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Yan, D.

Publications and source records attributed to Yan, D..

At least 37 records · Page 2Linked to original sources

G-CSF and IL-6 drive myeloid dysregulation during severe viral infection

Dysregulated myeloid states are associated with disease severity in both sepsis and COVID-19. However, their relevance in non-COVID-19 viral infection, the factors driving their induction, and their role in tissue injury remain poorly understood. We performed a meta-analysis of 1,622,180 myeloid cells from 890 COVID-19 or sepsis patients and controls across 19 published blood scRNA-seq datasets, which revealed severity-associated gene programs in both neutrophils and monocytes pointing to emergency myelopoiesis (EM). Using published bulk transcriptional data from 562 individuals with non-COVID-19 viral disease, we show that these signatures are similarly upregulated during severe influenza and RSV infection. Analysis of transcriptional and proteomic responses in tocilizumab-treated COVID-19 patients show that IL-6 signaling blockade results in a partial reduction of EM signatures and a compensatory increase in the growth factor G-CSF. Using a cellular model of human myelopoiesis, we show that both IL-6 and G-CSF stimulate the production of myeloid cells that express EM signatures in vitro. Using a mouse model of severe influenza infection, we demonstrate the effect of IL-6 and G-CSF signaling blockade on EM-associated myeloid cells, and highlight the opposing effects of EM-induced neutrophils and monocytes on tissue injury. Our study demonstrates the link between systemic cytokines and myeloid dysregulation during severe infection in humans, and highlights the cooperative role of IL-6 and G-CSF signaling in driving infection-induced myelopoiesis.

immunology↗

Self-Assembled Origami Neural Probes for Scalable, Multifunctional, Three-Dimensional Neural Interface

Flexible intracortical neural probes have drawn attention for their enhanced longevity in high-resolution neural recordings due to reduced tissue reaction. However, the conventional monolithic fabrication approach has met significant challenges in: (i) scaling the number of recording sites for electrophysiology; (ii) integrating of other physiological sensing and modulation; and (iii) configuring into three-dimensional (3D) shapes for multi-sided electrode arrays. We report an innovative self-assembly technology that allows for implementing flexible origami neural probes as an effective alternative to overcome these challenges. By using magnetic-field-assisted hybrid self-assembly, multiple probes with various modalities can be stacked on top of each other with precise alignment. Using this approach, we demonstrated a multifunctional device with scalable high-density recording sites, dopamine sensors and a temperature sensor integrated on a single flexible probe. Simultaneous large-scale, high-spatial-resolution electrophysiology was demonstrated along with local temperature sensing and dopamine concentration monitoring. A high-density 3D origami probe was assembled by wrapping planar probes around a thin fiber in a diameter of 80[~]105 m using optimal foldable design and capillary force. Directional optogenetic modulation could be achieved with illumination from the neuron-sized micro-LEDs (LEDs) integrated on the surface of 3D origami probes. We could identify angular heterogeneous single-unit signals and neural connectivity 360{degrees} surrounding the probe. The probe longevity was validated by chronic recordings of 64-channel stacked probes in behaving mice for up to 140 days. With the modular, customizable assembly technologies presented, we demonstrated a novel and highly flexible solution to accommodate multifunctional integration, channel scaling, and 3D array configuration.

neuroscience↗

Genetic purging of strongly deleterious mutations underlies black-necked crane's unusual escape from an extinction vortex

Many species are undergoing rapid demographic declines, necessitating an examination of the resulting genetic impacts. The prevailing small population paradigm posits an elevated genetic load and extinction risk. However, instances of fast recovery from severe population bottlenecks suggest alternative outcomes. To investigate this issue, we performed a population genomic analysis on the black-necked crane, analyzing 42 modern and 11 historical genomes. This study revealed substantial evidence of large-effect allele purging underlying the unexpectedly rapid population recovery following an abrupt bottleneck during the 1980s. Nevertheless, forward simulations supposing a prolonged bottleneck (e.g., five generations) predicted a reversion with negative prospects, implying that rapid population recovery served as both the cause and consequence of the species escaping from an extinction vortex. These findings shed light on a potential positive microevolutionary response to current widespread population collapses and underscore the urgency of implementing active and effective conservation strategies to reverse this trend before it becomes irreversible.

evolutionary biology↗

Let-7i-5p regulates osteogenic differentiation of bone marrow mesenchymal stem cells induced by psoralen by targeting CKIP-1

AbstractPrevious studies have shown that psoralen can treat osteoporosis by inducing osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs). CKIP-1 is an inhibitor of bone formation and plays a negative regulatory role in the osteogenic differentiation of BMSCs. MicroRNA (miRNA) is a kind of small single-stranded non-coding RNA, which regulates the occurrence and development process of cell proliferation, apoptosis, differentiation and metabolism. Studies have shown that miRNA can affect the proliferation and differentiation of BMSCs by targeting and regulating the expression of related genes. However, whether microRNA can regulate psoralen-induced osteogenic differentiation and its specific mechanisms remain unclear. The aim of this study was to identify miRNA target genes regulating the osteogenic differentiation of BMSCs induced by psoralen and further analyze the mechanism.miRNA microarray was performed to screen the differentially expressed mirnas induced by psoralen, and bioinformatics analysis and luciferase reporter gene assay were used to determine the target sites of let-7i-5p. The mechanism of let-7i-5p in psoralen induced osteogenic differentiation of BMSCs was investigated using in vitro overexpression or inhibition approaches.The results showed that miRNA microarray analysis and reverse transcription quantitative PCR further confirmed that let-7i-5p was up-regulated during the process of psoralin-induced osteogenic differentiation of BMSCs.Bioinformatics analysis and luciferase reporter gene assay confirmed CKIP-1 as a potential target of let-7i-5p.Overexpression of CKIP-1 by transfection with CKIP-1 mimetic significantly inhibited the expression of BMP, Runx2, and Smad1, while CKIP-1 inhibitor promoted their expression.Overexpression of let-7i-5p partially relieved the inhibitory effect of CKIP-1 on osteogenic differentiation of BMSCs.Our results suggest that let-7i-5p regulates psoralen-induced osteogenic differentiation of BMSCs by targeting CKIP-1, which provides a new therapeutic target for osteoporosis.

genetics↗

Heat Shock Proteins Function as Signaling Molecules to Mediate Neuron-Glia Communication During Aging

The nervous system is primarily composed of neurons and glia, and the communication between them plays profound roles in regulating the development and function of the brain. Neuron-glia signal transduction is known to be mediated by secreted or juxtacrine signals through ligand-receptor interactions on the cell membrane. Here, we report a novel mechanism for neuron-glia signal transduction, wherein neurons transmit proteins to glia through extracellular vesicles, activating glial signaling pathways. We find that in the amphid sensory organ of Caenorhabditis elegans, different sensory neurons exhibit varying aging rates. This discrepancy in aging is governed by the crosstalk between neurons and glia. We demonstrate that early-aged neurons can transmit heat shock proteins (HSP) to glia via extracellular vesicles. These neuronal HSPs activate the IRE1-XBP1 pathway, further increasing their expression in glia, forming a positive feedback loop. Ultimately, the activation of the IRE1-XBP-1 pathway leads to the transcriptional regulation of chondroitin synthases to protect glia-embedded neurons from aging-associated functional decline. Therefore, our studies unveil a novel mechanism for neuron-glia communication in the nervous system and provide new insights into our understanding of brain aging.

neuroscience↗

FFCM-MRF: An accurate and generalizable cerebrovascular segmentation pipeline for humans and rhesus monkeys based on TOF-MRA

PurposeCerebrovascular segmentation and quantification of vascular morphological features on humans and rhesus monkeys are essential for prevention, diagnosis, and treatment of brain diseases. However, current automated whole-brain vessel segmentation methods are often not generalizable to independent datasets, limiting their usefulness in real-world environments with their heterogeneity in participants, scanners, and species. Materials and MethodsIn this study, we proposed an automated, accurate and generalizable segmentation method for magnetic resonance angiography images called FFCM-MRF. This method integrates fast fuzzy c-means clustering and Markov random field optimization using blood vessel shape priors and spatial constraints. We used a total of 123 human and 44 macaque MRA images scanned at 1.5 T, 3 T, and 7 T MRI from 9 datasets to develop and validate the method. ResultsThe average Dice score coefficients for multiple independent datasets were 69.16-89.63%, with the improvements in FFCM-MRF ranged from 0.16-16.14% compared with state-of-the-art machine learning methods. Quantitative analysis showed that FFCM-MRF can accurately segment major arteries in the Circle of Willis at the base of the brain and smaller distal pial arteries while effectively suppressing noise. Test-retest analysis showed that the model yielded high vascular volume and diameter reliability. ConclusionsOur results demonstrate that the proposed method is highly accurate and reliable and largely independent of variations in field strength, scanner platforms, acquisition parameters, and species. The macaque MRA data and user-friendly open-source toolbox are freely available at OpenNeuro and GitHub to facilitate studies of imaging biomarkers for cerebrovascular and neurodegenerative diseases.

neuroscience↗

Pan-cancer characterization and clinical outcome of ecDNA-enhancer mediated transcriptional dysregulation

Extrachromosomal circular DNAs (ecDNAs) are extrachromosomal circular DNA elements harboring enhancers that can function as mobile transcriptional enhancers to promote oncogene amplification in diverse human cancers. However, little is known about the mechanism how ecDNA and relevant enhancers influence the transcription control in different cancers. Here, we performed a comprehensive pan-cancer analysis to explore the potential transcriptional regulatory network between ecDNA and enhancer, and the association with clinical outcomes in different human cancers. Firstly, we identified 18,385 enhancers on ecDNA in 1,921 samples of 20 TCGA tumors. By integrating corresponding RNA-seq data, we found that the expression levels of enhancer RNAs (eRNAs) were significantly different between ecDNA regions and corresponding linear chromosomes. This suggested that enhancers on ecDNA may employ eRNA to perform transcriptional regulation. Furthermore, we identified enhancer co-amplification with oncogenes and found that enhancers on ecDNA significantly affect the expression of co-amplified cancer genes and consequently influence the survival of cancer patients. In addition, we found that, compared to corresponding linear chromosomes, enhancers on ecDNA generally have lower levels of methylation, which may be mainly caused by the enhancer co-amplification with oncogenes. Genome-wide analysis of open chromatin regions revealed significantly higher levels of chromatin accessibility and certain chromosomal preferences in ecDNA regions comparing corresponding linear chromosomes. Moreover, we found that the downregulation of antigen presentation genes and the suppression of antigen presentation pathways may cause the tumor immune evasion mediated by ecDNA. Finally, we identified tumor-infiltrating activated mast cells in ecDNA-positive patients, which are associated with poorer prognosis. In summary, through pan-cancer analysis, we illustrate the characteristics and regulatory mechanisms of ecDNA and its enhancers in cancers, providing insights for future cancer treatment.

bioinformatics↗

Development and optimisation of cationic lipid nanoparticles for mRNA delivery

Messenger RNA (mRNA) has been proposed as a therapeutic agent for various diseases, including cancer. To ensure effective transfection of cancer cells, mRNA needs to be transported with a delivery system that protects its integrity and functionality. In this regard, cationic lipid nanoparticles composed of dioleoylphosphatidylethanolamine (DOPE) and 3{beta}-[N-(N,N-dimethylaminoethane)-carbamoyl] cholesterol (DC-Chol) have emerged as common vectors to deliver mRNA. In this project, we aim to use luciferase mRNA as a reporter to synthesise mRNA-loaded cationic lipid nanoparticles, and optimise their mRNA encapsulation and transfection efficiency in ovarian cancer cells. The optimisation process included: 1) adjusting the lipid formulation; 2) adjusting the input mRNA concentration before lipid nanoparticle extrusion; and 3) adjusting the extrusion methods. After optimisation, the encapsulation efficiency was optimised to 62%, thus achieving a relatively high transfection luminescence signal (9.4 times compared to baseline). The lipid nanoparticles also demonstrated stable physical characteristics and high biocompatibility (above 75% cell viability after treatment) within 24 hours. Overall, this project evaluated the synthesis of DOPE/DC-Chol cationic lipid nanoparticles, and optimised their mRNA encapsulation and transfection efficiency in ovarian cancer cell lines. The optimised lipid nanoparticles can be utilised as an ideal system for mRNA delivery, which could be further developed as a potential platform for the immunotherapy in ovarian cancer.

bioengineering↗

Omicron booster in ancestral strain vaccinated mouse augments protective immunities against both the Delta and Omicron variants

A booster vaccination is called for constraining the evolving epidemic of SARS-CoV-2. However, the necessity of a new COVID-19 vaccine is currently unclear. To compare the effect of an Omicron-matched S DNA vaccine and an ancestral S DNA vaccine in boosting cross-reactive immunities, we firstly immunized mice with two-dose of a DNA vaccine encoding the spike protein of the ancestral Wuhan strain. Then the mice were boosted with DNA vaccines encoding spike proteins of either the Wuhan strain or the Omicron variant. Specific antibody and T cell responses were measured at 4 weeks post boost. Our data showed that the Omicron-matched vaccine efficiently boosted RBD binding antibody and neutralizing antibody responses against both the Delta and the Omicron variants. Of note, antibody responses against the Omicron variant elicited by the Omicron-matched vaccine were much stronger than those induced by the ancestral S DNA vaccine. Meanwhile, CD8+ T cell responses against both the ancestral Wuhan strain and the Omicron strain also tended to be higher in mice boosted by the Omicron-matched vaccine than those in mice boosted with the ancestral S DNA vaccine, albeit no significant difference was observed. Our findings suggest that an Omicron-matched vaccine is preferred for boosting cross-reactive immunities.

immunology↗

Ultra-flexible and Stretchable Intrafascicular Peripheral Nerve Recording Device with Axon-dimension, Cuff-less Microneedle Electrode Array

Peripheral nerve mapping tools with higher spatial resolution are needed to advance systems neuroscience, and potentially provide a closed-loop biomarker in neuromodulation applications. Two critical challenges of microscale neural interfaces are (i) how to apply them to small peripheral nerves, and (ii) how to minimize chronic reactivity. We developed a flexible microneedle nerve array (MINA), which is the first high-density penetrating electrode array made with axon-sized silicon microneedles embedded in low-modulus thin silicone. We present the design, fabrication, acute recording, and chronic reactivity to an implanted MINA. Distinctive units were identified in the rat peroneal nerve. We also demonstrate a long-term, cuff-free, and suture-free fixation manner using rose bengal as a light-activated adhesive for two timepoints. The tissue response at 1-week included a sham (N=5) and MINA-implanted (N=5) group, and the response at 6-week also included a sham (N=3) and MINA-implanted (N=4) group. These conditions were quantified in the left vagus nerve of rats using histomorphometry. Micro-CT was added to visualize and quantify tissue encapsulation around the implant. MINA demonstrated a reduction in encapsulation thickness over previously quantified interfascicular methods. Future challenges include techniques for precise insertion of the microneedle electrodes and demonstrating long-term recording.

neuroscience↗

Novel inhibitors of E. coli lipoprotein diacylglyceryl transferase are insensitive to resistance caused by lpp deletion

Lipoprotein diacylglyceryl transferase (Lgt) catalyzes the first step in the biogenesis of Gram-negative bacterial lipoproteins which play crucial roles in bacterial growth and pathogenesis. We demonstrate that Lgt depletion in a clinical uropathogenic Escherichia coli strain leads to permeabilization of the outer membrane and increased sensitivity to serum killing and antibiotics. Importantly, we identify the first ever described Lgt inhibitors that potently inhibit Lgt biochemical activity in vitro and are bactericidal against wild-type Acinetobacter baumannii and E. coli strains. Unlike inhibition of other steps in lipoprotein biosynthesis, deletion of the major outer membrane lipoprotein, lpp, is not sufficient to rescue growth after Lgt depletion or provide resistance to Lgt inhibitors. Our data validate Lgt as a novel druggable antibacterial target and suggest that inhibition of Lgt may not be sensitive to one of the most common resistance mechanisms that invalidate inhibitors of downstream steps of bacterial lipoprotein biosynthesis and transport.

microbiology↗

Regulation of lipid accumulation-induced ROS in myeloid-derived suppressor cells via targeting fatty-acid transport protein 2 enhanced anti-PD-L1 tumor immunotherapy

Despite the remarkable success and efficacy of immune checkpoint blockade (ICB) therapy against the PD-1/PD-L1 axis, it induces sustained responses in a sizeable minority of cancer patients due to the activation of immunosuppressive factors such as myeloid-derived suppressor cells (MDSCs). Inhibiting the immunosuppressive function of MDSCs is critical for successful cancer ICB therapy. Interestingly, lipid metabolism is a crucial factor in modulating MDSCs function. Fatty acid transport protein 2 (FATP2) conferred the function of PMN-MDSCs in cancer via the upregulation of arachidonic acid metabolism. However, whether regulating lipid accumulation in MDSCs by targeting FATP2 could block MDSCs reactive oxygen species (ROS) production and enhance PD-L1 blockade-mediated tumor immunotherapy remains unexplored. Here we report that FATP2 regulated lipid accumulation, ROS, and immunosuppressive function of MDSCs in tumor-bearing mice. Tumor cells-derived granulocyte macrophage-colony stimulating factor (GM-CSF) induced FATP2 expression in MDSCs by activation of STAT3 signaling pathway. Pharmaceutical blockade of FATP2 expression in MDSCs by lipofermata decreased lipid accumulation, reduced ROS, blocked immunosuppressive activity, and consequently inhibited tumor growth. More importantly, lipofermata inhibition of FATP2 in MDSCs enhanced anti-PD-L1 tumor immunotherapy via the upregulation of CD107a and reduced PD-L1 expression on tumor-infiltrating CD8+ T-cells. Furthermore, the combination therapy blocked MDSCs suppressive role on T-cells thereby enhanced T-cells ability for the production of IFN-{gamma}. These findings indicate that FATP2 plays a key role in modulating lipid-induced ROS in MDSCs and targeting FATP2 in MDSCs provides a novel therapeutic approach to enhance anti-PD-L1 cancer immunotherapy. Research highlightsO_LIFATP2 inhibition in MDSCs blocked ROS-mediated immunosuppressive function and promoted MDSCs differentiation to immune-stimulatory phenotype. C_LIO_LIGM-CSF mediated FATP2 expression via activation of STAT3 signaling leading to lipid accumulation-induced ROS in MDSCs. C_LIO_LIFATP2 inhibition enhanced anti-PD-L1 tumor immunotherapy by reducing MDSCs immunosuppressive activity in blocking T-cells ability to produce IFN-{gamma}. C_LIO_LICombination of anti-PD-L1 antibody and FATP2 inhibitor decreased MDSCs accumulation and PD-L1 surface expression in the spleens of LLC-bearing mice. C_LIO_LICombined treatment of FATP2 inhibitor and PD-L1 blockade-mediated tumor immunotherapy enhanced tumor-infiltrating CD8+ T cells activation via increased CD107a and reduced PD-L1 surface expression. C_LI

immunology↗

SpRY greatly expands the genome editing scope in rice with highly flexible PAM recognition

Plant genome engineering mediated by various CRISPR-based tools requires specific protospacer adjacent motifs (PAMs), such as the well-performed NGG, NG, NNG, etc., to initiate the target recognition, which notably restricts the editable range of the plant genome. In this study, we thoroughly investigated the nuclease activity and the PAM preference of two structurally-engineered SpCas9 variants (SpG and SpRY) in transgenic rice. Our study shows that SpG nuclease favors NGD PAMs, albeit less efficiently than the previously described SpCas9-NG and that SpRY nuclease achieves efficient editing across a wide range of genomic loci, exhibiting a preference of NGD as well as NAN PAMs. Furthermore, SpRY-fused cytidine deaminase hAID*{Delta} and adenosine deaminase TadA8e were generated, respectively. These constructs efficiently induced C-to-T and A-to-G conversions in the target genes toward various non-canonical PAMs, including non-G PAMs. Remarkably, high-frequency self-editing events (indels and DNA fragments deletion) in the integrated T-DNA fragments as a result of the nuclease activity of SpRY were observed, whereas the self-editing of SpRY nickase-mediated base editor was quite low in transgenic rice lines. In conclusion, the broad PAM compatibility of SpRY greatly expands the targeting scope of CRISPR-based tools in plant genome engineering.

plant biology↗

Dynamic Configuration of Coactive Micropatterns in the Default Mode Network during Wakefulness and Sleep

The default mode network (DMN) is a prominent intrinsic network that is observable in many mammalian brains. However, few studies have investigated the temporal dynamics of this network based on direct physiological recordings. Herein, we addressed this issue by characterizing the dynamics of local field potentials (LFPs) from the rat DMN during wakefulness and sleep with an exploratory analysis. We constructed a novel coactive micropattern (CAMP) algorithm to evaluate the configurations of rat DMN dynamics and further revealed the relationship between DMN dynamics with different wakefulness and alertness levels. From the gamma activity (40-80 Hz) in the DMN across wakefulness and sleep, three spatially stable CAMPs were detected: a common low-activity level micropattern (cDMN), an anterior high-activity level micropattern (aDMN) and a posterior high-activity level micropattern (pDMN). A dynamic balance across CAMPs emerged during wakefulness and was disrupted in sleep stages. In the slow-wave sleep (SWS) stage, cDMN became the primary activity pattern, whereas aDMN and pDMN were the major activity patterns in the rapid eye movement sleep (REM) stage. Additionally, further investigation revealed phasic relationships between CAMPs and the up-down states of the slow DMN activity in the SWS stage. Our study revealed that the dynamic configurations of CAMPs were highly associated with different stages of wakefulness and provided a potential three-state model to describe the DMN dynamics for wakefulness and alertness. Impact StatementIn the current study, a novel coactive micropattern method (CAMP) was developed to elucidate fast DMN dynamics during wakefulness and sleep. Our findings demonstrated that the dynamic configurations of DMN activity are specific to different wakefulness stages and provided a three-state DMN CAMP model to depict wakefulness levels, thus revealing a potentially new neurophysiological representation of alertness levels. This work could elucidate the DMN dynamics underlying different stages of wakefulness and have important implications for the theoretical understanding of the neural mechanism of wakefulness and alertness.

bioinformatics↗

In vivo fluorescence imaging with a flat, lensless microscope

Fluorescence imaging over large areas of the brain in freely behaving animals would allow researchers to better understand the relationship between brain activity and behavior; however, traditional microscopes capable of high spatial resolution and large fields of view (FOVs) require large and heavy lenses that restrict animal movement. While lensless imaging has the potential to achieve both high spatial resolution and large FOV with a thin lightweight device, lensless imaging has yet to be achieved in vivo due to two principal challenges: (a) biological tissue typically has lower contrast than resolution targets, and (b) illumination and filtering must be integrated into this non-traditional device architecture. Here, we show that in vivo fluorescence imaging is possible with a thin lensless microscope by optimizing the phase mask and computational reconstruction algorithms, and integrating fiber optic illumination and thin-film color filters. The result is a flat, lensless imager that achieves better than 10 m spatial resolution and a FOV that is 30x larger than other cellular resolution miniature microscopes.

neuroscience↗

Circadian Rhythm Is Disrupted by ZNF704 in Breast Carcinogenesis

Copy number gain in chromosome 8q21 is considered as the prototype of genetic abnormalities associated with development of breast cancer, yet the oncogenic potential underlying this amplicon in breast carcinogenesis remains to be delineated. We report here that ZNF704, a gene mapped to 8q21, is recurrently amplified in various malignancies including breast cancer. We found that ZNF704 acts as transcription repressor and interacts with the transcription corepressor SIN3A complex. Genome-wide interrogation of the transcriptional targets identifies that the ZNF704/SIN3A complex represses a panel of genes including PER2 that are critically involved in the function of circadian clock. Indeed, ZNF704 overexpression prolongs the period and dampens the amplitude of circadian clock. We showed that ZNF704 promotes the proliferation and invasion of breast cancer cells in vitro and accelerates the growth and metastasis of breast cancer in vivo. Consistently, the level of ZNF704 expression is inversely correlated with that of PER2 in breast carcinomas, and high level of ZNF704 correlates with advanced histological grades, lymph node positivity, and poor prognosis of breast cancer patients, especially those with HER2+ and basal-like subtypes. These results indicate that ZNF704 is an important regulator of circadian clock and a potential driver for breast carcinogenesis.

cancer biology↗

Large-scale transgenic Drosophila resource collections for loss- and gain-of-function studies

The Transgenic RNAi Project (TRiP), a Drosophila functional genomics platform at Harvard Medical School, was initiated in 2008 to generate and distribute a genome-scale collection of RNAi fly stocks. To date, the TRiP has generated >15,000 RNAi fly stocks. As this covers most Drosophila genes, we have largely transitioned to development of new resources based on CRISPR technology. Here, we present an update on our libraries of publicly available RNAi and CRISPR fly stocks, and focus on the TRiP-CRISPR overexpression (TRiP-OE) and TRiP-CRISPR knockout (TRiP-KO) collections. TRiP-OE stocks express sgRNAs targeting upstream of a gene transcription start site. Gene activation is triggered by co-expression of catalytically dead Cas9 (dCas9) fused to an activator domain, either VP64-p65-Rta (VPR) or Synergistic Activation Mediator (SAM). TRiP-KO stocks express one or two sgRNAs targeting the coding sequence of a gene or genes, allowing for generation of indels in both germline and somatic tissue. To date, we have generated more than 5,000 CRISPR-OE or -KO stocks for the community. These resources provide versatile, transformative tools for gene activation, gene repression, and genome engineering.

genetics↗

Transcriptomic profiling of plaque psoriasis and cutaneous T cell subsets during treatment with secukinumab

The IL17A inhibitor secukinumab is efficacious for the treatment of psoriasis. In order to define its mechanism of action, it is important to understand its impact on psoriatic whole skin tissue as well as specific skin-resident immune cell populations such as T lymphocytes. In this study, we treated 15 moderate-to-severe plaque psoriasis patients with secukinumab and characterized the longitudinal transcriptomic changes of whole lesional skin tissue and cutaneous CD4+ T effector cells (Teffs), CD4+ T regulatory cells (Tregs), and CD8+ T effector cells during 12 weeks of treatment. Secukinumab was clinically effective, with 100%, 47%, and 27% of patients in the study achieving PASI75, PASI90, and PASI100 by week 12, respectively. At baseline prior to treatment, we observed that IL17A overexpression predominates in psoriatic CD8+ T cells rather than Teffs, supporting the importance of IL-17-secreting CD8+ T cells (Tc17) compared to IL-17-secreting CD4+ T cells (Th17) cells in the pathogenesis of psoriasis. Although secukinumab targets only IL17A, we observed rapid reduction of IL17A, IL17F, IL23A, IL23R, and IFNG expression in lesional skin as soon as 2 weeks after initiation of treatment and normalization of expression by week 12. Secukinumab treatment resulted in resolution of 89-97% of psoriasis-associated expression differences in both bulk tissue and T cell subsets by week 12 of treatment. Overall, secukinumab appears to rapidly reverse many of the molecular hallmarks of psoriasis.

genomics↗