Histone/Protein Deacetylase 11 Targeting Promotes Foxp3+ Treg Function.

Current interest in Foxp3+ T-regulatory (Treg) cells as therapeutic targets in transplantation is largely focused on their harvesting pre-transplant, expansion and infusion post-transplantation. An alternate strategy of pharmacologic modulation of Treg function using histone/protein deacetylase inhibitors (HDACi) may allow more titratable and longer-term dosing. However, the effects of broadly acting HDACi vary, such that HDAC isoform-selective targeting is likely required. We report data from mice with constitutive or conditional deletion of HDAC11 within Foxp3+ Treg cells, and their use, along with small molecule HDAC11 inhibitors, in allograft models. Global HDAC11 deletion had no effect on health or development, and compared to WT controls, Foxp3+ Tregs lacking HDAC11 showed increased suppressive function, and increased expression of Foxp3 and TGF-β. Likewise, compared to WT recipients, conditional deletion of HDAC11 within Tregs led to long-term survival of fully MHC-mismatched cardiac allografts, and prevented development of transplant arteriosclerosis in an MHC class II-mismatched allograft model. The translational significance of HDAC11 targeting was shown by the ability of an HDAC11i to promote long-term allograft allografts in fully MHC-disparate strains. These data are powerful stimuli for the further development and testing of HDAC11-selective pharmacologic inhibitors, and may ultimately provide new therapies for transplantation and autoimmune diseases

T-regulatory cells require Sin3a for stable expression of Foxp3

Histone deacetylases 1 and 2 play a major role in the transcriptional regulation of T-regulatory (Treg) cells via interactions with a myriad of coregulatory factors. Sin3a has been well established as a Hdac1/2 cofactor, while its role within Tregs has not been established. In this study, the effects of conditional deletion of Sin3a within Foxp3+ Tregs were evaluated. Developmental deletion of Sin3a from Foxp3+ Tregs resulted in the rapid onset of fatal autoimmunity. Treg numbers were greatly reduced, while residual Tregs had impaired suppressive function. Mice also showed effector T-cell activation, autoantibody production, and widespread tissue injury. Mechanistically, Sin3a deletion resulted in decreased transcription of Foxp3 with a complete lack of CNS2 CpG demethylation. In addition, Foxp3 protein stability was impaired with an increased ex-Treg population. Thus, Sin3a plays a critical role in the maintenance of Treg identity and function and is essential for the expression and stability of Foxp3

Two Lysines in the Forkhead Domain of Foxp3 Are Key to T Regulatory Cell Function

Background: The forkhead box transcription factor, Foxp3, is master regulator of the development and function of CD4+CD25+ T regulatory (Treg) cells that limit autoimmunity and maintain immune homeostasis. The carboxyl-terminal forkhead (FKH) domain is required for the nuclear localization and DNA binding of Foxp3. We assessed how individual FKH lysines contribute to the functions of Foxp3 in Treg cells. Methodology/Principal Findings: We found that mutation of FKH lysines at position 382 (K17) and at position 393 (K18) impaired Foxp3 DNA binding and inhibited Treg suppressive function in vivo and in vitro. These lysine mutations did not affect the level of expression of Foxp3 but inhibited IL-2 promoter remodeling and had important and differing effects on Treg-associated gene expression. Conclusions/Significance: These data point to complex effects of post-translational modifications at individual lysines within the Foxp3 FKH domain that affect Treg function. Modulation of these events using small molecule inhibitors ma

25th annual computational neuroscience meeting: CNS-2016

The same neuron may play different functional roles in the neural circuits to which it belongs. For example, neurons in the Tritonia pedal ganglia may participate in variable phases of the swim motor rhythms [1]. While such neuronal functional variability is likely to play a major role the delivery of the functionality of neural systems, it is difficult to study it in most nervous systems. We work on the pyloric rhythm network of the crustacean stomatogastric ganglion (STG) [2]. Typically network models of the STG treat neurons of the same functional type as a single model neuron (e.g. PD neurons), assuming the same conductance parameters for these neurons and implying their synchronous firing [3, 4]. However, simultaneous recording of PD neurons shows differences between the timings of spikes of these neurons. This may indicate functional variability of these neurons. Here we modelled separately the two PD neurons of the STG in a multi-neuron model of the pyloric network. Our neuron models comply with known correlations between conductance parameters of ionic currents. Our results reproduce the experimental finding of increasing spike time distance between spikes originating from the two model PD neurons during their synchronised burst phase. The PD neuron with the larger calcium conductance generates its spikes before the other PD neuron. Larger potassium conductance values in the follower neuron imply longer delays between spikes, see Fig. 17.Neuromodulators change the conductance parameters of neurons and maintain the ratios of these parameters [5]. Our results show that such changes may shift the individual contribution of two PD neurons to the PD-phase of the pyloric rhythm altering their functionality within this rhythm. Our work paves the way towards an accessible experimental and computational framework for the analysis of the mechanisms and impact of functional variability of neurons within the neural circuits to which they belong

Modeling and Parameter Design of Voltage-Controlled Inverters Based on Discrete Control

Grid-connected inverters are widely used to interface renewable energy and energy storage resources into the grid. Voltage-controlled inverters have attracted more and more attention due to their grid-friendly characteristics. The mathematical models of the voltage and current loops are developed in this paper, considering especially the discrete control delay caused by calculation and modulation. In order to suppress the resonance peak in the current loop, the frequency characteristics of the current loop are analyzed in detail. The optimum design flow of the current controller and voltage controller parameters are presented based on numerical analysis, and the stability, dynamic performance and the resonance peak suppression in voltage loop are also considered. Finally, the validity of the mathematical model and the effectiveness of the controller parameters design method are verified by simulation and experimental results

Improvement of Temperature Performance of Singlemode-Multimode-Singlemode Fiber Structure

A theoretical model for studying the temperature properties of singlemode-multimode-singlemode (SMS) fiber structure fabricated by absorptive multimode fiber (MMF) cladding is established. Moreover, an SMS-based temperature sensor is fabricated and experimentally demonstrated. Experimental results show that the dip wavelength of the transmission spectrum changes linearly with temperature, which is in good agreement with the simulated results obtained by using the model. Further, a comprehensive study of temperature characteristics affected by the thermo-optic effect, thermal expansion effect, and thermal effect of absorption characteristics is performed for SMS fiber optic structures with different refractive indexes, thermo-optic coefficients, and absorption properties of MMF cladding, MMF core diameters, and thermal expansion coefficients of packaging shell. According to the obtained rules, investigations are carried out into the thermal response of an SMS fiber structure resulting from combined thermal effects for temperature performance optimization. Excellent temperature stability with a temperature sensitivity of 0 pm/°C or good temperature sensitivity of −441.58 pm/°C is achieved accordingly

Foxp3 mutants impair Foxp3 DNA binding ability.

<p>293T cells were transfected with EV, WT Foxp3, K17R or K18R without or with p300 expression vectors, and 48 h later, cell lysates were harvested. (<b>A</b>) Equal amounts of cell lysates were incubated with biotin-labeled Foxp3 binding site nucleotide, and Foxp3 DNA binding was detected with anti-Foxp3 or anti-acetyl-lysine Abs. The protein expression levels of Foxp3 and loading control β-actin were detected by western blotting; arrow indicates acetylated Foxp3 bound to DNA, and star indicates non-specific binding. (<b>B–D</b>) The densities of Foxp3 DNA-binding bands were measured using Image-J software and normalized with Foxp3 input levels. (<b>B</b>) The relative Foxp3 DNA binding ability in the absence of p300 is shown. (<b>C</b>) Foxp3 and mutant DNA binding ability was increased in the presence of p300. (<b>D</b>) Comparison of relative Foxp3 DNA binding between WT and mutants in the presence of p300 is shown. (<b>E</b>) Comparison of relative acetylated Foxp3 binding level between WT and mutants is shown. Results are representative of 2 independent experiments.</p

Foxp3 mutants impair Treg function in vivo.

<p>(<b>A</b>) 1×10⁶ Thy1.1+ CD4+CD25− T cells were co-transferred with 1×10⁶ Thy1.2+ CD4+ T cells transduced with WT Foxp3, K16-19R, K17R, K18R or EV, or with purified normal B6 Treg cells, into Rag1−/− mice. At 7 d post-transfer, single-cell suspensions from lymph node or spleen samples were stained for FACS analyses; the numbers (<b>A</b>) or percentages (<b>B</b>) of CD4+ Thy1.1+ cells are shown. Results are representative of 2 independent experiments, and *p<0.05 compared to WT Foxp3.</p

Single Foxp3 lysine mutations affect Treg suppressive function and gene expression.

<p>(A) CD4+ CD25− T cells transduced with retroviruses encoding WT Foxp3, K16R, K17R, K18R, K19R or EV; Foxp3 staining showed >80% transduction efficiency. (B) Effects of single lysine mutations on Treg suppressive activity. (C) RNA derived from CD4+CD25− T cells transduced with WT Foxp3, K17R, K18R or EV were analyzed for CTLA4 (in the absence of CD3/CD28 mAbs) and IL-2, IL-4, IL-17 and IL-21 (in the presence of CD3/CD28 mAbs) gene expression by qPCR. Data were normalized to 18S; *p<0.05, **p<0.01 compared to WT Foxp3. Graphs show means ± SD and results are representative of 3 independent experiments.</p

Foxp3 lysine mutations affect Treg suppressive function and gene expression.

<p>(A) Comparison of amino acid sequences of the C-terminal regions of mouse (m) and human (h) Foxp3, with FKH domains in gray. Lysine at position 332 is K16 and the FKH domain contains K17–K21 (consecutive lysines numbered in green), with residues important for Runx1 (R), DNA (D) or NFAT (N) binding indicated in purple and non-conserved residues shown in red; adapted from <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0029035#pone.0029035-Tao1" target="_blank">[16]</a>. (B) CD4⁺CD25⁻ T cells were transduced with retroviruses encoding WT Foxp3, Foxp16-19R, Foxp16-19Q or EV; Foxp3 staining showed >85% transduction efficiency (%transduced cells shown in blue in each panel). (C) In vitro Treg suppression assays in which 5×10⁵ CFSE-labeled Teff cells were stimulated for 72 h with CD3 mAb in the presence of 5×10⁵ irradiated APC and the indicated ratios of Treg to Teff cells. Data are mean ± SD of duplicate measurements of the percentages of dividing Teff cells, and results are representative of 3 independent experiments; *p value<0.05, **p<0.01 compared to empty vector (EV) in left panel or compared to WT Foxp3 in right panel. (D) RNA derived from CD4⁺CD25⁻ T cells transduced with WT Foxp3, Foxp3 K16-19R, K16-19Q or EV were analyzed for CTLA4, GITR, IL-2, and IL-10 gene expression by qPCR and data were normalized to 18S; *p value<0.05, **p<0.01 compared to WT Foxp3. Graphs show means ± SD and results are representative of 3 independent experiments.</p