Biomedical subjects

Michael C Carroll

Publications and source records attributed to Michael C Carroll.

At least 19 recordsLinked to original sources

PubMed2006

Involvement of NFAT1 in B cell self-tolerance.

B cells from anti-lysozyme Ig/soluble lysozyme double-transgenic mice are chronically exposed to self-Ag in the periphery, resulting in an anergic phenotype. Chronic exposure to self-Ag leads to nuclear translocation of NFAT1 and NFAT2, suggesting that they are involved in anergy. To directly test a role for NFAT1 in B cell anergy, NFAT1-deficient mice were crossed with anti-lysozyme Ig transgenic mice. As expected, B cell anergy was evident in the presence of self-Ag based on reduced serum anti-lysozyme levels, percentage and number of mature B cells, and reduced B cell responsiveness. By contrast, B cell anergy was relieved in NFAT1(-/-) mice expressing soluble self-Ag. Bone marrow development was equivalent in NFAT1-sufficient and -deficient mice, suggesting that loss of anergy in the latter is due to selection later in development. Taken together, these studies provide direct evidence that the transcription factor NFAT1 is involved in B cell anergy.

Animals↗

PubMed2006

Alternative complement pathway activation is essential for inflammation and joint destruction in the passive transfer model of collagen-induced arthritis.

Activation of each complement initiation pathway (classical, alternative, and lectin) can lead to the generation of bioactive fragments with resulting inflammation in target organs. The objective of the current study was to determine the role of specific complement activation pathways in the pathogenesis of experimental anti-type II collagen mAb-passive transfer arthritis. C57BL/6 mice were used that were genetically deficient in either the alternative pathway protein factor B (Bf(-/-)) or in the classical pathway component C4 (C4(-/-)). Clinical disease activity was markedly decreased in Bf(-/-) compared with wild-type (WT) mice (0.5 +/- 0.22 (n = 6) in Bf(-/-) vs 8.83 +/- 0.41 (n = 6) in WT mice (p < 0.0001)). Disease activity scores were not different between C4(-/-) and WT mice. Analyses of joints showed that C3 deposition, inflammation, pannus, cartilage, and bone damage scores were all significantly less in Bf(-/-) as compared with WT mice. There were significant decreases in mRNA levels of C3, C4, CR2, CR3, C3aR, and C5aR in the knees of Bf(-/-) as compared with C4(-/-) and WT mice with arthritis; mRNA levels for complement regulatory proteins did not differ between the three strains. These results indicate that the alternative pathway is absolutely required for the induction of arthritis following injection of anti-collagen Abs. The mechanisms by which these target organ-specific mAbs bypass the requirements for engagement of the classical pathway remain to be defined but do not appear to involve a lack of alternative pathway regulatory proteins.

Animals↗

PubMed2006

Natural antibody mediated innate autoimmune response.

Recent advance in autoimmunity research reveals that the innate immune system is able to recognize self-targets and initiate inflammatory response in a similar way as with pathogens. This review describes one novel example of this innate autoimmunity, ischemia-reperfusion (I/R) injury. Studies of intestinal, skeletal muscle, and heart I/R models showed that reperfusion of ischemic tissues elicits an acute inflammatory response involving serum complement system which is activated by natural IgM. The recent identification of a monoclonal natural IgM that initiates I/R led to the identification of non-muscle myosin heavy chain type II A and C as the self-targets in two different tissues. New evidence further suggests that IgM binds initially to ischemic antigen providing a binding site for mannan binding lectin (MBL) which subsequently leads to activation of complement and results in tissue injury. Therefore, natural IgM mediated innate autoimmunity is likely responsible for the detrimental consequences in ischemic diseases.

Animals↗

PubMed2006

The role of natural IgM in myocardial ischemia-reperfusion injury.

Myocardial ischemia-reperfusion injury represents a combination of factors, namely the intrinsic cellular response to ischemia and the extrinsic acute inflammatory response. Recent studies in mesenteric and skeletal muscle reperfusion models identified natural IgM as a major initiator of pathology through the activation of the complement system and inflammatory cells. To determine whether a similar mechanism is involved in myocardial tissues, mice bearing an altered natural IgM repertoire (Cr2-/-) were examined in a murine model of coronary artery ischemia. Notably, these mice were significantly protected based on the reduced infarct size, limited apoptosis of cardiomyocytes, and decreased neutrophil infiltration. Protection was IgM-dependent as reconstitution of these mice with wild-type IgM restored myocardial reperfusion injury. These results support a model in which natural IgM initiates the acute inflammatory response in the myocardium following ischemia and reperfusion.

Animals↗

PubMed2006

T cell-independent and T cell-dependent immunoglobulin G responses to polyomavirus infection are impaired in complement receptor 2-deficient mice.

Polyomavirus (PyV) infection induces protective T cell-independent (TI) IgM and IgG antibody responses in T cell-deficient mice, but these responses are not generated by immunization with viral proteins or virus like particles. We hypothesized that innate signals contribute to the generation of isotype-switched antiviral antibody responses. We studied the role of complement receptor (CR2) engagement in TI and T cell-dependent (TD) antibody responses to PyV using CR2-deficient mice. Antiviral IgG responses were reduced by 80-40% in CR2-/- mice compared to wild type. Adoptive transfer experiments demonstrated the need for CR2 not only in TD, but also in TI IgG responses to PyV. Transfer of CR2-/- B lymphocytes to SCID mice resulted in TI antiviral IgG responses that corresponded to 10% of that seen in wild-type B cell-reconstituted mice. Thus, our studies revealed a profound dependence of TI and TD antiviral antibody responses on CR2-mediated signals in PyV-infected mice, where the viral antigen is abundant and persistent.

Adoptive Transfer↗

PubMed2006

Factor B of the alternative complement pathway regulates development of airway hyperresponsiveness and inflammation.

Exposure to inhaled allergens leads to increases in airway hyperresponsiveness (AHR) and inflammation, associated with increased levels of biologically active fragments derived from the complement C3 and C5 family of proteins. Further, complement activation during allergen challenge in sensitized animals is necessary for the development of AHR and airway inflammation. To define the complement pathway involved, we studied mice deficient in complement factor 4 (C4-/-), a critical component of the classical pathway, or factor B (fB-/-), an essential protein in the alternative complement pathway. WT, C4-/-, and fB-/- mice were sensitized to ovalbumin and subsequently exposed to nebulized ovalbumin (1% in saline) on 3 consecutive days. After allergen sensitization and challenge, fB-/- mice demonstrated significantly lower airway responsiveness to methacholine and less airway inflammation. In contrast, C4-/- mice showed no reduction in AHR and airway inflammation compared with WT mice. Tissue inflammation, goblet cell hyperplasia, and IL-4, IL-5, and IL-13 levels in BAL fluid were significantly reduced in fB-/- mice compared with C4-/- and WT mice. The development of AHR and airway inflammation in sensitized fB-/- mice could be restored after intranasal administration of purified factor B before the airway challenge. In addition, administration of a neutralizing anti-factor B mAb to sensitized mice before airway challenge reduced the development of AHR and airway inflammation. These results demonstrate that in sensitized hosts complement activation through the alternative pathway after allergen exposure is critical to the development of AHR and airway inflammation.

Animals↗

PubMed2006

Emerging patterns in complement-mediated pathogen recognition.

Protection from acute infection requires an immediate response by the host innate immune system. Two recent Cell papers, including one in this issue, provide new insight into the role of the complement system in response to blood-borne pathogens.

Animals↗

PubMed2006

Identification of the target self-antigens in reperfusion injury.

Reperfusion injury (RI), a potential life-threatening disorder, represents an acute inflammatory response after periods of ischemia resulting from myocardial infarction, stroke, surgery, or trauma. The recent identification of a monoclonal natural IgM that initiates RI led to the identification of nonmuscle myosin heavy chain type II A and C as the self-targets in two different tissues. These results identify a novel pathway in which the innate response to a highly conserved self-antigen expressed as a result of hypoxic stress results in tissue destruction.

Animals↗

PubMed2006

Attenuation of skeletal muscle reperfusion injury with intravenous 12 amino acid peptides that bind to pathogenic IgM.

BACKGROUND: The injury sustained by reperfused skeletal muscle is inflammatory and is initiated by binding of pre-formed IgM to involved tissue, followed by local complement activation and further inflammation. A clone of natural IgM has been described that initiates this injury, suggesting that specific antigens are exposed on ischemic tissues that act as ligands for this pathogenic antibody. In these experiments, we examine the properties of short peptide sequences, and their homologues, that bind to the antigen-combining site of this pathogenic IgM clone. METHODS: A 12-mer phage display library was biopanned with the pathogenic IgM clone and then negatively selected against an inactive natural IgM clone. All 8 clones that bound specifically to the pathogenic IgM had closely related amino acid sequences. P8 is the clone that bound most avidly. Tissue lysates from ischemic tissue were reacted with pathogenic IgM, and immune complexes isolated and analyzed on SDS-PAGE. Bands were excised and sequenced, identifying non-muscle myosin as the protein reacting with pathogenic antibody in ischemic gut and glycogen phosphorylase as the counterpart in ischemic skeletal muscle. Both proteins contain sequence homologous to P8; N2 and GP1 are the natural 12-mers homologues that are contained within non-muscle myosin and glycogen phosphorylase, respectively. Wild-type C57/Bl6 mice, divided into groups receiving saline, P8, N2, GP1, or a random peptide at the start of the experiment, were subjected to 2 hours of tourniquet induced hind limb ischemia and 3 hours of reperfusion. Muscle was assessed for injury with histology and for immune activation with histochemistry. RESULTS: Intravenous administration of P8, N2, and GP1 led to significant attenuation of muscle injury (13 +/- 1.8 injured fibers/50 counted, 12 +/- 0.81, 8.0 +/- 0.73 respectively) after reperfusion injury compared to animals receiving saline (26 +/- 2.3) or the same mass of a random peptide (22 +/- 2.3), P less than .05. This level of protection from injury is comparable to that seen in the absence of antibody altogether. As well, P8-treated animals exhibited a marked decrease in deposition of IgM (as well as C3) in comparison to saline treated controls. CONCLUSIONS: Specific peptide blockade of an injury-inducing IgM clone decreased the local consequences of skeletal muscle ischemia/reperfusion injury in wild-type animals that have the full repertoire of IgM specificities. This indicates that the antibodies that initiate reperfusion injury have specificity only for P8-related antigens. This could also indicate that the variety of relevant ischemic antigens is quite restricted.

Animals↗

PubMed2006

Transcriptional basis of lymphocyte tolerance.

Signaling through lymphocyte antigen receptors has the potential to initiate several distinct outcomes: proliferation, differentiation, apoptosis, or functional unresponsiveness. Expansion and differentiation of effector T cells is required for defense against foreign antigens, whereas functional unresponsiveness, termed anergy, is a cell-intrinsic mechanism that contributes to peripheral self-tolerance. Other mechanisms of peripheral tolerance include the 'dominant' tolerance imposed by regulatory T cells and immunosuppression mediated by interleukin-10 and transforming growth factor-beta. T- and B-cell antigen receptor ligation induces an increase in intracellular calcium levels as well as activating additional signaling pathways that are further potentiated by costimulatory receptors. In this review, we argue that cell-intrinsic programs of peripheral anergy and tolerance are imposed by sustained calcium signaling in lymphocytes. We address in particular the role of the calcium-dependent transcription factor nuclear factor for activation of T cells, which is activated by antigen receptor stimulation and, depending on the presence or absence of input from its transcriptional partner, activator protein-1, dictates two distinct transcriptional programs: activation or tolerance.

Animals↗

PubMed2006

Optimal long-term humoral responses to replication-defective herpes simplex virus require CD21/CD35 complement receptor expression on stromal cells.

Replication-defective herpes simplex virus (HSV) strains elicit durable immune responses and protect against virulent HSV challenge in mice, despite being unable to establish latent infection in neuronal cells. Mechanisms for generating long-lived immunity in the absence of viral persistence remain uncertain. In animals immunized with replication-defective HSV, durable serum immunoglobulin G (IgG) responses were elicited. Surprisingly, Western blot analyses revealed that the specificities of antiviral IgG changed over time, and antibody reactivity to some viral proteins was detected only very late. Thus, some of the durable IgG activity appeared to be contributed by either new or significantly enhanced antibody responses at late times. Following immunization, radiation bone marrow-chimeric mice lacking complement receptors CD21 and CD35 on stromal cells elicited only short-lived serum IgG and failed to mount recall responses to subsequent HSV exposure. Our results suggest that complement-mediated retention of viral antigens by stromal cells, such as follicular dendritic cells, is critical for optimal maintenance of antibody responses and B-cell memory following vaccination with replication-defective HSV.

Animals↗

PubMed2005

CD21/CD19 coreceptor signaling promotes B cell survival during primary immune responses.

The adaptive immune response is tightly regulated to limit responding cells in an Ag-specific manner. On B cells, coreceptors CD21/CD19 modulate the strength of BCR signals, potentially influencing cell fate. The importance of the CD95 pathway was examined in response of B cells to moderate affinity Ag using an adoptive transfer model of lysozyme-specific Ig transgenic (HEL immunoglobulin transgene (MD4) strain) B cells. Although adoptively transferred Cr2+/+ MD4 B cells are activated and persist within splenic follicles of duck egg lysozyme-immunized mice, Cr2-/- MD4 B cells do not. In contrast, Cr2-/- MD4 lpr B cells persist after transfer, suggesting that lack of CD21/CD35 signaling results in CD95-mediated elimination. Cr2 deficiency did not affect CD95 levels, but cellular FLIP (c-FLIP) protein and mRNA levels were reduced 2-fold compared with levels in Cr2+/+ MD4 B cells. In vitro culture with Cr2+/+ MD4 B cells demonstrated that equimolar amounts of rHEL-C3d3 were more effective than hen egg lysozyme alone in up-regulating c-FLIP levels and for protection against CD95-mediated apoptosis. Collectively, this study implies a mechanism for regulating B cell survival in vivo whereby the strength of BCR signaling (including coreceptor) determines c-FLIP levels and protection from CD95-induced death.

Animals↗

PubMed2005

Mast cell protease 5 mediates ischemia-reperfusion injury of mouse skeletal muscle.

Ischemia with subsequent reperfusion (IR) injury is a significant clinical problem that occurs after physical and surgical trauma, myocardial infarction, and organ transplantation. IR injury of mouse skeletal muscle depends on the presence of both natural IgM and an intact C pathway. Disruption of the skeletal muscle architecture and permeability also requires mast cell (MC) participation, as revealed by the fact that IR injury is markedly reduced in c-kit defective, MC-deficient mouse strains. In this study, we sought to identify the pathobiologic MC products expressed in IR injury using transgenic mouse strains with normal MC development, except for the lack of a particular MC-derived mediator. Histologic analysis of skeletal muscle from BALB/c and C57BL/6 mice revealed a strong positive correlation (R(2) = 0.85) between the extent of IR injury and the level of MC degranulation. Linkage between C activation and MC degranulation was demonstrated in mice lacking C4, in which only limited MC degranulation and muscle injury were apparent. No reduction in injury was observed in transgenic mice lacking leukotriene C(4) synthase, hemopoietic PGD(2) synthase, N-deacetylase/N-sulfotransferase-2 (enzyme involved in heparin biosynthesis), or mouse MC protease (mMCP) 1. In contrast, muscle injury was significantly attenuated in mMCP-5-null mice. The MCs that reside in skeletal muscle contain abundant amounts of mMCP-5 which is the serine protease that is most similar in sequence to human MC chymase. We now report a cytotoxic activity associated with a MC-specific protease and demonstrate that mMCP-5 is critical for irreversible IR injury of skeletal muscle.

Animals↗

PubMed2005

Defective B cell responses in the absence of SH2D1A.

More than half of patients with X-linked lympho-proliferative disease, which is caused by a defect in the intracellular adapter protein SH2D1A, suffer from an extreme susceptibility to Epstein-Barr virus. One-third of these patients, however, develop dysgammaglobulenemia without an episode of severe mononucleosis. Here we show that in SH2D1A(-/-) mice, both primary and secondary responses of all Ig subclasses are severely impaired in response to specific antigens. Because germinal centers were absent in SH2D1A(-/-) mice upon primary immunization, and because SH2D1A was detectable in wt germinal center B cells, we examined whether SH2D1A(-/-) B cell functions were impaired. Using the adoptive cotransfer of B lymphocytes from hapten-primed SH2D1A(-/-) mice with CD4(+) T cells from primed wt mice into irradiated wt mice provided evidence that signal transduction events controlled by SH2D1A are essential for B cell activities resulting in antigen specific IgG production. Defects in naive SH2D1A(-/-) B cells became evident upon cotransfer with non-primed wt CD4(+) cells into Rag2(-/-) recipients. Thus, both defective T and B cells exist in the absence of SH2D1A, which may explain the progressive dysgammaglobulinemia in a subset of X-linked lympho-proliferative disease patients without involvement of Epstein-Barr virus.

Adoptive Transfer↗

PubMed2005

Innate autoimmunity.

The adaptive immune system has evolved highly specific pattern recognition proteins and receptors that, when triggered, provide a first line of host defense against pathogens. Studies reveal that these innate recognition proteins are also self-reactive and can initiate inflammation against self-tissues in a similar manner as with pathogens. This specific event is referred to as "innate autoimmunity." In this review, we describe two classes of autoimmune responses, that is, reperfusion injury and fetal loss syndrome, in which the recognition and injury are mediated by innate immunity. Both disorders are common and are clinically important. Reperfusion injury (RI) represents an acute inflammatory response after a reversible ischemic event and subsequent restoration of blood flow. Findings that injury is IgM and complement dependent and that a single natural antibody prepared from a panel of B-1 cell hybridomas can restore injury in antibody-deficient mice suggest that RI is an autoimmune-type disorder. Fetal loss syndrome is also an antibody- and complement-dependent disorder. Although both immune and natural antibodies are likely involved in recognition of phospholipid self-antigens, inhibition of the complement pathway in rodent models can block fetal loss. As new innate recognition proteins and receptors are identified, it is likely that innate responses to self represent frequent events and possibly underlie many of the known chronic autoimmune disorders normally attributable to dysregulation of adaptive immunity.

Animals↗

PubMed2004

Identification of a specific self-reactive IgM antibody that initiates intestinal ischemia/reperfusion injury.

Reperfusion injury of ischemic tissue represents an acute inflammatory response that can cause significant morbidity and mortality. The mechanism of injury is not fully elucidated, but recent studies indicate an important role for natural antibody and the classical pathway of complement. To test the hypothesis that injury is initiated by specific IgM, we have screened a panel of IgM-producing hybridomas prepared from peritoneal cells enriched in B-1 cells. One clone, CM22, was identified that could restore pathogenic injury in RAG-1(-/-) mice in an intestinal model of ischemia/reperfusion (I/R). In situ activation of the classical pathway of complement was evident by deposition of IgM, complement C4, and C3 in damaged tissue after passive transfer of CM22 IgM. Sequence analysis of CM22 Ig heavy and light chains showed germ-line configurations with high homology to a V(H) sequence from the B-1 repertoire and a V(K) of a known polyreactive natural IgM. These data provide definitive evidence that I/R injury can be initiated by clonally specific natural IgM that activates the classical pathway of complement. This finding opens an avenue for identification of I/R-specific self-antigen(s) and early prevention of injury.

Animals↗

PubMed2004

B cell receptor signal strength determines B cell fate.

B cell receptor (BCR)-mediated antigen recognition is thought to regulate B cell differentiation. BCR signal strength may also influence B cell fate decisions. Here, we used the Epstein-Barr virus protein LMP2A as a constitutively active BCR surrogate to study the contribution of BCR signal strength in B cell differentiation. Mice carrying a targeted replacement of Igh by LMP2A leading to high or low expression of the LMP2A protein developed B-1 or follicular and marginal zone B cells, respectively. These data indicate that BCR signal strength, rather than antigen specificity, determines mature B cell fate. Furthermore, spontaneous germinal centers developed in gut-associated lymphoid tissue of LMP2A mice, indicating that microbial antigens can promote germinal centers independently of BCR-mediated antigen recognition.

Animals↗

PubMed2004

The complement system in B cell regulation.

Early studies of animals bearing natural deficiencies in complement C3 and C4 and mice transiently deficient in C3 suggested that the complement system played a role in humoral immunity. Identification and characterization of the complement receptors CD21 and CD35 and their expression on B lymphocytes provided evidence for a direct role for complement in "linkage of innate and adaptive immunity". More recent studies of mice bearing targeted deficiencies in complement proteins C3, C4 or the receptors CD21/CD35 has confirmed the importance of complement in B cell responses in vivo and extended our understanding to distinct stages in B cell differentiation in which complement participates in humoral immunity. In this review, a role for complement is described in five distinct stages of B cell differentiation.

Animals↗