Epithelial–immune cell interactions in allergic diseases

Epithelial/immune interactions are characterized by the different properties of the various epithelial tissues, the mediators involved, and the varying immune cells that initiate, sustain, or abrogate allergic diseases on the surface. The intestinal mucosa, respiratory mucosa, and regular skin feature structural differences according to their primary function and surroundings. In the context of these specialized functions, the active role of the epithelium in shaping immune responses is increasingly recognizable. Crosstalk between epithelial and immune cells plays an important role in maintaining homeostatic conditions. While cells of the myeloid cell lineage, mainly macrophages, are the dominating immune cell population in the skin and the respiratory tract, lymphocytes comprise most intraepithelial immune cells in the intestine under healthy conditions. Common to all surface epithelia is the fact that innate immune cells represent the first line of immunosurveillance that either directly defeats invading pathogens or initiates and coordinates more effective successive immune responses involving adaptive immune cells and effector cells. Pharmacological approaches for the treatment of allergic and chronic inflammatory diseases involving epithelial barriers target immunological mediators downstream of the epithelium (such as IL-4, IL-5, IL-13, and IgE). The next generation of therapeutics involves upstream events of the inflammatory cascade, such as epithelial-derived alarmins and related mediators.     

γδ T Cells in Host Defense and Epithelial Cell Biology

Recent studies have demonstrated increased numbers of γδ T cells in a variety of human infectious as well as noninfectious diseases. In some cases γδ T cells could be shown to destroy infected or transformed cells. Advances in the identification of ligands recognized by γδ T cells and the development of animal model systems to study these cellsin vivoshould overcome some of the major obstacles currently preventing a better understanding of γδ T cell function in immune responses. As we gain this knowledge it may become possible to design therapeutic strategies exploiting unique properties of γδ T cells to promote more effective immunity.     

Myoid Gonadal Stromal Tumor with Epithelial Differentiation (? Testicular Myoepithelioma)

A 46-year-old man presented with a cytologically bland testicular tumor composed of spindle cells that showed both epitheliallike (ie, true desmosomes and tonofilamentlike structures) and myogenous differentiation (ie, thin filaments with focal densities and α-smooth muscle actin immunoreac-tivity). Tumor cells were immunoreactive for vimen-tin and S-100 protein but negative for cytokeratin and desmin. Peritubular myoid cells are present in the normal testis; contain subplasmalemmal micro-pinocytotic vesicles; show thin filaments with focal densities; and are reactive with desmin, vimentin, and α-smooth muscle actin. They have no desmosomes and lie outside the basement membrane of the seminiferous tubules; thus they are not true myoepithelial cells (a cell type not present in the testis). Paradoxically, the current tumor appeared to show bidirectional differentiation, mimicking both a peritubular myoid spindle cell and an epitheliallike cell (possibly similar to the granulosa cell or rete testis epithelial cell). Although the findings suggest myoepithelial differentiation, the cytogenesis of this tumor remains uncertain.     

Dermal Transforming Growth Factor-β Responsiveness Mediates Wound Contraction and Epithelial Closure

Stromal-epithelial interactions are important during wound healing. Transforming growth factor-β (TGF-β) signaling at the wound site has been implicated in re-epithelization, inflammatory infiltration, wound contraction, and extracellular matrix deposition and remodeling. Ultimately, TGF-β is central to dermal scarring. Because scarless embryonic wounds are associated with the lack of dermal TGF-β signaling, we studied the role of TGF-β signaling specifically in dermal fibroblasts through the development of a novel, inducible, conditional, and fibroblastic TGF-β type II receptor knockout (Tgfbr2^dermalKO) mouse model. Full thickness excisional wounds were studied in control and Tgfbr2^dermalKO back skin. The Tgfbr2^dermalKO wounds had accelerated re-epithelization and closure compared with controls, resurfacing within 4 days of healing. The loss of TGF-β signaling in the dermis resulted in reduced collagen deposition and remodeling associated with a reduced extent of wound contraction and elevated macrophage infiltration. Tgfbr2^dermalKO and control skin had similar numbers of myofibroblastic cells, suggesting that myofibroblastic differentiation was not responsible for reduced wound contraction. However, several mediators of cell-matrix interaction were reduced in the Tgfbr2^dermalKO fibroblasts, including α1, α2, and β1 integrins, and collagen gel contraction was diminished. There were associated deficiencies in actin cytoskeletal organization of vasodilator-stimulated phosphoprotein-containing lamellipodia. This study indicated that paracrine and autocrine TGF-β dermal signaling mechanisms mediate macrophage recruitment, re-epithelization, and wound contraction.Skin wound healing in adult humans after full-thickness injury results in scar tissue, which is prone to contracture, loss in elasticity, and altered tensile strength. The characteristics of a healed wound, unlike the original dermal tissue, include an extracellular matrix without the normal basket weave organization of collagen fibers that can be further exaggerated as a hypertrophic scar. In scars, there is a loss of normal tissue architecture and function due, at least in part, to altered cell-matrix interaction and the assembly of the actin cytoskeleton that occurs during wound repair.^1,2 The actin cytoskeleton is important in processes that are fundamental to wound healing: migration; contraction; adhesion; and proliferation.³ The vasodilator-stimulated phosphoprotein (VASP) is associated with filamentous actin formation and likely plays a role in cell adhesion and motility.⁴ VASP may also be involved in the intracellular signaling pathways that regulate integrin-extracellular matrix interactions.The importance of stromal-epithelial interactions in development and wound healing is well established. These interactions likely involve autocrine and paracrine action of multiple growth factors, including members of the transforming growth factor-β (TGF-β) family. TGF-β1, β2, and β3 isoforms signal by binding to the TGF-β receptors type I and type I at the cell surface. The ligand-receptor complex formed stimulates multiple parallel signal pathways in the cytoplasm. TGF-β is a growth inhibitor for many cell types and stimulates extracellular matrix expression in mesenchymal cells. In the injured skin, macrophages, endothelium, fibroblasts, and epithelia are all sources of elevated TGF-β expression. TGF-β signaling at the wound site is thought to be important for extracellular matrix deposition and remodeling. The application of TGF-β1 to wounds has been shown to accelerate wound healing.⁵ In apparent contradiction, a genetic knockout of the TGF-β effector, Smad3, in mice exhibited accelerated skin and lens wound healing.^6–8 However, the epithelial and stromal compartments can respond differentially to TGF-β signaling, and the specific mechanism of TGF-β action on any individual cell type within tissues is still not understood. Wounding of embryonic skin, in contrast to that in adults, is not associated with remodeling deficiencies associated with scarring. Interestingly, embryonic dermal fibroblasts lack expression of TGF-β receptor type II.^9,10In the present study, we specifically address the role of TGF-β signaling in dermal fibroblasts by the development of a novel dermal fibroblast-inducible, conditional, TGF-β type II receptor knockout (Tgfbr2^dermalKO) mouse model. The generation of Tgfbr2^dermalKO mice provided an opportunity to test the role of TGF-β in stromal cells in wound healing. The full-thickness skin excision wounds in Tgfbr2^dermalKO mice had accelerated re-epithelization compared with control, impaired collagen organization associated with a lack of wound contraction, and reduced macrophage recruitment. The studies indicate that paracrine and autocrine TGF-β dermal responsiveness mechanisms impact skin wound healing.     

Role of the Urokinase-Fibrinolytic System in Epithelial–Mesenchymal Transition during Lung Injury

Alveolar type II epithelial (ATII) cell injury precedes development of pulmonary fibrosis. Mice lacking urokinase-type plasminogen activator (uPA) are highly susceptible, whereas those deficient in plasminogen activator inhibitor (PAI-1) are resistant to lung injury and pulmonary fibrosis. Epithelial–mesenchymal transition (EMT) has been considered, at least in part, as a source of myofibroblast formation during fibrogenesis. However, the contribution of altered expression of major components of the uPA system on ATII cell EMT during lung injury is not well understood. To investigate whether changes in uPA and PAI-1 by ATII cells contribute to EMT, ATII cells from patients with idiopathic pulmonary fibrosis and chronic obstructive pulmonary disease, and mice with bleomycin-, transforming growth factor β–, or passive cigarette smoke–induced lung injury were analyzed for uPA, PAI-1, and EMT markers. We found reduced expression of E-cadherin and zona occludens-1, whereas collagen-I and α-smooth muscle actin were increased in ATII cells isolated from injured lungs. These changes were associated with a parallel increase in PAI-1 and reduced uPA expression. Further, inhibition of Src kinase activity using caveolin-1 scaffolding domain peptide suppressed bleomycin-, transforming growth factor β–, or passive cigarette smoke–induced EMT and restored uPA expression while suppressing PAI-1. These studies show that induction of PAI-1 and inhibition of uPA during fibrosing lung injury lead to EMT in ATII cells.Idiopathic pulmonary fibrosis (IPF) and other interstitial lung diseases are characterized by destruction of lung architecture due to excessive deposition of extracellular matrix proteins by activated fibroblasts or myofibroblasts, leading to progressive dyspnea and loss of lung function.^1–3 The origins of myofibroblasts participating in the pathological remodeling of IPF lungs are not clear. Histopathological evaluation demonstrates that myofibroblasts accumulate in fibroblastic foci. Emerging evidence suggests that polarized type II alveolar epithelial (ATII) cells undergo epithelial–mesenchymal transitions (EMT) after lung injury. The ATII cells assume phenotypic changes such as increased migration, invasion, resistance to apoptosis, and production of elevated levels of extracellular matrix proteins^4,5 and therefore serve as a source of myofibroblasts. Understanding the possible mechanisms contributing to EMT in ATII cells may help identify new targets to treat or at least limit fibrogenesis after lung injury.A number of molecular processes are involved in the initiation of EMT in ATII cells.⁵ Components of the fibrinolytic system such as urokinase-type plasminogen activator (uPA), uPA plasma membrane receptor (uPAR), and its major inhibitor, plasminogen activator inhibitor (PAI-1) are all elaborated by ATII cells. These proteins independently influence a broad range of biological processes germane to lung injury and its repair.⁶ However, their role in fibrogenesis via EMT is unclear. Recent publications using bleomycin (BLM)⁷ and a passive cigarette smoke (PCS)⁸ or adenovirus expressing constitutively active transforming growth factor β (Ad-TGF-β)^1,9 exposure model of lung injury indicate that a coordinate increase in PAI-1 and a decrement in uPA by ATII cells promote lung injury and subsequent pulmonary fibrosis (PF). We also found that caveolin-1 scaffolding domain peptide (CSP) acts as a competitor to caveolin-1, restores expression of uPA and uPAR, and inhibits PAI-1 in ATII cells after lung injury. These changes prevent development of PF after lung injury.⁷ Recent literature suggests that up to 30% to 50% of myofibroblasts may be derived via EMT during fibrogenesis.^10–12 However, an in vivo genetic lineage tracing study reported by Rock et al¹³ contradicts these findings. Our objective in the current study is to elucidate the role of altered expression of uPA, uPAR, and PAI-1 after lung injury in EMT, and further evaluate whether reinstatement of baseline expression of uPA, uPAR, and PAI-1 by CSP intervention after lung injury reduces EMT in ATII cells.     

Epithelial–mesenchymal transition (EMT) of renal tubular cells in canine glomerulonephritis

Tubulo-interstitial fibrosis in dogs may result from primary injury to the interstitium or develop secondary to other renal diseases. As in human renal pathology, tubular epithelial cells (TEC) are believed to actively participate in the mechanisms of renal fibrosis. In this study, we examined the changes in the tubular epithelial component in two specific canine diseases. Immunohistochemistry showed the expression of the epithelial marker cytokeratin, the smooth muscle marker α-SMA, the mesenchymal marker vimentin and PCNA in 20 dogs with membranous glomerulonephritis and membrano-proliferative glomerulonephritis. Results showed that the loss of the epithelial marker in TEC was directly correlated to the grade of tubulo-interstitial disease present and independent of the type of glomerulonephritis. Varying degrees of vimentin positivity were detected in tubular epithelium in areas of inflammation, and low numbers of scattered α-SMA-positive cells were also observed. Immunohistochemistry showed that epithelial tubular cells lose their cytokeratin staining characteristics and transdifferentiate into cells exhibiting key mesenchymal immunophenotypic feature of vimentin-positive staining in both diseases investigated. The integrity of the tubular basement membrane is likely to be fundamental in maintaining the epithelial phenotype of TEC. Animal models provide opportunities for investigating the pathogenesis of renal fibrosis in humans.     

Hertwig’s Epithelial Root Sheath Fate during Initial Cellular Cementogenesis in Rat Molars

To elucidate the fate of the epithelial root sheath during initial cellular cementogenesis, we examined developing maxillary first molars of rats by immunohistochemistry for keratin, vimentin, and tissue non-specific alkaline phosphatase (TNALP) and by TdT-mediated dUTP nick end labeling (TUNEL). The advancing root end was divided into three sections, which follow three distinct stages of initial cellular cementogenesis: section 1, where the epithelial sheath is intact; section 2, where the epithelial sheath becomes fragmented; and section 3, where initial cellular cementogenesis begins. After fragmentation of the epithelial sheath, many keratin-positive epithelial sheath cells were embedded in the rapidly growing cellular cementum. A few unembedded epithelial cells located on the cementum surface. Dental follicle cells, precementoblasts, and cementoblasts showed immunoreactivity for vimentin and TNALP. In all three sections, there were virtually no cells possessing double immunoreactivity for vimentin-keratin or TNALP-keratin and only embedded epithelial cells showed TUNEL reactivity. Taken together, these findings suggest that: (1) epithelial sheath cells divide into two groups; one group is embedded in the cementum and thereafter dies by apoptosis, and the other survives on the cementum surface as epithelial cell rests of Malassez; and (2) epithelial sheath cells do not undergo epithelial-mesenchymal transition during initial cellular cementogenesis.     

The Role of Epithelial Cells in the Pathogenesis of Sjögren’s Syndrome

Sjögren’s syndrome (SS) is a chronic autoimmune exocrinopathy that is characterized by periductal mononuclear cell infiltrates in the affected exocrine glands. Epithelial cells are thought to play an important pathogenetic role, as suggested by the occurrence of infiltrating lesions in various epithelial tissues (described as autoimmune epithelitis) as well as the increased epithelial expression of several inflammatory proteins in the histopathologic lesions of patients. In the recent decade, the application of long-term cultured nonneoplastic salivary gland epithelial cell (SGEC) lines has permitted the more explicit analysis of the role of these cells in SS pathophysiology. In such studies, cultured SGEC have been demonstrated to express constitutively or inducibly various molecules that are implicated in innate and acquired immune responses, a fact that underscores the inherent capacity of these epithelial cells to induce and promote chronic inflammatory reactions. Furthermore, the parallel analysis of SGEC lines obtained from SS patients and disease controls has revealed the significantly increased constitutive expression of several molecules in cells derived from SS patients. This fact strongly suggests the operation of intrinsic activation mechanisms in the epithelia of patients and further supports the active participation of epithelia in SS pathogenesis.     

A Randomized Study of Epithelial Ovarian Cancer: Is Chemotherapy Useful after Complete Remission?

Objective. The aim of this study is to verify whether consolidation chemotherapy with Cisplatin improves disease-free survival and/or overall survival in patients affected by epithelial ovarian cancer.Methods. A multicenter study examined 122 randomized patients in complete remission as judged by laparoscopy or laparotomy following first-line chemotherapy consisting of ACy (Adriamycin + Cyclophosphamide), PCy (Cisplatin + Cyclophosphamide), or Mitoxantrone + Carboplatin. Sixty-one of these patients were treated with 3 cycles of 5-Fluorouracil (FU) 500 mg/m2 for 5 days followed by Cisplatin at 100 mg/m2 on the 6th or 7th day every 28 days; the other 61 received no further treatment (nihil group).Results. Sixty patients in the Cisplatin arm were evaluable. There were 36 relapses in the FU+Cisplatin arm and 30 in the nihil arm. Peritoneal relapses were 25% for Cisplatin treatment vs. 16.4 % for nihil. There were 29 deaths in the Cisplatin arm vs. 27 for nihil. Median overall survival time (95 months with Cisplatin vs. 96 months in the nihil group) and median disease-free survival (66 months with Cisplatin vs. 73 in the nihil group) were similar in both arms (p=0.66 and p=0.41, respectively). There were no significant differences in tumor stage and grade between the two arms. Seven patients presented a second neoplasm during follow-up: six in the nihil arm, but only one patient in the Cisplatin arm. Death in these patients was due to the second neoplasm and not to progression of ovarian cancer.Conclusion. Three courses of additional platinum+FU treatment after five cycles of first-line chemotherapy without FU produced no increase in overall survival or disease-free survival.     

Ophiopogonin D Inhibiting Epithelial NF-κB Signaling Pathway Protects Against Experimental Colitis in Mice

Inflammation - The sustained activation of the nuclear factor κB (NF-κB) signaling pathway has been observed in human inflammatory bowel disease (IBD). Ophiopogonin D (OP-D) is a small...     

Glutamine Reduces TNF-α by Enhancing Glutathione Synthesis in Lipopolysaccharide-Stimulated Alveolar Epithelial Cells of Rats

To investigate the role of glutathione (GSH) synthesis in the regulation on nuclear factor (NF)-κB activity and tumor necrosis factor-alpha (TNF-α) release by glutamine (GLN) in lipopolysaccharide (LPS)-stimulated alveolar type II (AT-II) epithelial cells of rat lungs. Primary cultured AT-II cells were pre-treated with various doses of GLN for 2, 8, 16, 24 h. At the 8 h time point before LPS stimulation, various doses of l-buthionine-(S,R)-sulfoximine (BSO), an inhibitor of GSH synthesis, were added with 10 mM GLN. Then the cells were stimulated with 1 μg/ml LPS for 24 h. The cells were obtained for GSH measurement. TNF-α level in the supernatant was determined by enzyme-linked immunosorbent assay. NF-κB activity was assessed by electrophoretic mobility shift assay. Eight hours before LPS exposure was the best time point for GLN’s enhancing GSH synthesis. LPS could significantly decrease the GSH level, increase NF-κB activation and TNF-α release in AT-II cells. Supplementation of GLN could increase the GSH level and attenuate the release of TNF-α in LPS-stimulated AT-II cells in a dose-dependant manner. And NF-κB activation also could be prevented by GLN. BSO could block the effect of GLN. As a precursor of GSH, glutamine could prevent the NF-κB activation and attenuate the release of TNF-α in LPS-stimulated AT-II cells and the effect may be mediated via GSH synthesis.     

Effects of Laminins 332 and 411 on the Epithelial—Mesenchymal Status of Colorectal Cancer Cells

Bulletin of Experimental Biology and Medicine - The effects of laminins 332 and 411 (LM-332 and LM-411) on the epithelial—mesenchymal transformation of colorectal cancer cells (lines HT-29,...