Epidermal growth factor partially restores colonic ion transport responses in mouse models of chronic colitis

BACKGROUND & AIMS: Epidermal growth factor receptor (EGFR) activation, which plays an important role in regulating intestinal ion transport, can alleviate clinical symptoms such as diarrhea in patients with ulcerative colitis and promote mucosal restitution in animal models of colitis. Here, we investigate whether EGFR can regulate colonic ion transport in the setting of colitis. METHODS: Distal colon from control mice and mice with colitis was retained for immunohistochemistry or mounted in Ussing chambers. Ion transport responses across the tissues to the calcium agonist carbachol and the adenosine 3',5'-cyclic monophosphate agonist forskolin were measured with or without epidermal growth factor (EGF) pretreatment. RESULTS: EGF pretreatment of normal colonic mucosa inhibited ion transport responses to carbachol and forskolin but potentiated the reduced ion transport responses seen in dextran sulfate sodium (DSS)-treated and mdr1a knockout mouse colon. Ion substitution studies and the sodium transport inhibitor amiloride showed that sodium movement primarily accounted for the potentiating effect of EGF in DSS-treated tissues, despite decreased sodium channel expression. EGF potentiation of transport responses in DSS-treated colon was completely blocked by the cytoskeletal disruptor cytochalasin D and the phosphatidylinositol 3-kinase inhibitor wortmannin, whereas the novel and conventional protein kinase C isoform inhibitor G?6850 and the extracellular signal-regulated kinase inhibitor PD98059 partially reduced EGF effects. EGFR epithelial distribution and transforming growth factor alpha expression were also altered in DSS-treated tissues. CONCLUSIONS: Chronic inflammation uncovers a potentiating effect of EGFR activation on epithelial electrogenic sodium absorption that would be expected to ameliorate diarrheal symptoms associated with colitis.     

Loss of cooperative function of transforming growth factor-beta signaling proteins, smad3 with embryonic liver fodrin, a beta-spectrin, in primary biliary cirrhosis.

Modulation of fibrogenesis, epithelial, and mesenchymal cell fates are prominent effects of transforming growth factor-beta (TGF-beta) signaling by Smad proteins. We have previously shown that Smad2 and Smad3 insufficiency leads to a loss of bile ducts. In addition, Smad3/4 activity is mediated by embryonic liver fodrin (ELF), a beta-Spectrin. In mouse elf(-/-) mutants and in liver explant cultures, loss of ELF function results in T lymphocytic proliferation and absent intrahepatic bile ducts. A similar phenotype is seen in a number of cholestatic diseases with progressive loss of intrahepatic bile ducts and fibrosis. However, the expression patterns of Smads or role of ELF in cholestatic and fibrotic liver diseases are not yet known. METHODS/RESULTS: We investigated the role of ELF in primary biliary cirrhosis (PBC), autoimmune hepatitis C, chronic viral hepatitis and in livers from mice deficient in Smad2/Smad3. We generated elf(+/-) mutant mice and analyzed for chronic liver disease and hepatocellular cancer (HCC) from 6 to 12 months. Perturbations in ELF expression were consistently seen only in PBC tissues. ELF expression was similarly aberrant in tissues from Smad2(+/-)/Smad3(+/-) mutant mice. Further studies indicated that ELF mislocalization is correlated with aberrant localization of Smad3 in some PBC tissues. Thirteen of 17 elf(+/-) mutant mice developed steatosis, fibrosis, hepatic dysplasia, with HCC in two mice. CONCLUSIONS: These results suggest that a compromised cytoarchitecture and polarized trafficking of TGF-beta signaling molecules, ELF and Smad3 are involved in the pathogenesis of PBC as well as HCC.     

Activated microglia in a rat stroke model express NG2 proteoglycan in peri‐infarct tissue through the involvement of TGF‐β1

We investigated activated microglia in ischemic brain lesions from rats that had been subjected to transient middle cerebral artery occlusion. Activated microglia expressing NG2 chondroitin sulfate proteoglycan (NG2) were found only in the narrow zone (demarcation zone) that demarcated the peri‐infarct tissue and ischemic core. NG2^? activated microglia were abundantly distributed in the peri‐infarct tissue outside the demarcation zone. NG2⁺ microglia but not NG2^? microglia expressed both CD68 and a triggering receptor expressed on myeloid cells 2 (TREM‐2), suggesting that NG2⁺ microglia eliminated apoptotic neurons. In fact, NG2⁺ microglia often attached to degenerating neurons and sometimes internalized NeuN⁺ or neurofilament protein⁺ material. Kinetic studies using quantitative real‐time RT‐PCR revealed that expression of transforming growth factor‐β1 (TGF‐β1) was most evident in the ischemic core; with this marker produced mainly by macrophages located in this region. TGF‐β receptor mRNA expression peaked at 3 days post reperfusion (dpr) in the peri‐infarct tissue, including the demarcation zone. Primary cultured rat microglia also expressed the receptor mRNA. In response to TGF‐β1, primary microglia enhanced the expression of NG2 protein and TREM‐2 mRNA as well as migratory activity. A TGF‐β1 inhibitor, SB525334, abolished these effects. The present results suggest that TGF‐β1 produced in the ischemic core diffused toward the peri‐infarct tissue, driving activated microglial cells to eliminate degenerating neurons. Appropriate control of NG2⁺ microglia in the demarcation zone might be a novel target for the suppression of secondary neurodegeneration in the peri‐infarct tissue. GLIA 2014;62:185–198     

Immunohistochemical study of tubular epithelial cells and vascular endothelial cells in glomerulonephritis

Background and aims: In order to assess the role played by tubular epithelial cells (TEC) and interstitial vascular endothelial cells (VEC) in interstitial fibrogenesis in human glomerulonephritis, we studied the expression of markers of activated fibroblasts (α-smooth muscle actin (αSMA) and vimentin (Vim)) and of the transforming growth factor β (TGFβ), at the level of these cells. Methods: We studied retrospectively 41 renal biopsies from patients with primary and secondary glomerulonephritis [24 males, 17 females, mean age 45.5?±?12.9?years]. Immunohistochemistry using monoclonal antibodies (SMA, Vim, TGFβ) was assessed using a semiquantitative score, that was correlated with biological and histological data (quantified using a scoring system in order to assess active-inflammatory and chronic–sclerotic/fibrotic lesions). Results: The presence of SMA and Vim as markers of myofibroblasts was found in TECs and VECs. TEC Vim expression correlated with interstitial Vim expression (r?=?0.38; p?=?0.008), interstitial infiltrate (r?=?0.31; p?=?0.027), interstitial fibrosis (R?=?0.25; p?=?0.042), GFR (r?=??0.35; p?=?0.016), SMA (r?=??0.42; p?=?0.015), TGFβ (r?=?0.25; p?=?0.046), and hemoglobin (r?=??0.55; p?r?=??0.32; p?=?0.023) and interstitial fibrosis (r?=??0.34; p?=?0.017). Conclusion: Our study reflects the complexity of the involvement of VEC and mainly of TEC in fibrosis. The expression of mesenchymal markers at the tubular cell level (especially Vim) correlates with histological interstitial changes, with the decrease of renal function and more strongly with anemia.     

New advances in the pathophysiologic and radiologic basis of the exstrophy spectrum

The exstrophy–epispadias complex is a rare spectrum of anomalies affecting the genitourinary system, anterior abdominal wall, and pelvis. Recent advances in the repair of classic bladder exstrophy (CBE) and cloacal exstrophy (CE) have resulted in significant changes in outcomes of surgical management (including higher continence rate, fewer surgical complications, and better cosmesis) and health-related quality of life in these patients. These noteworthy changes resulted from advances in the pathophysiological and genetic backgrounds of this disease and better radiologic assessment of the three-dimensional anatomy of the bony pelvis and its musculature. A PubMed search was performed with the keyword exstrophy. The resulting literature pertaining to genetics, stem cells, imaging, tissue engineering, epidemiology, and endocrinology was reviewed. The following represents an overview of the advances in basic science understanding and imaging of the exstrophy–epispadias spectrum and discusses their possible and future effects on the management of CBE and CE.