no cure exists. Over the last decade, our group has shown that human amnion epithelial cells (hAECs), derived from healthy amniotic membranes can prevent and reverse lung injury in small and large animal models of adult and neonatal lung disease, including BPD. Aim(s): To assess the safety of intravenously administered allogeneic hAECs in preterm infants with established BPD. Method(s): Preterm infants born before 28 weeks of gestation were eligible if they were still significantly dependent on ventilatory support and oxygen at 36 weeks postconceptional age. Primary safety outcome parameters included short term measures, i.e. local reaction, anaphylaxis, infection, systemic rejection effects, and long term measures, i.e. tumourigenesis. cGMP-compliant hAECs were isolated from healthy term placenta donors and prepared at the Monash Health Cell Therapy Platform. The allogeneic hAECs were delivered to infants at Monash Newborn neonatal intensive care unit via a slow intravenous infusion at a dose of 1 million/ kg suspended in saline. Secondary outcomes included changes in respiratory function, other morbidities and death. Result(s): Six preterm infants (median birth weight 814 (450- 990) grams, gestation 27 (25-28) weeks) with established severe BPD were recruited into the trial. Three were intubated and three on continuous positive airway pressure (CPAP). There were no immediate or short term adverse effects associated with cell infusion. Respiratory support parameters in the first few days after cell infusion either improved or showed no significant change from pre-treatment levels. Long term results (two years post-infusion) of the first recruit indicate that the allogeneic hAECs are well-tolerat

STUDY QUESTION: Do obese levels of advanced glycation end products (AGEs) within the uterine cavity detrimentally alter tissue function in embryo implantation and placental development? SUMMARY ANSWER: Obese levels of AGEs activate inflammatory signaling (p65 NF?B) within endometrial epithelial cells and alter their function, cause endoplasmic reticulum (ER) stress in endometrial stromal cells and impair decidualization, compromise implantation of blastocyst mimics and inhibit trophoblast invasion. WHAT IS KNOWN ALREADY: Obese women experience a higher incidence of infertility, recurrent miscarriage and pregnancy complications compared with lean women. Oocyte donation cycles suggest a detrimental uterine environment plays a role in these outcomes. STUDY DESIGN, SIZE, DURATION: Uterine lavage and tissues from lean (BMI 19.524.9, n = 17) and obese (BMI > 30, n = 16) women examined. Cell culture experiments utilizing human endometrial epithelial, trophectoderm and trophoblast cell lines and primary human stromal cells used to examine the functional impact of obese levels of AGEs. PARTICIPANTS/MATERIALS, SETTING, METHODS: Levels of AGEs examined within uterine lavage assessed by ELISA to determine differences between lean and obese women. Expression and localization of AGEs, receptor for AGEs (RAGE) and NF?B within endometrial tissues obtained from lean and obese women determined by immunohistochemistry. Endometrial epithelial cells (ECC-1), primary human stromal cells and trophoblast cells (HTR8-SVneo) treated with lean (2000 nmol/mol lysine) or obese (8000 nmol/mol lysine) uterine levels of AGEs and p65 NF?B (western immunoblot), real-time adhesion, proliferation migration and invasion (xCelligence real-time cell function analysis), decidualization (cell morphology and prolactin release), ER stress (western immunoblot for p-PERK) determined. Co-cultures of endometrial epithelial cells and blastocyst mimics (trophectoderm spheroids) similarly treated with lean or obese

Radiographic contrast agents cause acute kidney injury (AKI), yet the underlying pathogenesis is poorly understood. Nod-like receptor pyrin containing 3-deficient (Nlrp3-deficient) mice displayed reduced epithelial cell injury and inflammation in the kidney in a model of contrast-induced AKI (CI-AKI). Unexpectedly, contrast agents directly induced tubular epithelial cell death in vitro that was not dependent on Nlrp3. Rather, contrast agents activated the canonical Nlrp3 inflammasome in macrophages. Intravital microscopy revealed diatrizoate (DTA) uptake within minutes in perivascular CX3CR1+ resident phagocytes in the kidney. Following rapid filtration into the tubular luminal space, DTA was reabsorbed and concentrated in tubular epithelial cells via the brush border enzyme dipeptidase-1 in volume-depleted but not euvolemic mice. LysM-GFP+ macrophages recruited to the kidney interstitial space ingested contrast material transported from the urine via direct interactions with tubules. CI-AKI was dependent on resident renal phagocytes, IL-1, leukocyte recruitment, and dipeptidase-1. Levels of the inflammasome-related urinary biomarkers IL-18 and caspase-1 were increased immediately following contrast administration in patients undergoing coronary angiography, consistent with the acute renal effects observed in mice. Taken together, these data show that CI-AKI is a multistep process that involves immune surveillance by resident and infiltrating renal phagocytes, Nlrp3-dependent inflammation, and the tubular reabsorption of contrast via dipeptidase-1.Copyright © 2018 American Society for Clinical Investigation. All rights reserved.

Aim: To determine whether Protease Activated-Receptor-2 (PAR-2) contributes to tubular cell activation and renal fibrosis in the obstructed kidney. Background(s): Activation of PAR2 has been implicated in the development of renal inflammation and fibrosis. In particular, activation of PAR-2 in cultured tubular epithelial cells induces cell proliferation and secretion of fibronectin, CCL2 and TGF-beta1; suggesting that PAR-2 signalling may promote tubulointerstitial inflammation and fibrosis. We tested this hypothesis in the unilateral ureteric obstruction (UUO) model in which activation of tubular epithelial cells drives tubulointerstitial inflammation and fibrosis. Method(s): UUO surgery was performed in groups (n=10) of Par2-/- and WT (littermate control) C57BL6/J mice which were killed 7 days later. Result(s): WT mice exhibited a 5-fold increase in PAR-2 mRNA levels in the UUO kidney. In situ hybridisation localised PAR-2 mRNA expression to tubular epithelial cells in normal kidney, with a marked increase in PAR-2 mRNA expression by tubular cells, including damaged tubular cells, in WT UUO kidney. Renal fibrosis was not different between Par2-/- and WT UUO in terms of collagen IV deposition and accumulation of alpha-SMA+ myofibroblasts, while increased mRNA levels for col I, col IV, alpha-SMA, CTGF and TGF-beta1 were also not different. Similarly, no difference in up-regulation of macrophage markers (CD68 and CD206) or inflammation markers (TNF-alpha, NOS-2, MCP- 1, IL-36alpha) was evident. Tubular damage was equivalent between Par2-/- and WT mice based on the percentage of damaged tubules on PAS stained sections, increased levels of KIM-1 mRNA and decreased levels of alpha-Klotho mRNA. Conclusion(s): PAR-2 expression is substantially up-regulated in tubular epithelial cells in the obstructed kidney, but this does not contribute to the development of renal inflammation or fibrosis.

Background and aim: Chronic liver disease is characterised by progressive hepatocyte injury, inflammation and accumulation of extracellularmatrix. Treatment for end stage liver disease is limited to liver transplantation. Liver progenitor cells (LPC) are bipotent cells induced during liver injury that can regenerate both hepatocytes and cholangiocytes. In addition, macrophages play a pivotal role in fibrogenesis and fibrosis resolution. Classically, macrophages are classified into M1 (pro-inflammatory macrophages) and M2 (proresolution macrophages). Recently, Ly6C expression has been used to identify distinct macrophage populations; Ly6Chi (inflammatory) and Ly6Clow (resolution). We are developing a therapy for chronic liver fibrosis using human amnion epithelial cells (hAECs). The cholinedeficient ethionine- supplemented (CDE) diet is a non-invasive and quick method to induce liver injury and LPC proliferation. Here, we aimed to investigate the effect of hAECs on macrophages and LPC during liver injury induced by CDE diet. Method(s): Male C57Bl/6J mice (6-8-weeks)were fed a CDE diet for up to 42 days; 2 or 4 million hAECs were infused intra-peritoneally 24h following initiation of the CDE diet. Outcomes were assessed at days 1, 3, 5, 14, 21 and 42 of the CDE diet following hAEC administration. The extent of liver fibrosis was determined by Sirius red staining and computer assisted morphometry was used to quantify the percentage of positively stained liver (expressed as % fibrosis area). LPC activation was determined by staining with wide spectrum screening cytokeratin (Pan-CK). Macrophages were further identified and characterised using flow cytometery sorting and PCR. Monocyte-derived macrophages (MDM) were distinguished from resident hepatic macrophages (Kupffer cells) based on expression CD45+ CD11bHi F4\80intermediate Ly6G- CD3- B220- NK1.1-. Further, MDM were divided into two subpopulations based on high and low Ly6C expression. Result(s): In CDE treated mice

Background: Bronchopulmonary dysplasia (BPD) is a common condition in extreme premature infants. Current management of BPD is largely supportive and no cure exists. Over the last decade, our group has shown that human amnion epithelial cells (hAECs), derived from healthy amniotic membranes can prevent and reverse lung injury in small and large animal models of adult and neonatal lung disease, including BPD. Aim(s): To assess the safety of intravenously administered allogeneic hAECs in preterm infants with established BPD. Method(s): Preterm infants born before 28 weeks of gestation were eligible if they were dependent on pressure support and oxygen at 36 weeks postconceptional age. Primary safety outcome parameters included short term effects - local reaction, anaphylaxis, infection, systemic rejection, and long term effects, including tumorigenesis. CGMP-compliant hAECs were isolated from healthy term placenta donors and delivered to infants using a slow intravenous infusion at a dose of 1 million/ kg suspended in saline. Secondary outcomes included changes in respiratory function. Result(s): Six preterm infants (median birth weight 795 (450- 990) grams, gestation 26 (24-28) weeks) with established severe BPD were administered hAECs at 89 (59-187) days of life. There were no immediate or short term adverse effects following hAEC transfusion. Respiratory support parameters in the first few days after cell infusion either improved or showed no significant change from pre-treatment levels. Conclusion(s): hAECs seem to be well tolerated when given to preterm infants with established severe BPD. This first-in-human study will inform future clinical trials of this novel therapy for BPD.

Background: Patients with inflammatory bowel disease (IBD) experience diarrhoea, bleeding, abdominal pain and weight loss. The chronic nature of IBD requires lifelong immunosuppressive therapy with numerous associated side effects. Patients may become refractory to conventional therapy, necessitating surgical intervention. Research has focused on stem cells as a disease modifying and regenerative option. The pluripotent human amnion epithelial cell (hAEC) is an excellent candidate due to its plentiful and ethical supply from placentas, anti-inflammatory properties and no known safety issues. We examined the efficacy of hAECs in a 2,4,6-trinitrobenzene sulfonic acid (TNBS) model of acute colitis. Method(s): For pre-sensitisation of BALB/c mice, a 1.5 x 1.5 cm area of skin was shaved for administration of 1% (w/v) 2,4,6-trinitrobenzene sulfonic acid (TNBS) in an acetone olive oil emulsion. Control mice received the same solution without TNBS. At Day 8, mice received a 150 mul enema of 2.5% TNBS (w/v) in 100% ethanol. Ethanol control mice received a 150 mul enema of absolute ethanol. At Day 9, TNBS-hAEC mice received an intravenous injection of 2 million hAECs. Control TNBS mice were untreated. Mice were monitored using the disease activity index (DAI) and culled at Day 12. Colitis severity was determined histologically through haematoxylin and eosin staining of the colon. IL-1beta, TNF-alpha, IL-6, IFN-gamma, IL-17, IL-23 were measured by ELISA. Neutrophil infiltration was evaluated by myeloperoxidase (MPO) activity assay. Result(s): Compared with untreated TNBS mice, bodyweight loss was significantly lower in hAEC-treated mice (p < 0.0001) with an improved DAI (p < 0.0001). In addition, hAEC treatment also prevented colon shortening by 17% (p < 0.05) with a similar trend for colon weight (p = 0.058). Histological severity scores were significantly higher in untreated TNBS mice compared with hAEC treated TNBS mice (p < 0.05). Similarly, IL-1beta, TNF-alpha, IL-6, IFN

Background: Inflammatory bowel disease encompasses a group of conditions characterised by inflammation and ulceration of the colon resulting in diarrhoea, haematochezia and weight loss. This often requires hospitalisation, corticosteroids and long-term immunosuppressants. However, these treatments are associated with numerous side effects and a subset of patients remain refractory to treatment. Stem cells, such as the pluripotent human amnion epithelial cells (hAECs), have been explored as a potential therapy. Ethically sourced from placentas, hAECs possess anti-inflammatory properties and no known safety issues. We examined the efficacy of hAECs in a dextran sulfate sodium (DSS) induced murine model of acute colitis. Method(s): C57BL/6J mice received 2% DSS (w/v) in their normal drinking water for 7 days. At Day 2, Group 1 (n = 17) received a tail vein of 2 x 106 hAECs. At Day 4, Group 2 (n = 17) received an injection of 2 x 106 hAECs. DSS control mice (n = 17) were untreated. Mice were monitored daily and culled at Day 8. Colitis severity was scored histologically using H&E stained colon sections. Colonic neutrophil infiltration was determined by myeloperoxidase (MPO) activity assay. IL-1beta, TNF-alpha, IL-6, IFN-gamma and IL-10 were measured through enzyme-linked immunosorbent assays. Colon sections were stained with anti-F/480 antibody to identify macrophages. Result(s): Untreated DSS mice lost significantly more body weight (-17.4%) compared with hAEC treated mice (Group 1: -5.6% (p < 0.0001), Group 2: -8.8% (p < 0.0001)). Treatment prevented colon shortening in Group 1 (6.7 cm p < 0.001) and Group 2 (6.3 cm, p < 0.01) compared with controls (5.3 cm). Similarly, hAEC treated mice had significantly lower histological severity scores (Group 1: p < 0.0001, Group 2: p < 0.001). MPO activity was almost halved in Groups 1 (p < 0.0001) and 2 (p < 0.0001). Treatment also significantly reduced concentrations of pro-inflammatory cytokines (IL-1beta, TNF-alpha, IL-6, IFN

Background and Purpose - Human amnion epithelial cells (hAECs) are nonimmunogenic, nontumorigenic, antiinflammatory cells normally discarded with placental tissue. We reasoned that their profile of biological features, wide availability, and the lack of ethical barriers to their use could make these cells useful as a therapy in ischemic stroke. Methods - We tested the efficacy of acute (1.5 hours) or delayed (1-3 days) poststroke intravenous injection of hAECs in 4 established animal models of cerebral ischemia. Animals included young (7-14 weeks) and aged mice (20-22 months) of both sexes, as well as adult marmosets of either sex. Results - We found that hAECs administered 1.5 hours after stroke in mice migrated to the ischemic brain via a CXC chemokine receptor type 4-dependent mechanism and reduced brain inflammation, infarct development, and functional deficits. Furthermore, if hAECs administration was delayed until 1 or 3 days poststroke, long-term functional recovery was still augmented in young and aged mice of both sexes. We also showed proof-of-principle evidence in marmosets that acute intravenous injection of hAECs prevented infarct development from day 1 to day 10 after stroke. Conclusions - Systemic poststroke administration of hAECs elicits marked neuroprotection and facilitates mechanisms of repair and recovery.Copyright © 2018 American Heart Association, Inc.

Background: Inflammatory bowel disease (IBD) requires lifelong therapy to control symptoms. Despite therapeutic advances, some patients experience debilitating side effects or are refractory to conventional treatment. Current treatments do not reduce or reverse established intestinal fibrosis. Isolated from placentas, human amnion epithelial cells (hAECs) are pluripotent stem cells that possess anti-inflammatory and anti-fibrotic properties. With no tumourogenicity and an excellent safety profile, hAECs are a promising novel therapy for IBD. We examined the efficacy of hAECs and its conditioned media (CM) in a dextran sulphate sodium (DSS) induced murine model of chronic colitis. Method(s): C57BL/6J mice received a cycle of 2% DSS (w/v) in normal water for one week then normal water for two weeks. This threeweek cycle was repeated twice until cull at week 10. Beginning at week 6, the Chronic DSS group administered hAECs (Chr-hAEC, n = 14) received weekly tail vein injections of 2million hAECs; the Chronic DSS group administered CM (Chr-CM, n = 15) group received 250uL CM three times weekly. Normal mice (n = 7) and DSS control mice (Chr-DSS, n = 13) received no therapy. Mice were monitored daily. Colitis severity was scored using H&E stained colon sections. IL-1beta, TNF-alpha, IL-6, TGF-beta, IFN-gamma, IL-10 were measured by ELISA. Fibrosis extent was measured via computer-assisted morphometry of picro-sirius stained colon sections. Fibroblasts and macrophages were identified by immunohistochemistry (alpha-SMA, F4/80). Result(s): Untreated control DSS mice gained significantly less body weight (4.2%) compared with hAEC (10%, p < 0.05) and CM (12.5%, p < 0.001) treated mice. hAECs and CM prevented colon shortening (6.75 cm, p < 0.001 and 7.3 cm, p < 0.0001 respectively) compared with controls (5.9cm). Splenomegaly was greater in Chr-DSS (212 mg) than hAEC (160 mg, p < 0.05) and CM (134 mg, p < 0.001) treated mice. Compared with controls, hAEC (p < 0.0001) and CM tre

Background: Sellar arachnoid cysts are a rare occurrence but may impinge on vital parasellar anatomy and thus are often symptomatic. The etiology of sellar arachnoid cysts is contentious, fueled by heterogeneity in cyst wall structure and contents between cases. The "ball-valve" mechanism is 1 of 2 predominant theories describing their formation, which contends that an aperture in the diaphragm allows cerebrospinal fluid to enter the cyst, propelled by pulsatile flow, but its egress is obscured by the pituitary during the ebb of the pressure wave. Case Description: Here we present a case of a 51-year-old female with a symptomatic sellar arachnoid cyst. She underwent an endoscopic transsphenoidal fenestration which alleviated her symptoms. Conclusion(s): Intraoperative video evidence during arachnoid cyst fenestration supports the "ball-valve" theory of sellar arachnoid cyst development.Copyright © 2018 Elsevier Inc.

The std locus of Salmonella enterica, an operon acquired by horizontal transfer, encodes fimbriae that permit adhesion to epithelial cells in the large intestine. Expression of the std operon is bistable, yielding a major subpopulation of StdOFF cells (99.7%) and a minor subpopulation of StdON cells (0.3%). In addition to fimbrial proteins, the std operon encodes two proteins, StdE and StdF, that have DNA binding capacity and control transcription of loci involved in flagellar synthesis, chemotaxis, virulence, conjugal transfer, biofilm formation, and other cellular functions. As a consequence of StdEF pleiotropic transcriptional control, StdON and StdOFF subpopulations may differ not only in the presence or absence of Std fimbriae but also in additional phenotypic traits. Separation of StdOFF and StdON lineages by cell sorting confirms the occurrence of lineage-specific features. Formation of StdOFF and StdON lineages may thus be viewed as a rudimentary bacterial differentiation program.