A role for CCL28–CCR3 in T-cell homing to the human upper airway mucosa

Lymphocyte recruitment to peripheral tissues is fundamental for immune surveillance and homeostasis, but the chemokines and chemokine receptors responsible for tissue-specific homing of T cells to the upper airway mucosa have not been determined. To address this, we analyzed the chemokines expressed in the normal human nasal mucosa and found that CCL28 is preferentially expressed at a high level on the lumenal face of vascular endothelial cells in the mucosa. Analysis of the cognate chemokine receptors revealed that close to 50% of the CD4⁺ T cells in the human nasal mucosa expressed the CCL28 receptor CCR3, whereas CCR3 was hardly detectable on T cells in the small intestine and skin. In the circulation, CCR3⁺ T cells comprised a small subset that did not express homing receptors to the intestine or skin. Moreover, depletion of CCR3⁺CD4⁺ T cells abrogated the proliferative response of human blood CD4⁺ T cells against the opportunistic nasopharyngeal pathogen Haemophilus influenzae, indicating that the CCR3⁺CD4⁺ T-cell subset in the circulation contains antigen specificities relevant for the upper airways. Together, these findings indicate that CCL28–CCR3 interactions are involved in the homeostatic trafficking of CD4⁺ T cells to the upper airways.     

CCR6− regulatory T cells blunt the restoration of gut Th17 cells along the CCR6–CCL20 axis in treated HIV-1-infected individuals

The gut CD4⁺ T cells, particularly the T helper type 17 (Th17) subset, are not completely restored in most HIV-1-infected individuals despite combined antiretroviral therapy, when initiated at the chronic phase of infection. We show here that the CCR6–CCL20 chemotactic axis is altered, with reduced CCL20 production by small intestine epithelial cells in treated HIV-1-infected individuals. This leads to impaired CCR6⁺CD4⁺ T-cell homing, particularly Th17 cells, to the small intestine mucosa. In contrast, the frequency of gut FoxP3⁺ T regulatory (Treg) cells, specifically the CCR6⁻ subset, was increased. The resulting imbalance in the Th17/CCR6⁻ Treg ratio and the associated shift from interleukin (IL)-17 to IL-10 and transforming growth factor-β (TGF-β) blunts CCL20 production by enterocytes, perpetuating a negative feedback for the recruitment of CCR6⁺CD4⁺ T cells to the small intestine in treated HIV-1-infected individuals.     

A role for the heat shock protein–CD91 axis in the initiation of immune responses to tumors

For over 100 years, it has been established that tumor-specific immune responses can frequently be detected in the tumor-bearing host. Whether or not these immune responses are capable of controlling the growth of the tumor is influenced by many factors. However, the mechanism by which the immune responses are initiated in the first place has remained a dilemma. In this chapter, we present evidence that heat shock protein-peptide complexes released by tumor cells are the entity responsible for initiating the immune responses. Interaction of the extracellular HSP with its receptor CD91 is necessary for priming the immune response. We propose that the disruption of the HSP-CD91 interaction may be an active mechanism by which tumors prevent the generation of immune responses against it.     

Estradiol-17β-responsive A1 and A2 noradrenergic cells of the brain stem project to the bed nucleus of the stria terminalis in the ewe brain: A possible route for regulation of gonadotropin releasing hormone cells

We have studied brain stem cells in the ewe brain that project to the bed nucleus of the stria terminalis (BNST) and determined if these cells are activated by estradiol-17β. This would predicate an indirect role in the estradiol-17β regulation of gonadotropin releasing hormone (GnRH) cells, since these receive input from the BNST. Ovariectomized ewes received 50 μg estradiol-17β benzoate (i.m.) 1 h prior to brain collection, so that activated cells could be identified by Fos immunohistochemistry. Retrograde tracer (FluoroGold; FG), was injected into the three divisions of the BNST and labeled cells were mapped to the A1 and A2 regions and the parabrachial nucleus (PBN) of the brain stem. With FG injection into the dorsal and lateral BNST, all FG-containing cells in the caudal A1 and 45% of those in A2 stained for dopamine-β-hydroxylase (DBH), indicating noradrenergic type. No FG-labelled cells in the PBN were DBH-positive. In A1 and A2 respectively, 42% and 46% of FG-labelled cells were Fos-positive, with no double-labeling in cells of the PBN. In ewes receiving FG injections into the ventral BNST, estrogen receptor (ER)α-immunoreactive nuclei were found in 82% of A1-FG labeled and 38% of A2-FG labeled cells. No FG-labelled cells of the PBN were ERα-positive. Anterograde tracing from A1 with microruby injection identified projections to the PBN, BNST and preoptic area (POA). Thus, A1 and A2 noradrenergic neurons project to the BNST in the ewe brain, express ERα and are activated by estradiol-17β. These noradrenergic, estrogen-responsive cells may provide indirect input to GnRH cells, via the BNST.     

The SMG5–SMG7 heterodimer directly recruits the CCR4–NOT deadenylase complex to mRNAs containing nonsense codons via interaction with POP2

Nonsense-mediated mRNA decay (NMD) is a eukaryotic quality control mechanism that detects aberrant mRNAs containing nonsense codons and induces their rapid degradation. This degradation is mediated by SMG6, an NMD-specific endonuclease, as well as the SMG5 and SMG7 proteins, which recruit general mRNA decay enzymes. However, it remains unknown which specific decay factors are recruited and whether this recruitment is direct. Here, we show that SMG7 binds directly to POP2, a catalytic subunit of the CCR4–NOT deadenylase complex, and elicits deadenylation-dependent decapping and 5′-to-3′ decay of NMD targets. Accordingly, a catalytically inactive POP2 mutant partially suppresses NMD in human cells. The SMG7–POP2 interaction is critical for NMD in cells depleted of SMG6, indicating that SMG7 and SMG6 act redundantly to promote the degradation of NMD targets. We further show that UPF1 provides multiple binding sites for decapping factors. These data unveil a missing direct physical link between NMD and the general mRNA decay machinery and indicate that NMD employs diverse and partially redundant mechanisms to ensure robust degradation of aberrant mRNAs.     

Response of human bone marrow stromal cells to a resorbable P2O5–SiO2–CaO–MgO–Na2O–K2O phosphate glass ceramic for tissue engineering applications

This work focuses on the synthesis and characterization of a novel bioresorbable glass ceramic phosphate-based material (GC-ICEL). More specifically, its solubility in different aqueous media (water, Tris–HCl and acellular simulated body fluid) and the response of human stromal cells cultured on it were investigated. X-ray diffraction analysis showed the presence of two crystalline phases identified as Na₂Mg(PO₄)₃ and Ca₂P₂O₇ and dissolution tests highlighted a preferential dissolution of the Na₂Mg(PO₄)₃ phase and of the residual amorphous phase in all the chosen media. Soaking tests in simulated body fluid showed precipitation of a hydroxyapatite layer, demonstrating the bioactivity of GC-ICEL, which is partially due to the reported bioactivity of Ca₂P₂O₇. The effect of GC-ICEL on adhesion, proliferation and osteoblastic gene expression of human bone marrow-derived stromal cells was also studied. Combining molecular and biochemical analyses, it was found that bone marrow cell differentiation was stimulated over proliferation on GC-ICEL. Moreover, the expression of bone-related genes in cells cultured on GC-ICEL confirmed the bioactivity of this phosphate-based glass ceramic, which might have a stimulatory effect on osteogenesis.     

Decreased circulating CXCR3 + CCR9+T helper cells are associated with elevated levels of their ligands CXCL10 and CCL25 in the salivary gland of patients with Sjögren’s syndrome to facilitate their concerted migration

CCR9 + T helper (Th) cells can induce Sjögren-like symptoms in mice and both CCR9 + Th cells and their ligand CCL25 are increased in the salivary glands of primary Sjögren's syndrome (pSS) patients. Increased circulating CCR9 + Th cells are present in pSS patients. CCR9 + Th cells are hyperresponsive to IL-7, secrete high levels of IFN-γ, IL-21, IL-17 and IL-4 and potently stimulate B cells in both patients and healthy individuals. Our aim was to study co-expression of chemokine receptors on CCR9 + Th cells and whether in pSS this might differentially affect CCR9 + Th cell frequencies. Frequencies of circulating CCR9 + and CCR9- Th cells co-expressing CXCR3, CCR4, CCR6 and CCR10 were studied in pSS patients and healthy controls. CCL25, CXCL10, CCL17, CCL20 and CCL27 mRNA and protein expression of salivary gland tissue of pSS and non-Sjögren's sicca (non-SS) patients was assessed. Chemotaxis assays were performed to study migration induced by CXCL10 and CCL25. Higher expression of CXCR3, CCR4 and CCR6 but not CCR10 was observed on CCR9 + Th cells as compared to cells lacking CCR9. Decreased frequencies of circulating memory CCR9 + CXCR3+ Th cells were found in pSS patients, which was most pronounced in the effector memory subset. Increased salivary gland CCL25 and CXCL10 expression significantly correlated and both ligands functioned synergistically based on in vitro induced chemotaxis. Decreased memory CXCR3 + CCR9+ Th cells in blood of pSS patients may be due to a concerted action of overexpressed ligands at the site of inflammation in the salivary glands facilitating their preferential migration and positioning in the lymphocytic infiltrates.     

New insights into the pathogenesis and treatment of peritoneal fibrosis: A potential role of Wnt/β-catenin induced epithelial to mesenchymal transition and stem cells for therapy

Peritoneal fibrosis is a chronic, progressive progress, which is associated with ultrafiltration failure. In the development of peritoneal fibrosis, Epithelial to mesenchymal transition is an important cellular process whereby epithelial cells transform into mesenchymal cells under physiology and pathology conditions, along with change of cell morphology and expression of related genes. It plays an important role in embryogenesis and development of tissues and organs, as well as organ fibrosis and tumorigenesis. Several intracellular signal transduction pathways induce the process of Epithelial to mesenchymal transition. In recent researches, Wnt/β-catenin induced epithelial to mesenchymal transition was suggested to be an important reason for tissues and organs fibrosis. The following paper reviews the potential role of Wnt/β-catenin induced epithelial to mesenchymal transition in peritoneal fibrosis. New potential therapeutic interventions of peritoneal fibrosis are discussed.     

Log-linear models for assessing gene–age interaction and their application to case-control studies of the apolipoprotein E (apoE) gene in Alzheimer's disease

Case-control studies provide a powerful approach for detecting disease-susceptibility genes or assessing gene–environment interactions. We investigated the situation in which the gene being studied plays a role in several diseases, and the allele frequency among subjects free of the disease of interest consequently decreases with age as subjects die from other diseases. The logistic model is one approach frequently used for analyzing case-control data, but it cannot accommodate this dependence of genotype and age. Using a log-linear model, we therefore proposed a hierarchical procedure that could be used as a valid method for assessing interactions in such situations. We then applied this procedure to observed data on Alzheimer's disease and the apolipoprotein E gene in Japan. We were able to derive an appropriate inference on whether the interaction was a gene–age interaction or merely a bias due to death from other diseases.     

Ligation of the left aorta in alligators affects acid–base balance: A role for the R → L shunt

This study investigated the effects of preventing bulk flow from the right ventricle to the body via the left aorta (LAo; right to left shunt, R → L) on acid–base status in alligators following feeding, during long-term fasting and a cold temperature exposure. Post-feeding pHv and Pv_CO2${\text{Pv}}_{{\text{CO}}_2}$ were not significantly different between S and C. Post-feeding pHv increased in both groups of alligators, but not significantly. During fasting, all acid–base variables were similar between the two groups of alligators. A 10 °C reduction in environmental temperature resulted in a significant difference in pHv and HCO₃⁻ between S and C. Both pHv and HCO₃⁻ were significantly higher in C animals. Pv_CO2${\text{Pv}}_{{\text{CO}}_2}$ significantly decreased in both groups during the cold exposure. Preventing the R → L shunt via the LAo had significant effects on acid–base balance in alligators indicating incomplete compensation for its loss and a role for the LAo in metabolic homeostasis.     

Effects of water-soluble cigarette smoke extracts upon the release of β-hexosaminidase from RBL-2H3 basophilic leukaemia cells in response to substance P,compound 48/80, concanavalin A and antigen stimulation

Objective and design: To determine whether water-soluble constituents of cigarette smoke affect mast cell function using an in vitro model, RBL-2H3 basophilic leukaemia cells. Materials and methods: RBL-2H3 cells were induced to degranulate in response to compound 48/80 and substance P, as assessed by monitoring the release of the granular enzyme -hexosaminidase, by treatment for 7 days with 20 M quercetin. Responses to concanavalin A and antigen were determined by measuring the -hexosaminidase release from cells cultured on fibronectin-coated plates. Results: The -hexosaminidase release response to compound 48/80 induced by quercetin treatment was accompanied by a release of lactate dehydrogenase, suggesting that degranulation is not the only process triggered by compound 48/80 under these conditions. Quercetin treatment reduced the -hexosaminidase release response to concanavalin A. Precoating of the culture wells with rat fibronectin enhanced the -hexosaminidase response to calcimycin, but not to concanavalin A. Under these conditions, concanavalin A did not induce a release of lactate dehydrogenase. The responses to c48/80, substance P, calcimycin, concanavalin A and antigen (after IgE pretreatment) were reduced by treatment with cigarette smoke solution obtained from standard and low-tar cigarettes (IR3 and IR5F). The effect of cigarette smoke solution from IR5F cigarettes upon the -hexosaminidase release elicited by compound 48/80 (in quercetin-treated cells) and by concanavalin A (in cells cultured on fibronectin-coated wells) could be prevented by N-acetyl-L-cysteine, but not with either hemoglobin, -tocopherol, catalase or palmitoylethanolamide. N-acetyl-L-cysteine also reduced the effect of cigarette smoke solution upon the degranulation response to antigen. Conclusion: Under the conditions used, oxidants present in cigarette smoke solution from IR5F cigarettes reduce the ability of RBL-2H3 cells to degranulate in response to both immunological and non-immunological stimuli.Received 6 December 2002; returned for revision 1 April 2003; accepted by I. Ahnfelt-RøJune 2003     

Induction of an ATP-polymerizing enzyme in TMV-infected tobacco and its homology to the human 2′–5′ a synthetase

Several reports have indicated that tobacco carries an enzyme (APE) that, in the presence of poly (rI):(rC), polymerizes ATP to oligoadenylates. This paper demonstrates that the tobacco APE system comprises several proteins (estimated sizes: 32, 42, 67, and 84±10% kD). Only one of these proteins (the 67-kD form) binds to poly (rI):(rC). This APE form has been purified by affinity chromatography on a synthetic ds-RNA column. Four tobacco proteins, including the purified one, crossreact with antibodies against the human enzyme, 2–5 A synthetase. The ATP-binding capacity of some of these proteins has also been demonstrated. The amount of plant oligoadenylates obtained by polymerizing ATP with the purified APE form allows, for the first time, their direct analysis by TLC. The TLC analysis indicated that the oligomer produced by APE is not identical to the 2–5 oligoadenylate.     

A 5th type of hypersensitivity reaction: Does incidental recruitment of autoreactive effector memory T-cells in response to minute amounts of PAMPs or DAMPs,underlie inflammatory episodes in the seronegative arthropathies and acute anterior uveitis?

There are many documented associations of the HLA B27-related diseases – the seronegative arthropathies (SNAs) and acute anterior uveitis (AAU). However, to date no single pathogenic mechanism has been proposed which can account for more than a few of these associations.The hypothesis presented in this paper is that individual inflammatory episodes in the SNAs and HLA B27 AAU are initiated locally within the inflamed site, in response to tiny quantities of innately recognised molecules – pathogen associated molecular patterns (PAMPs), and damage-associated molecular patterns (DAMPs). However, clinically significant responses to such small amounts of PAMPs/DAMPs only occur in those individuals who possess pre-existing tissue-specific autoreactive effector memory T-cells, which once recruited into the tissue where their autoantigen resides, can mediate inflammatory damage.The mechanism put forward here could explain much of the research evidence and distinguishing clinical features of the HLA B27-associated diseases. These include: the well documented involvement of micro-organisms and tissue damage in these diseases; the more frequent involvement of certain organs compared with others; the often local and asymmetrical presentation of the SNAs/AAU, which contrasts with other autoimmune diseases that effect many sites in a more diffuse/symmetrical pattern; the fact that these diseases which are thought to be mediated by T-cells, are nonetheless refractory to treatment with T-cell activation blockers.Finally, if correct, the mechanism would define a 5th type of clinical hypersensitivity reaction, one manifested by an excessive reaction to small pro-inflammatory signals, in individuals who possess tissue-specific autoreactive effector memory T-cells.     

Extending David Horrobin’s membrane phospholipid theory of schizophrenia: Overactivity of cytosolic phospholipase A2 in the brain is caused by overdrive of coupled serotonergic 5HT2A/2C receptors in response to stress

David Horrobin’s membrane phospholipid theory of schizophrenia has held up well over time because his therapeutic prediction that dietary supplementation with eicosapentaenoic acid (EPA) would have a therapeutic effect has been partially verified and undergoes continued testing. In the final version of his theory, he hypothesized that there was hyperactivity of phosphoslipase A₂ (PLA₂) or a related enzyme but did not explain how the hyperactivity came about. It is known that serotonergic 5HT_2A/2C receptors are coupled to PLA₂, which hydrolyzes both arachidonic acid (AA) and EPA from diacylglycerides at the sn-2 position. In this paper, Horrobin’s theory is combined with a previously published theory of chronic stress in which it was hypothesized that a disinhibited dorsal raphe nucleus, the principal nucleus of the serotonergic system, can organize the neuropathology of diseases such as migraine, hypertension, and the metabolic syndrome. The new or combined theory is that schizophrenia is a disease of chronic stress in which a disinhibited DRN causes widespread serotonergic overdrive in the cerebral cortex. This in turn causes overdrive of cPLA₂ and both central and peripheral depletion of AA and EPA. Because EPA is present in smaller amounts, it falls below threshold for maintaining an intracellular balance between AA-derived and EPA-derived second messenger cascades, which leads to abnormal patterns of neuronal firing. There are two causes of neuronal dysfunction: the disinhibited DRN and EPA depletion. Schizophrenia is statistically associated with metabolic syndrome, hypertension, and migraine because they form a cluster of diseases with similar pathophysiology.The theory provides an explanation for both the central and peripheral phospholipid abnormalities in schizophrenia. It also explains the role of stress in schizophrenia, elevated serum PLA₂ activity in schizophrenia, the relationship between untreated schizophrenia and metabolic syndrome, and the therapeutic rationale for EPA.     

Protein phosphatase 2A catalytic subunit α (PP2Acα) maintains survival of committed erythroid cells in fetal liver erythropoiesis through the STAT5 pathway

Suppression of programmed cell death is critical for the final maturation of red blood cells and depends largely on the anti-apoptotic effects of EpoR-STAT5-Bcl-x(L) signaling. As the major eukaryotic serine/threonine phosphatase, protein phosphatase 2A (PP2A) regulates multiple cellular processes, including apoptosis. However, whether PP2A plays a role in preventing erythroid cells from undergoing apoptosis remains to be elucidated. We conditionally inactivated the catalytic subunit α of PP2A (PP2Acα), which is the predominant form of PP2Ac, during early embryonic hematopoiesis. Loss of PP2Acα in hematopoietic cells perturbed definitive erythropoiesis characterized by fetal liver atrophy, reduced Ter119(+) cell number, abnormal expression patterns of molecular markers, less colony formation, and a reduction in definitive globin expression. Levels of erythropoiesis-promoting cytokines and initial seeding with hematopoietic progenitors remained unchanged in PP2Acα(TKO) fetal livers. We noted impaired expansion of the fetal erythroid compartment, which was associated with increased apoptosis of committed erythroid cells. Mechanistically, PP2Acα depletion markedly reduced Tyr(694) phosphorylation of STAT5 and expression of Bcl-x(L). Unexpectedly, PP2Acα-deficient embryos did not manifest any early embryonic vascular defects. Collectively, these data provide direct loss-of-function evidence demonstrating the importance of PP2Acα for the survival of committed erythroid cells during fetal liver erythropoiesis.