Blocking FcRn in humans reduces circulating IgG levels and inhibits IgG immune complex-mediated immune responses.

The neonatal crystallizable fragment receptor (FcRn) functions as an intracellular protection receptor for immunoglobulin G (IgG). Recently, several clinical studies have reported the lowering of circulating monomeric IgG levels through FcRn blockade for the potential treatment of autoimmune diseases. Many autoimmune diseases, however, are derived from the effects of IgG immune complexes (ICs). We generated, characterized, and assessed the effects of SYNT001, a FcRn-blocking monoclonal antibody, in mice, nonhuman primates (NHPs), and humans. SYNT001 decreased all IgG subtypes and IgG ICs in the circulation of humans, as we show in a first-in-human phase 1, single ascending dose study. In addition, IgG IC induction of inflammatory pathways was dependent on FcRn and inhibited by SYNT001. These studies expand the role of FcRn in humans by showing that it controls not only IgG protection from catabolism but also inflammatory pathways associated with IgG ICs involved in a variety of autoimmune diseases.

Auditory evoked fields measured noninvasively with small-animal MEG reveal rapid repetition suppression in the guinea pig.

In animal models, single-neuron response properties such as stimulus-specific adaptation have been described as possible precursors to mismatch negativity, a human brain response to stimulus change. In the present study, we attempted to bridge the gap between human and animal studies by characterising responses to changes in the frequency of repeated tone series in the anesthetised guinea pig using small-animal magnetoencephalography (MEG). We showed that 1) auditory evoked fields (AEFs) qualitatively similar to those observed in human MEG studies can be detected noninvasively in rodents using small-animal MEG; 2) guinea pig AEF amplitudes reduce rapidly with tone repetition, and this AEF reduction is largely complete by the second tone in a repeated series; and 3) differences between responses to the first (deviant) and later (standard) tones after a frequency transition resemble those previously observed in awake humans using a similar stimulus paradigm.

Depth-dependent temporal response properties in core auditory cortex.

The computational role of cortical layers within auditory cortex has proven difficult to establish. One hypothesis is that interlaminar cortical processing might be dedicated to analyzing temporal properties of sounds; if so, then there should be systematic depth-dependent changes in cortical sensitivity to the temporal context in which a stimulus occurs. We recorded neural responses simultaneously across cortical depth in primary auditory cortex and anterior auditory field of CBA/Ca mice, and found systematic depth dependencies in responses to second-and-later noise bursts in slow (1-10 bursts/s) trains of noise bursts. At all depths, responses to noise bursts within a train usually decreased with increasing train rate; however, the rolloff with increasing train rate occurred at faster rates in more superficial layers. Moreover, in some recordings from mid-to-superficial layers, responses to noise bursts within a 3-4 bursts/s train were stronger than responses to noise bursts in slower trains. This non-monotonicity with train rate was especially pronounced in more superficial layers of the anterior auditory field, where responses to noise bursts within the context of a slow train were sometimes even stronger than responses to the noise burst at train onset. These findings may reflect depth dependence in suppression and recovery of cortical activity following a stimulus, which we suggest could arise from laminar differences in synaptic depression at feedforward and recurrent synapses.

Stimulus-specific adaptation occurs in the auditory thalamus.

Neurons in the primary auditory cortex respond less strongly to a commonly occurring "standard" tone than to the same tone when it is rare or "deviant." This phenomenon, called "stimulus-specific adaptation" (SSA), has been proposed as a possible single-neuron correlate of the mismatch negativity, a cortical evoked potential associated with stimulus novelty. Previous studies in cat did not observe SSA in single neurons in the auditory thalamus. However, these reports did not differentiate between the auditory thalamic subdivisions and did not examine the effects of changing the stimulus presentation rate. To explore the possibility of thalamic SSA more completely, we recorded extracellularly from 30 single units and 22 multiunit clusters in the ventral, medial, and dorsal subdivisions of the mouse medial geniculate body (MGB), while presenting the anesthetized animals with sequences of standard and deviant tones at interstimulus intervals of 400, 500 and 800 ms. We found SSA in the auditory thalamus at all three stimulus presentation rates, primarily in the medial subdivision but to a lesser degree also in the ventral MGB. Thalamic SSA was evident from the earliest onset of tone-evoked activity, although the latencies of responses to standard and deviant tones were not significantly different. Together with related findings of SSA in neurons of the "belt" regions of the inferior colliculus, these results demonstrate that SSA is present at subcortical levels, primarily in but not restricted to the nonlemniscal auditory pathway.

Mouse auditory cortex differs from visual and somatosensory cortices in the laminar distribution of cytochrome oxidase and acetylcholinesterase.

Cytochrome oxidase (CYO) and acetylcholinesterase (AChE) staining density varies across the cortical layers in many sensory areas. The laminar variations likely reflect differences between the layers in levels of metabolic activity and cholinergic modulation. The question of whether these laminar variations differ between primary sensory cortices has never been systematically addressed in the same set of animals, since most studies of sensory cortex focus on a single sensory modality. Here, we compared the laminar distribution of CYO and AChE activity in the primary auditory, visual, and somatosensory cortices of the mouse, using Nissl-stained sections to define laminar boundaries. Interestingly, for both CYO and AChE, laminar patterns of enzyme activity were similar in the visual and somatosensory cortices, but differed in the auditory cortex. In the visual and somatosensory areas, staining densities for both enzymes were highest in layers III/IV or IV and in lower layer V. In the auditory cortex, CYO activity showed a reliable peak only at the layer III/IV border, while AChE distribution was relatively homogeneous across layers. These results suggest that laminar patterns of metabolic activity and cholinergic influence are similar in the mouse visual and somatosensory cortices, but differ in the auditory cortex.

Cross-correlation in the auditory coincidence detectors of owls.

Interaural time difference (ITD) plays a central role in many auditory functions, most importantly in sound localization. The classic model for how ITD is computed was put forth by Jeffress (1948). One of the predictions of the Jeffress model is that the neurons that compute ITD should behave as cross-correlators. Whereas cross-correlation-like properties of the ITD-computing neurons have been reported, attempts to show that the shape of the ITD response function is determined by the spectral tuning of the neuron, a core prediction of cross-correlation, have been unsuccessful. Using reverse correlation analysis, we demonstrate in the barn owl that the relationship between the spectral tuning and the ITD response of the ITD-computing neurons is that predicted by cross-correlation. Moreover, we show that a model of coincidence detector responses derived from responses to binaurally uncorrelated noise is consistent with binaural interaction based on cross-correlation. These results are thus consistent with one of the key tenets of the Jeffress model. Our work sets forth both the methodology to answer whether cross-correlation describes coincidence detector responses and a demonstration that in the barn owl, the result is that expected by theory.

The consequences of response nonlinearities for interpretation of spectrotemporal receptive fields.

Neurons in the central auditory system are often described by the spectrotemporal receptive field (STRF), conventionally defined as the best linear fit between the spectrogram of a sound and the spike rate it evokes. An STRF is often assumed to provide an estimate of the receptive field of a neuron, i.e., the spectral and temporal range of stimuli that affect the response. However, when the true stimulus-response function is nonlinear, the STRF will be stimulus dependent, and changes in the stimulus properties can alter estimates of the sign and spectrotemporal extent of receptive field components. We demonstrate analytically and in simulations that, even when uncorrelated stimuli are used, interactions between simple neuronal nonlinearities and higher-order structure in the stimulus can produce STRFs that show contributions from time-frequency combinations to which the neuron is actually insensitive. Only when spectrotemporally independent stimuli are used does the STRF reliably indicate features of the underlying receptive field, and even then it provides only a conservative estimate. One consequence of these observations, illustrated using natural stimuli, is that a stimulus-induced change in an STRF could arise from a consistent but nonlinear neuronal response to stimulus ensembles with differing higher-order dependencies. Thus, although the responses of higher auditory neurons may well involve adaptation to the statistics of different stimulus ensembles, stimulus dependence of STRFs alone, or indeed of any overly constrained stimulus-response mapping, cannot demonstrate the nature or magnitude of such effects.

Preservation of spectrotemporal tuning between the nucleus laminaris and the inferior colliculus of the barn owl.

Performing sound recognition is a task that requires an encoding of the time-varying spectral structure of the auditory stimulus. Similarly, computation of the interaural time difference (ITD) requires knowledge of the precise timing of the stimulus. Consistent with this, low-level nuclei of birds and mammals implicated in ITD processing encode the ongoing phase of a stimulus. However, the brain areas that follow the binaural convergence for the computation of ITD show a reduced capacity for phase locking. In addition, we have shown that in the barn owl there is a pooling of ITD-responsive neurons to improve the reliability of ITD coding. Here we demonstrate that despite two stages of convergence and an effective loss of phase information, the auditory system of the anesthetized barn owl displays a graceful transition to an envelope coding that preserves the spectrotemporal information throughout the ITD pathway to the neurons of the core of the central nucleus of the inferior colliculus.

Noise reduction of coincidence detector output by the inferior colliculus of the barn owl.

A recurring theme in theoretical work is that integration over populations of similarly tuned neurons can reduce neural noise. However, there are relatively few demonstrations of an explicit noise reduction mechanism in a neural network. Here we demonstrate that the brainstem of the barn owl includes a stage of processing apparently devoted to increasing the signal-to-noise ratio in the encoding of the interaural time difference (ITD), one of two primary binaural cues used to compute the position of a sound source in space. In the barn owl, the ITD is processed in a dedicated neural pathway that terminates at the core of the inferior colliculus (ICcc). The actual locus of the computation of the ITD is before ICcc in the nucleus laminaris (NL), and ICcc receives no inputs carrying information that did not originate in NL. Unlike in NL, the rate-ITD functions of ICcc neurons require as little as a single stimulus presentation per ITD to show coherent ITD tuning. ICcc neurons also displayed a greater dynamic range with a maximal difference in ITD response rates approximately double that seen in NL. These results indicate that ICcc neurons perform a computation functionally analogous to averaging across a population of similarly tuned NL neurons.

Quantitative analysis of the immune response to mouse non-MHC transplantation antigens in vivo: the H60 histocompatibility antigen dominates over all others.

Minor histocompatibility Ags (minor H Ags) are substantial impediments to MHC-matched solid tissue and bone marrow transplantation. From an antigenic standpoint, transplantation between MHC-matched individuals has the potential to be remarkably complex. To determine the extent to which the immune response is simplified by the phenomenon of immunodominance, we used peptide/MHC tetramers based on recently discovered minor H Ags (H60, H13, and HY) and monitored in vivo CD8 T cell responses of female C57BL/6 mice primed with MHC-matched, but background-disparate, male BALB.B cells. CD8 T cells against H60 overwhelmed responses to the H13 and HY throughout primary and secondary challenge. H60 immunodominance was an inherent quality, overcoming a lower memory precursor frequency compared with that of H13 and evoking a T cell response with diverse TCRV beta usage. IFN-gamma staining examining congenically defined minor H Ags extended H60 dominance over additional minor H Ags, H28, H4, and H7. These four minor H Ags accounted for up to 85% of the CD8 T cell response, but H60 stood out as the major contributor. These findings show that immunodominance applies to antigenically complex transplantation settings in vivo and that the responses to the H60 minor H Ag dominates in this model. We suggest that immunodominant minor H Ags are those that result from the absence of a self analog.

Differences that matter: major cytotoxic T cell-stimulating minor histocompatibility antigens.

Despite thousands of genetic polymorphisms among MHC matched mouse strains, a few unknown histocompatibility antigens are targeted by the cytotoxic T cells specific for tissue grafts. We isolated the cDNA of a novel BALB.B antigen gene that defines the polymorphic H28 locus on chromosome 3 and yields the naturally processed ILENFPRL (IFL8) peptide for presentation by Kb MHC to C57BI/6 CTL. The CTL specific for the IFL8/Kb and our previously identified H60/Kb complexes represent a major fraction of the B6 anti-BALB.B immune response. The immunodominance of these antigens can be explained by their differential transcription in the donor versus the host strains and their expression in professional donor antigen-presenting cells.

Identification of a CD8 T cell that can independently mediate autoimmune diabetes development in the complete absence of CD4 T cell helper functions.

Previous work has indicated that an important component for the initiation of autoimmune insulin-dependent diabetes mellitus (IDDM) in the NOD mouse model entails MHC class I-restricted CD8 T cell responses against pancreatic beta cell Ags. However, unless previously activated in vitro, such CD8 T cells have previously been thought to require helper functions provided by MHC class II-restricted CD4 T cells to exert their full diabetogenic effects. In this study, we show that IDDM development is greatly accelerated in a stock of NOD mice expressing TCR transgenes derived from a MHC class I-restricted CD8 T cell clone (designated AI4) previously found to contribute to the earliest preclinical stages of pancreatic beta cell destruction. Importantly, these TCR transgenic NOD mice (designated NOD.AI4alphabeta Tg) continued to develop IDDM at a greatly accelerated rate when residual CD4 helper T cells were eliminated by introduction of the scid mutation or a functionally inactivated CD4 allele. In a previously described stock of NOD mice expressing TCR transgenes derived from another MHC class I-restricted beta cell autoreactive T cell clone, IDDM development was retarded by elimination of residual CD4 T cells. Hence, there is variability in the helper dependence of CD8 T cells contributing to the development of autoimmune IDDM. The AI4 clonotype represents the first CD8 T cell with a demonstrated ability to progress from a naive to functionally activated state and rapidly mediate autoimmune IDDM development in the complete absence of CD4 T cell helper functions.

Biochemical and immunogenetic analysis of an immunodominant peptide (B6dom1) encoded by the classical H7 minor histocompatibility locus.

Of the many minor histocompatibility (H) Ags that have been detected in mice, the ability to induce graft vs host disease (GVHD) after bone marrow transplantation is restricted to a limited number of immunodominant Ags. One such murine Ag, B6dom1, is presented by the H2-Db MHC class I molecule. We present biochemical evidence that the natural B6dom1 peptide is indistinguishable from AAPDNRETF, and we show that this peptide can be isolated from a wide array of tissues, with highest levels from the lymphoid organs and lung. Moreover, we employ a novel, somatic cell selection technique involving CTL-mediated immunoselection coupled with classical genetics, to show that B6dom1 is encoded by the H7 minor H locus originally discovered approximately 40 years ago. These studies provide a molecular genetic framework for understanding B6dom1, and exemplify the fact that mouse minor H loci that encode immunodominant CTL epitopes can correspond to classical H loci originally identified by their ability to confer strong resistance to tumor transplantation. Additionally, these studies demonstrate the utility of somatic cell selection approaches toward resolving H Ag immunogenetics.

Shaping the repertoire of cytotoxic T-lymphocyte responses: explanation for the immunodominance effect whereby cytotoxic T lymphocytes specific for immunodominant antigens prevent recognition of nondominant antigens.

The immunodominance effect, whereby the presence of immunodominant epitopes prevents recognition of nondominant determinants presented on the same antigen-presenting cell (APC) considerably restricts the repertoire of cytotoxic T lymphocyte (CTL) responses. To elucidate the molecular basis of the immunodominance effect, we compared the interactions of a dominant (B6(dom1)) and a nondominant epitope (H-Y) with their restricting class I molecule (H2-Db), and their ability to trigger cognate CTLs. We found that B6(dom1)/Db complexes behaved as optimal T-cell receptor (TCR) ligands and triggered a more rapid in vivo expansion of cognate CTLs than H-Y/Db complexes. The superiority of the dominant epitope was explained by its high cell surface density (1,012 copies/cell for B6(dom1) v 10 copies/cell for H-Y) and its optimal affinity for cognate TCRs. Based on these results, we conclude that dominant class I-associated epitopes are those that have optimal ability to trigger TCR signals in CTLs. We propose that the rapid expansion of CTLs specific for dominant antigens should enable them to compete more successfully than other CTLs for occupancy of the APC surface.

Positional cloning and molecular characterization of an immunodominant cytotoxic determinant of the mouse H3 minor histocompatibility complex.

Immune responses to minor histocompatibility antigens are poorly understood and present substantial barriers to successful solid tissue and bone marrow transplantation among MHC-matched individuals. We exploited a unique positional cloning approach relying on the potent negative selection capability of cytotoxic T cells to identify the H3a gene responsible for immunodominant H2-Db-restricted determinants of the classically defined mouse autosomal H3 complex. The allelic basis for reciprocal H3a antigens is two amino acid changes within a single nonamer H2-Db-binding peptide. The H3a gene, now called Zfp106, encodes a 1888-amino acid protein with three zinc fingers and a beta-transducin domain consistent with DNA/protein binding. A region of ZFP106 is identical to a 600-amino acid sequence implicated in the insulin receptor signaling pathway.

A molecular basis for how a single TCR interfaces multiple ligands.

CD8+ T cells respond to Ags when their clonotypic receptor, the TCR, recognizes nonself peptides displayed by MHC class I molecules. The TCR/ligand interactions are degenerate because, in its life time, the TCR interacts with self MHC class I-self peptide complexes during ontogeny and with self class I complexed with nonself peptides to initiate Ag-specific responses. Additionally, the same TCR has the potential to interact with nonself class I complexed with nonself peptides. How a single TCR interfaces multiple ligands remains unclear. Combinatorial synthetic peptide libraries provide a powerful tool to elucidate the rules that dictate how a single TCR engages multiple ligands. Such libraries were used to probe the requirements for TCR recognition by cloned CD8+ T cells directed against Ags presented by H-2Kb class I molecules. When H-2Kb contact residues were examined, position 3 of the peptides proved more critical than the dominant carboxyl-terminal anchor residue. Thus, secondary anchor residues can play a dominant role in determining the antigenicity of the epitope presented by class I molecules. When the four solvent-exposed potential TCR contact residues were examined, only one or two of these positions required structurally similar residues. Considerable structural variability was tolerated at the remaining two or three solvent-exposed residues of the Kb-binding peptides. The TCR, therefore, requires close physico-chemical complementarity with only a few amino acid residues, thus explaining why TCR/MHC interactions are of low affinity and degenerate.

Major histocompatibility complex class I-restricted T cells are required for all but the end stages of diabetes development in nonobese diabetic mice and use a prevalent T cell receptor alpha chain gene rearrangement.

Nonobese diabetic (NOD) mice develop insulin-dependent diabetes mellitus due to autoimmune T lymphocyte-mediated destruction of pancreatic beta cells. Although both major histocompatibility complex class I-restricted CD8(+) and class II-restricted CD4(+) T cell subsets are required, the specific role each subset plays in the pathogenic process is still unclear. Here we show that class I-dependent T cells are required for all but the terminal stages of autoimmune diabetes development. To characterize the diabetogenic CD8(+) T cells responsible, we isolated and propagated in vitro CD8(+) T cells from the earliest insulitic lesions of NOD mice. They were cytotoxic to NOD islet cells, restricted to H-2Kd, and showed a diverse T cell receptor beta chain repertoire. In contrast, their alpha chain repertoire was more restricted, with a recurrent amino acid sequence motif in the complementarity-determining region 3 loop and a prevalence of Valpha17 family members frequently joined to the Jalpha42 gene segment. These results suggest that a number of the CD8(+) T cells participating in the initial phase of autoimmune beta cell destruction recognize a common structural component of Kd/peptide complexes on pancreatic beta cells, possibly a single peptide.

The molecular and functional characterization of a dominant minor H antigen, H60.

Minor histocompatibility (H) Ags elicit T cell responses and thereby cause chronic graft rejection and graft-vs-host disease among MHC identical individuals. Although numerous independent H loci exist in mice of a given MHC haplotype, certain H Ags dominate the immune response and are thus of considerable conceptual and therapeutic importance. To identify these H Ags and their genes, lacZ-inducible CD8+ T cell hybrids were generated by immunizing C57BL/6 (B6) mice with MHC identical BALB.B spleen cells. The cDNA clones encoding the precursor for the antigenic peptide/Kb MHC class I complex were isolated by expression cloning using the BCZ39.84 T cell as a probe. The cDNAs defined a new H locus (termed H60), located on mouse chromosome 10, and encoded a novel protein that contains the naturally processed octapeptide LTFNYRNL (LYL8) presented by the Kb MHC molecule. Southern blot analysis revealed that the H60 locus was polymorphic among the BALB and the B6 strains. However, none of the H60 transcripts expressed in the donor BALB spleen were detected in the host B6 strain. The expression and immunogenicity of the LYL8/Kb complex in BALB.B and CXB recombinant inbred strains strongly suggested that the H60 locus may account for one of the previously described antigenic activity among these strains. The results establish the source of an immunodominant autosomal minor H Ag that, by its differential transcription in the donor vs the host strains, provides a novel peptide/MHC target for host CD8+ T cells.