- Disruptive Influences on Research in Academic Pathology Departments: Proposed Changes to the Common Rule Governing Informed Consent for Research Use of Biospecimens and to Rules Governing Return of Research Results. [Review]
- AJAm J Pathol 2016 Nov 30
- Academic pathology departments will be dramatically affected by proposed United States federal government regulatory initiatives. Pathology research will be substantially altered if proposed changes ...
Academic pathology departments will be dramatically affected by proposed United States federal government regulatory initiatives. Pathology research will be substantially altered if proposed changes to the Common Rule (Code of Federal Regulations: Protection of Human Subjects title 45 CFR 46) and regulations governing the return of individual research results are approved and finalized, even more so now that the Precision Medicine initiative has been launched. Together, these changes are disruptive influences on academic pathology research as we know it, straining limited resources and compromising advances in diagnostic and academic pathology. Academic research pathologists will be challenged over the coming years and must demonstrate leadership to ensure the continued availability of and the ethical use of research pathology specimens.
- Mouse Double Minute 2 Homolog Actively Suppresses p53 Activity in Oocytes during Mouse Folliculogenesis. [Journal Article]
- AJAm J Pathol 2016 Nov 29
- The p53 signaling network is indispensible in cellular stress responses and tumor suppression. Negative regulations of p53 by mouse double minute 2 and 4 homologs (MDM2) and (MDM4) are an integrated ...
The p53 signaling network is indispensible in cellular stress responses and tumor suppression. Negative regulations of p53 by mouse double minute 2 and 4 homologs (MDM2) and (MDM4) are an integrated component of the network and have been implicated in regulating the stress responses and the maintenance of normal development and homeostasis of multiple somatic cell lineages. However, the regulatory role of MDM2 on p53 and stress responses in female germ cells remains undetermined. Here, we used the Cre-loxP system to delete Mdm2 in oocytes at different stages of folliculogenesis in mice. Mdm2 deletion resulted in a clear p53 nuclear accumulation in the oocytes and impeded fertilities with early follicular loss in mice, resembling human premature ovarian failure phenotypes. These phenotypes were fully rescued by concurrent deletion of p53 in mice. In addition, Nutlin-3, a small molecule compound that inhibited the binding of MDM2 to p53, also promoted p53-dependent oocyte death. Although cancer therapeutic agents fluorouracil and doxorubicin could not induce a robust p53 activation in the wild-type oocytes, they induced p53 nuclear accumulation in the Mdm2 and Mdm4 double heterozygous oocytes. These results demonstrated a critical prosurvival role for MDM2 in the oocytes. Moreover, they suggested a more tightened and rigorous regulatory mode for the MDM2/MDM4-p53 network in female germ cells under stress situations.
- Nlrp3 Activation Induces Il-18 Synthesis and Affects the Epithelial Barrier Function in Reactive Cholangiocytes. [Journal Article]
- AJAm J Pathol 2016 Nov 29
- Microbial products are thought to influence the progression of cholangiopathies, in particular primary sclerosing cholangitis (PSC). Inflammasomes are molecular platforms that respond to microbial pr...
Microbial products are thought to influence the progression of cholangiopathies, in particular primary sclerosing cholangitis (PSC). Inflammasomes are molecular platforms that respond to microbial products through the synthesis of proinflammatory cytokines. We investigated the role of inflammasome activation in cholangiocyte response to injury. Nucleotide-binding oligomerization domain (NOD)-like receptor family, pyrin domain-containing protein 3 (Nlrp3) expression was tested in cholangiocytes of normal and cholestatic livers. Effects of Nlrp3 activation induced by incubation with lipopolysaccharide and ATP was studied in vitro in normal and siRNA-Nlrp3 knockdown cholangiocytes. Wild-type and Nlrp3 knockout (Nlrp3(-/-)) mice were fed 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC; a model of sclerosing cholangitis) for 4 weeks. Nlrp3 and its components were overexpressed in cholangiocytes of mice subjected to DDC and in patients affected by PSC. In vitro, Nlrp3 activation stimulated expression of Il-18 but not of Il-1β and Il-6. Nlrp3 activation had no effect on cholangiocyte proliferation but significantly decreased the expression of Zonulin-1 and E-cadherin, whereas Nlrp3 knockdown increased the permeability of cholangiocyte monolayers. In vivo, the DDC-stimulated number of cytokeratin-19-positive cells in the liver of wild-type animals was slightly reduced in Nlrp3(-/-) mice, and expression of E-cadherin was reestablished. In conclusion, Nlrp3 is expressed in reactive cholangiocytes, in both murine models and patients with PSC. Activation of Nlrp3 leads to synthesis of proinflammatory cytokines and influences epithelial integrity of cholangiocytes.
- Activating Transcription Factor 3 Is Essential for Cigarette Smoke-Induced Mucin Expression via Interaction with Activator Protein-1. [Journal Article]
- AJAm J Pathol 2016 Nov 29
- Mucus hypersecretion is an important pathologic feature of chronic obstructive pulmonary disease. Activating transcription factor 3 (ATF3) is an adaptive-response gene that participates in various ce...
Mucus hypersecretion is an important pathologic feature of chronic obstructive pulmonary disease. Activating transcription factor 3 (ATF3) is an adaptive-response gene that participates in various cellular processes. However, little is known about its role in cigarette smoke (CS)-induced mucus hyperproduction. This study aimed to investigate the role and molecular mechanisms of ATF3 in CS-induced Mucin 5AC (MUC5AC) expression. ATF3 was elevated in lung tissues of mice exposed to CS for 12 weeks. Treatment with CS extract significantly induced ATF3 expression and MUC5AC production in human bronchial epithelial cells, NCI-H292, and mouse tracheal epithelial cells. Interference of ATF3 significantly attenuated CS-induced MUC5AC expression in NCI-H292 and human bronchial epithelial cells. Mouse tracheal epithelial cells isolated from Atf3(-/-) mice also exhibited less MUC5AC production in response to CS extract treatment. In vivo, the Atf3(-/-) mice also displayed a significantly reduced mucus production relative to wild-type controls in response to chronic CS exposure. Furthermore, a chromatin immunoprecipitation assay revealed increased ATF3 binding to the MUC5AC promoter after CS treatment, and this transcriptional binding was significantly inhibited by knockdown of JUN, a subunit of activator protein-1. These results demonstrate that ATF3 may be involved in activator protein-1 signaling and transcriptional promotion of CS-induced MUC5AC expression in airway epithelial cells.
- SPARC Interacts with Actin in Skeletal Muscle in Vitro and in Vivo. [Journal Article]
- AJAm J Pathol 2016 Nov 28
- The cytoskeleton is an integral part of skeletal muscle structure, and reorganization of the cytoskeleton occurs during various modes of remodeling. We previously found that the extracellular matrix ...
The cytoskeleton is an integral part of skeletal muscle structure, and reorganization of the cytoskeleton occurs during various modes of remodeling. We previously found that the extracellular matrix protein secreted protein acidic and rich in cysteine (SPARC) is up-regulated and expressed intracellularly in developing muscle, during regeneration and in myopathies, which together suggests that SPARC might serve a specific role within muscle cells. Using co-immunoprecipitation combined with mass spectrometry and verified by staining for direct protein-protein interaction, we find that SPARC binds to actin. This interaction is present in regenerating myofibers of patients with Duchenne muscular dystrophy, polymyositis, and compartment syndrome. Analysis of the α-, β-, and γ-actin isoforms in SPARC knockout myoblasts reveals a changed expression pattern with dominance of γ-actin. In SPARC knockout mice, we performed an injury study to investigate whether lack of SPARC would compromise the ability to repair muscle. We report that these mice develop normal skeletal muscle with retained ability to regenerate. However, when we subject muscle from SPARC-deficient mice to an in vitro fatigue stimulation protocol, we find a defective force recovery. Therefore, SPARC appears to be an important modulator of the actin cytoskeleton, implicating maintenance of muscular function. This direct interaction with actin suggests a new role of SPARC during tissue remodeling.
- Correction. [Published Erratum]
- AJAm J Pathol 2016; 186(12):3316
- The Role of Substance P in Pulmonary Clearance of Bacteria in Comparative Injury Models. [Journal Article]
- AJAm J Pathol 2016; 186(12):3236-3245
- Neural input to the immune system can alter its ability to clear pathogens effectively. Patients suffering mild traumatic brain injury (mTBI) have shown reduced rates of pneumonia and a murine model ...
Neural input to the immune system can alter its ability to clear pathogens effectively. Patients suffering mild traumatic brain injury (mTBI) have shown reduced rates of pneumonia and a murine model replicated these findings, with better overall survival of TBI mice compared with sham-injured mice. To further investigate the mechanism of improved host response in TBI mice, this study developed and characterized a mild tail trauma model of similar severity to mild TBI. Both mild tail trauma and TBI induced similar systemic changes that normalized within 48 hours, including release of substance P. Examination of tissues showed that injuries are limited to the target tissue (ie, tail in tail trauma, brain in mTBI). Pneumonia challenge showed that mild TBI mice showed improved immune responses, characterized by the following: i) increased survival, ii) increased pulmonary neutrophil recruitment, iii) increased bacterial clearance, and iv) increased phagocytic cell killing of bacteria compared with tail trauma. Administration of a neurokinin-1-receptor antagonist to block substance P signaling eliminated the improved survival of mTBI mice. Neurokinin-1-receptor antagonism did not alter pneumonia mortality in tail trauma mice. These data show that immune benefits of trauma are specific to mTBI and that tail trauma is an appropriate control for future studies aimed at elucidating the mechanisms of improved innate immune responses in mTBI mice.
- Identification of an Epitope from Adenine Nucleotide Translocator 1 That Induces Inflammation in Heart in A/J Mice. [Journal Article]
- AJAm J Pathol 2016; 186(12):3160-3175
- Heart failure, a leading cause of death in humans, can emanate from myocarditis. Although most individuals with myocarditis recover spontaneously, some develop chronic dilated cardiomyopathy. Myocard...
Heart failure, a leading cause of death in humans, can emanate from myocarditis. Although most individuals with myocarditis recover spontaneously, some develop chronic dilated cardiomyopathy. Myocarditis may result from both infectious and noninfectious causes, including autoimmune responses to cardiac antigens. In support of this notion, intracellular cardiac antigens, like cardiac myosin heavy chain-α, cardiac troponin-I, and adenine nucleotide translocator 1 (ANT1), have been identified as autoantigens in cardiac autoimmunity. Herein, we demonstrate that ANT1 can induce autoimmune myocarditis in A/J mice by generating autoreactive T cells. We show that ANT1 encompasses multiple immunodominant epitopes (namely, ANT1 21-40, ANT1 31-50, ANT1 171-190, and ANT1 181-200). Although all four peptides induce comparable T-cell responses, only ANT1 21-40 was found to be a major myocarditogenic epitope in immunized animals. The myocarditis-inducing ability of ANT1 21-40 was associated with the generation of T cells producing predominantly IL-17A, and the antigen-sensitized T cells could transfer the disease to naïve recipients. These data indicate that cardiac mitochondrial proteins can be target autoantigens in myocarditis, supporting the notion that the antigens released as a result of primary damage may contribute to the persistence of chronic inflammation through autoimmunity.
- miR-182-5p Inhibition Ameliorates Ischemic Acute Kidney Injury. [Journal Article]
- AJAm J Pathol 2016 Nov 18
- Acute kidney injury (AKI) remains a major clinical event with high mortality rates. We previously identified renal miR-182 as the main driver of post-transplantation AKI. Therefore, we tested the cau...
Acute kidney injury (AKI) remains a major clinical event with high mortality rates. We previously identified renal miR-182 as the main driver of post-transplantation AKI. Therefore, we tested the causal inference of miR-182 by inhibiting its renal expression in vivo. In 45 rats AKI was induced by right nephrectomy and contralateral clamping of the renal pedicle for 40 minutes. Systemically administered antisense oligonucleotide (ASO) inhibited miR-182 in the kidneys up to 96 hours. The maximum creatinine elevation was on day 2 after injury (ASO 2.5 mg/kg: median, 1.9 mg/dL [interquartile range (IQR), 1.3 to 3.2 mg/dL]; ASO 25 mg/kg: median, 2.8 mg/dL [IQR, 0.7 to 5.0 mg/dL]; mismatch oligonucleotide (MM) 25 mg/kg: median, 5.7 mg/dL [IQR, 5.0 to 5.8 mg/dL]; saline: 4.4 mg/dL [IQR, 3.5 to 5.8 mg/dL]; P = 0.016, analysis of variance). Blinded semiquantitative histologic evaluation of renal biopsies showed better preserved morphology in both ASO groups than saline- and MM- treated kidneys [in overall injury scores, ASO both concentrations: median, 1 (IQR, 1 to 1); saline: median, 3 (IQR, 3 to 3); MM: median, 3 (IQR, 3 to 3); P < 0.001, analysis of variance]. ASO facilitated cell proliferation, metabolism, and angiogenesis on a genome-wide level. ASO when applied in normothermic kidney machine perfusion reduced renal miR-182 expression by more than two magnitudes. In summary, we showed that in vivo inhibition of miR-182 by ASO improved kidney function and morphology after AKI. This technique may be applicable to reduce the high rate of AKI in the human renal transplantation setting.
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- CD30 Induces Heat Shock Protein 90 and Signal Integration in Classic Hodgkin Lymphoma Cells. [Journal Article]
- AJAm J Pathol 2016 Nov 18
- Previous studies report deregulation of multiple signaling pathways in classic Hodgkin lymphoma (cHL) cells. However, the mechanisms of how these pathways are integrated are not fully understood. Her...
Previous studies report deregulation of multiple signaling pathways in classic Hodgkin lymphoma (cHL) cells. However, the mechanisms of how these pathways are integrated are not fully understood. Herein, we show involvement of cHL hallmark antigen CD30 in this process. CD30 facilitates phosphorylation of heat shock factor 1, activates heat shock promoter element, and induces heat shock protein (HSP) 90. CD30 repression and subsequent inhibition of HSP90 suppresses NF-κB, extracellular signal-regulated kinase, AKT, and STAT pathways in cHL cell lines. Thus, CD30-mediated induction of HSP90 appears to serve as a central hub for integration of intracellular signaling in cHL cells. We also show that CD30 induces HSP90 through phosphorylation of heat shock factor 1 via c-Jun N-terminal kinase in cHL cells. Although anaplastic large-cell lymphoma (ALCL) also is associated with CD30 overexpression, our experiments reveal that HSP90 induction in ALCL-bearing nucleophosmin-anaplastic lymphoma kinase (ALK) does not depend on CD30 but instead on ALK via c-Jun N-terminal kinase. Together, these results highlight a novel role for CD30 in mediating integration of signaling pathways of cHL cells while being replaced in this function by ALK in ALCL cells.