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A thorough understanding of the cell behaviors of the human neural grafts is fundamental to exploit them to achieve cell therapy for recovering brain functions. Here by using immunohistological staining, we trace the cell fate of the intrastriatal human neural progenitor cell (NPC) grafts up to 9 months in adult rats, with multiple examining time points to provide a unified working time frame for future transplantation study. Lots of Nestin+/Sox2+ human cells continuously migrate along the white matter tracts into distal brain parenchyma even long time after transplantation, providing a potential for curing diffuse brain damage. Further analysis reveals a significant heterogeneity of the long-term sustained neural stem cells (NSC)/NPCs that progressing throughout different stages, mimicking the neural development of human forebrain. More importantly, the initial GFAP expression in human grafts marks the NSC progression instead of terminal astrocyte differentiation. The distally migrating human cells continuously show the capability to produce new neurons, albeit at a low efficiency in the intact brain. Further investigations in neural disease models are needed. Such study would benefit neural cell therapy with regarding to the optimization of the transplantation strategy and choosing of acting mode by neural grafts (e.g. via cell replacement orex vivogene therapy).

More than 30 % of individuals with epilepsy are refractory to currently available drugs, highlighting the urgent need to develop novel candidate drugs. Accumulating evidence implicates the key role of ferroptosis in the pathophysiology of epileptic seizuresand its potential as a new drug target. Drug repurposing is a promising strategy for the rapid generation of new candidate drugs from the market drugs with new therapeutic indications, such as the best-selling drug thalidomide. Herein, we reported the discovery of Seratrodast, a market drug of thromboxane A2 receptor antagonist as a new ferroptosis inhibitor (IC

Background and purpose Anti-N-methyl-D-aspartate receptor (NMDAR) encephalitis is a severe central nervous system disorder mediated by NMDAR antibodies that damages neurons. We investigated the correlation between cytoskeletal autoantibodies and the clinical severity in patients with anti-NMDAR encephalitis. Methods Non-NMDAR autoantibodies were identified by screening matched cerebrospinal fluid (CSF) and the serum samples of 45 consecutive patients with anti-NMDAR encephalitis and 60 healthy individuals against N-methyl-D-aspartate receptor 1-transfected and nontransfected human embryonic kidney 293T cells. Immunocytochemistry was performed to assess antibody binding in rat brain sections and primary cortical neurons. Cell-based assays and Western blotting were applied to identify autoantibodies targeting medium neurofilaments (NFMs). We compared clinical characteristics between patients with NMDAR encephalitis who were positive and negative for anti-NFM-autoantibodies. Results Anti-NFM autoantibodies were detected in both the serum and CSF in one patient (2%) and in the serum only in six patients (13%). No antibodies were detected in the serum of healthy controls (7/45 vs. 0/60, p=0.0016). Four of the seven patients with anti-NFM autoantibodies in serum were children (57%), and three (43%) had abnormalities in brain magnetic resonance imaging. These patients responded well to immunotherapy, and either no significant or only mild disability was observed at the last follow-up. Anti-NMDAR encephalitis did not differ with the presence of anti-NFM autoantibodies. Conclusions Anti-NFM autoantibodies may be present in patients with anti-NMDAR encephalitis, indicating underlying neuronal damage. A large cohort study is warranted to investigate the clinical differences between patients with NMDAR encephalitis according to their anti-NFM antibody status.

Neurogenesis, a highly orchestrated process, entails the transition from a pluripotent to neural state and involves neural progenitor cells (NPCs) and neuronal/glial subtypes. However, the precise epigenetic mechanisms underlying fate decision remain poorly understood. Here, we delete KDM6s (JMJD3 and/or UTX), the H3K27me3 demethylases, in human embryonic stem cells (hESCs) and show that their deletion does not impede NPC generation from hESCs. However, KDM6-deficient NPCs exhibit poor proliferation and a failure to differentiate into neurons and glia. Mechanistically, both JMJD3 and UTX are found to be enriched in gene loci essential for neural development in hNPCs, and KDM6 impairment leads to H3K27me3 accumulation and blockade of DNA accessibility at these genes. Interestingly, forced expression of neuron-specific chromatin remodelling BAF (nBAF) rescues the neuron/glia defect in KDM6-deficient NPCs despite H3K27me3 accumulation. Our findings uncover the differential requirement of KDM6s in specifying NPCs and neurons/glia and highlight the contribution of individual epigenetic regulators in fate decisions in a human development model., Neurogenesis is an ordered transition from pluriptotent cells to neural precursor cells (NPCs) to neurons. Here the authors show that loss of the lysine demethylases JMJD3 and UTX leads reduced DNA accessibility at neurogenesis loci in human NPCs, and that the chromatin remodeller BAF can rescue differentiation defects.

Transdifferentiation of other cell type into human neuronal cells (hNCs) provides a platform for neural disease modeling, drug screening and potential cell-based therapies. Among all of the cell donor sources, human urine cells (hUCs) are convenient to obtain without invasive harvest procedure. Here, we report a novel approach for the transdifferentiation of hUCs into hNCs. Our study demonstrated that a combination of seven small molecules (CAYTFVB) cocktail induced transdifferentiation of hUCs into hNCs. These chemical-induced neuronal cells (CiNCs) exhibited typical neuron-like morphology and expressed mature neuronal markers. The neuronal-like morphology revealed in day 1, and the Tuj1-positive CiNCs reached to about 58% in day 5 and 38.36% Tuj1+/MAP2+ double positive cells in day 12. Partial electrophysiological properties of CiNCs was obtained using patch clamp. Most of the CiNCs generated using our protocol were glutamatergic neuron populations, whereas motor neurons, GABAergic or dopaminergic neurons were merely detected. hUCs derived from different donors were converted into CiNCs in this work. This method may provide a feasible and noninvasive approach for reprogramming hNCs from hUCs for disease models and drug screening.

Polycomb repressive complexes (PRCs) are essential in mouse gastrulation and specify neural ectoderm in human embryonic stem cells (hESCs), but the underlying molecular basis remains unclear. Here in this study, by employing an array of different approaches, such as gene knock-out, RNA-seq, ChIP-seq, et al., we uncover that EZH2, an important PRC factor, specifies the normal neural fate decision through repressing the competing meso/endoderm program. EZH2−/− hESCs show an aberrant re-activation of meso/endoderm genes during neural induction. At the molecular level, EZH2 represses meso/endoderm genes while SOX2 activates the neural genes to coordinately specify the normal neural fate. Moreover, EZH2 also supports the proliferation of human neural progenitor cells (NPCs) through repressing the aberrant expression of meso/endoderm program during culture. Together, our findings uncover the coordination of epigenetic regulators such as EZH2 and lineage factors like SOX2 in normal neural fate decision. Supplementary Information The online version contains supplementary material available at 10.1186/s13619-021-00092-6.

BackgroundThere is a huge controversy about whether xenograft or allograft in the “immune-privileged” brain needs immunosuppression. In animal studies, the prevailing sophisticated use of immunosuppression or immunodeficient animal is detrimental for the recipients, which results in a short lifespan of animals, confounds functional behavioral readout of the graft benefits, and discourages long-term follow-up.MethodsNeuron-restricted neural progenitor cells (NPCs) were derived from human embryonic stem cells (ESCs, including H1, its gene-modified cell lines for better visualization, and HN4), propagated for different passages, and then transplanted into the brain of immunocompetent rats without immunosuppressants. The graft survivals, their cell fates, and HLA expression levels were examined over time (up to 4 months after transplantation). We compared the survival capability of NPCs from different passages and in different transplantation sites (intra-parenchyma vs. para- and intra-cerebroventricle). The host responses to the grafts were also investigated.ResultsOur results show that human ESC-derived neuron-restricted NPCs survive extendedly in adult rat brain parenchyma with no need of immunosuppression whereas a late-onset graft rejection seems inevitable. Both donor HLA antigens and host MHC-II expression level remain relatively low with little change over time and cannot predict the late-onset rejection. The intra-/para-cerebroventricular human grafts are more vulnerable to the immune attack than the intrastriatal counterparts. Prevention of graft hyperplasia by using hypoproliferative late passaged human NPCs further significantly extends the graft survival time. Our new data also shows that a subpopulation of host microglia upregulate MHC-II expression in response to the human graft, but fail to present the human antigen to the host immune system, suggestive of the immune-isolation role of the blood–brain barrier (BBB).ConclusionsThe present study confirms the “immune privilege” of the brain parenchyma and, more importantly, unveils that choosing hypoproliferative NPCs for transplantation can benefit graft outcome in terms of both lower tumor-genic risk and the prolonged survival time without immunosuppression.

Hematopoietic stem cells (HSCs)/progenitor cells (HPCs) are generated from hemogenic endothelial cells (HECs) during the endothelial-to-hematopoietic transition (EHT); however, the underlying mechanism remains poorly understood. Here, using an array of approaches, including CRSPR/Cas9 gene knockouts, RNA-Seq, ChIP-Seq, ATAC-Seq etc., we report that vitamin C (Vc) is essential in HPC generation during human pluripotent stem cell (hPSC) differentiation in defined culture conditions. Mechanistically, we found that the endothelial cells generated in the absence of Vc fail to undergo the EHT because of an apparent failure in opening up genomic loci essential for hematopoiesis. Under Vc deficiency, these loci exhibited abnormal accumulation of histone H3 trimethylation at Lys-27 (H3K27me3), a repressive histone modification that arose because of lower activities of demethylases that target H3K27me3. Consistently, deletion of the two H3K27me3 demethylases, Jumonji domain-containing 3 (JMJD3 or KDM6B) and histone demethylase UTX (UTX or KDM6A), impaired HPC generation even in the presence of Vc. Furthermore, we noted that Vc and jmjd3 are also important for HSC generation during zebrafish development. Together, our findings reveal an essential role for Vc in the EHT for hematopoiesis, and identify KDM6-mediated chromatin demethylation as an important regulatory mechanism in hematopoietic cell differentiation.

Class IIa histone deacetylases (HDACs) are a subfamily of HDACs with important functions in development and adult tissue homeostasis. As opposed to other HDACs, they lack catalytic function and bind transcription factors to recruit transcriptional co-regulators, mostly co-repressors such as nuclear receptor co-repressor (NCoR)/silencing mediator of retinoid and thyroid hormone receptor (SMRT). Class IIa HDACs enhance mouse somatic cell reprogramming to induced pluripotent stem cells (iPSCs) by repressing the function of the pro-mesenchymal transcription factor myocyte enhancer factor 2 (MEF2), which is upregulated during this process. Here, we describe, using HDAC4 and 7 as examples, that class IIa HDACs exhibit nuclear-cytoplasmic trafficking in reprogramming, being mostly cytoplasmic in donor fibroblasts and intermediate cells but translocating to the nucleus in iPSCs. Importantly, over-expressing a mutant form of HDAC4 or 7 that becomes trapped in the nucleus enhances the early phase of reprogramming but is deleterious afterwards. The latter effect is mediated through binding to the exogenous reprogramming factors at pluripotency loci, and the subsequent recruitment of NCoR/SMRT co-repressors. Thus, our findings uncover a context-dependent function of class IIa HDACs in reprogramming and further reinforce the idea that recruitment of co-repressors by the exogenous factors is a major obstacle for reactivating the pluripotency network in this process. Keywords: Somatic cell reprogramming, iPSCs, Class IIa HDACs, NCoR/SMRT co-repressors, Mesenchymal-to-epithelial transition, Pluripotency

Retrograde monosynaptic tracing with EnvA-pseudotyped rabies virus has been employed to identify the afferent and efferent connectivity of transplanted human embryonic stem (hES) cell-derived neurons in animal models. Due to the protracted development of transplanted human neurons in host animals, it is important that those transplanted cells express avian leukosis and sarcoma virus subgroup A receptor (TVA) and rabies glycoprotein G (Rgp) for a period of up to several months to enable identification of the synaptic inputs from host neurons to grafted neurons through this rabies virus-based method. Here, we report the generation of an engineered hES cell line through CRISPR/Cas9-mediated targeting to the AAVS1 locus of an EnvA-pseudotyped rabies virus-based tool for retrograde monosynaptic tracing. This engineered hES cell line, named H1-CAG-GTRgp, expresses GFP, TVA and Rgp. Upon transplantation of H1-CAG-GTRgp-derived neural progenitor cells (NPCs) into the rat brain after traumatic injury, the grafted neurons derived from H1-CAG-GTRgp cells expressed GFP, TVA, and Rgp stably for up to 6 months post-transplantation and received robust synaptic inputs from host neurons in the target regions of the orthotopic neural circuitry. The retrograde monosynaptic tracing hES cell line provides an efficient approach to analyze transplant connectivity for the comprehensive assessment of host-donor cell innervation. Keywords: Neural progenitors, AAVS1 locus, EnvA-pseudotyped rabies virus, Retrograde monosynaptic tracing, Transplantation

Human pluripotent stem cells (hPSCs) exhibit very limited contribution to interspecies chimeras. One explanation is that the conventional hPSCs are in a primed state and so unable to form chimeras in pre-implantation embryos. Here, we show that the conventional hPSCs undergo rapid apoptosis when injected into mouse pre-implantation embryos. While, forced-expression of BMI1, a polycomb factor in hPSCs overcomes the apoptosis and enables hPSCs to integrate into mouse pre-implantation embryos and subsequently contribute to chimeras with both embryonic and extra-embryonic tissues. In addition, BMI1 also enables hPSCs to integrate into pre-implantation embryos of other species, such as rabbit and pig. Notably, BMI1 high expression and anti-apoptosis are also indicators for naïve hPSCs to form chimera in mouse embryos. Together, our findings reveal that the apoptosis is an initial barrier in interspecies chimerism using hPSCs and provide a rational to improve it., Conventional human pluripotent stem cells (hPSCs) fail to contribute to interspecies chimaeras when injected into mouse blastocysts. Here the authors show that forced expression of BMI1 overcomes apoptosis of hPSCs in blastocysts of mouse, rabbit and pig allowing them to contribute to chimaeras.

Chloroplast genomes are widely used in genetic engineering, molecular marker development, and phylogeny. In order to analyze the complete chloroplast genome of Dianthus caryophyllus, the complete chloroplast genome of D. caryophyllus was sequenced and annotated. On the other hand, phylogenetic analysis of the chloroplast genome of D. caryophyllus was carried out. The results showed that the whole length of the chloroplast genome of D. caryophyllus was 147,604 bp, and had a typical conserved quadripartite structure. The G and C basic content of D. caryophyllus chloroplast was 36.3%. The genome contained 83 protein-coding genes, 34 tRNA genes, and 6 rRNA genes. Among the protein-coding genes, 10 genes contain a single intron and 2 genes contain two introns. The phylogeny of D. caryophyllus indicated that the closest phylogenetic relationship was D. longicalyxanus. This study provides materials for the molecular study of D. caryophyllus may improve the carnation industry.

Definitive hematopoiesis generates hematopoietic stem/progenitor cells (HSPCs) that give rise to all mature blood and immune cells, but remains poorly defined in human. Here, we resolve human hematopoietic populations at the earliest hematopoiesis stage by single-cell RNA-seq. We characterize the distinct molecular profiling between early primitive and definitive hematopoiesis in both human embryonic stem cell (hESC) differentiation and early embryonic development. We identify CD44 to specifically discriminate definitive hematopoiesis and generate definitive HSPCs from hESCs. The multipotency of hESCs-derived HSPCs for various blood and immune cells is validated by single-cell clonal assay. Strikingly, these hESCs-derived HSPCs give rise to blood and lymphoid lineages in vivo. Lastly, we characterize gene-expression dynamics in definitive and primitive hematopoiesis and reveal an unreported role of ROCK-inhibition in enhancing human definitive hematopoiesis. Our study provides a prospect for understanding human early hematopoiesis and a firm basis for generating blood and immune cells for clinical purposes.

Human neural progenitor cell (hNPC) transplantation holds great potential to treat neurological diseases. However, hNPC grafts take a long time to differentiate into mature neurons due to their intrinsically prolonged developmental timetable. Here, we report that postoperative physical exercise (PE), a prevailing rehabilitation intervention, promotes the neuronal commitment, maturation, and integration of engrafted hNPCs, evidenced by forming more synapses, receiving more synaptic input from host neurons, and showing higher neuronal activity levels. More important, NPC transplantation, combined with PE, shows significant improvement in both structural and behavioral outcomes in stroke-damaged rats. PE enhances ingrowth of blood vessels around the infarction region and neural tract reorganization along the ischemic boundary. The combination of NPC transplantation and postoperative PE creates both a neurotrophic/growth factor-enriched proneuronal microenvironment and an ideal condition for activity-dependent plasticity to give full play to its effects. Our study provides a potential approach to treating patients with stroke injury.

Background: There is a huge controversy about whether xenograft or allograft in the “immune-privileged” brain needs immunosuppression. In animal studies, prevailing sophiscated immunosuppression or immunodeficiency is detrimental for the recipients, which results in short lifespan of animals, and confounds functional behavioral readout of the graft benefit, discouraging long-term follow-up. Methods: Neuron-restricted human neural progenitor cells (NPCs) were derived from human embryonic stem cells (ESCs, including H1, its gene-modified cell lines for better visualization, and HN4), propagated for different passages and then transplanted into the brain of immunocompetent rats without use of immunosuppressant. The graft survivals, their cell fates and HLA expression levels were examined over time (up to months after transplantation). We compared the survival capability of NPCs from different passages, and in different transplantation sites (brain parenchyma vs. para-and intra-ventricular sites). The host responses to the grafts were also investigated.Results: Our results show that human ESC-derived neuron-restricted NPCs survive extendedly in adult rat cerebro-parenchyma with no need of immunosuppression whereas a late-onset graft rejection seems inevitable. Both donor HLA expression level and host MHC-II expression level remain relatively low and little change over the long-term survival, and therefore can’t predict the late-onset rejection. Human grafts in or close to cerebroventricle are more vulnerable to the immune attack than the intra-striatum grafts. Prevention of graft hyperplasia by using hypoproliferative late-passaged human NPCs further significantly extends the graft survival time. Our new data also shows that a subpopulation of host microglia upregulate MHC-II expression in response to the human graft, but fail to present the human antigen to the host immune system, suggestive of the immune-isolation role of the blood–brain barrier (BBB). Conclusions: The present study confirms the “immune privilege” of the brain parenchyma, and more importantly, unveils that choosing hypoproliferative NPCs for transplantation can benefit graft outcome in terms of both lower tumor-genic risk and the prolonged survival time without immunosuppression.

Background Cultivated tea is one of the most important economic and ecological trees distributed worldwide. Cultivated tea suffer from long-term targeted selection of traits and overexploitation of habitats by human beings, which may have changed its genetic structure. The chloroplast is an organelle with a conserved cyclic genomic structure, and it can help us better understand the evolutionary relationship of Camellia plants. Results We conducted comparative and evolutionary analyses on cultivated tea and wild tea, and we detected the evolutionary characteristics of cultivated tea. The chloroplast genome sizes of cultivated tea were slightly different, ranging from 157,025 to 157,100 bp. In addition, the cultivated species were more conserved than the wild species, in terms of the genome length, gene number, gene arrangement and GC content. However, comparing Camellia sinensis var. sinensis and Camellia sinensis var. assamica with their cultivars, the IR length variation was approximately 20 bp and 30 bp, respectively. The nucleotide diversity of 14 sequences in cultivated tea was higher than that in wild tea. Detailed analysis on the genomic variation and evolution of Camellia sinensis var. sinensis cultivars revealed 67 single nucleotide polymorphisms (SNPs), 46 insertions/deletions (indels), and 16 protein coding genes with nucleotide substitutions, while Camellia sinensis var. assamica cultivars revealed 4 indels. In cultivated tea, the most variable gene was ycf1. The largest number of nucleotide substitutions, five amino acids exhibited site-specific selection, and a 9 bp sequence insertion were found in the Camellia sinensis var. sinensis cultivars. In addition, phylogenetic relationship in the ycf1 tree suggested that the ycf1 gene has diverged in cultivated tea. Because C. sinensis var. sinensis and its cultivated species were not tightly clustered. Conclusions The cultivated species were more conserved than the wild species in terms of architecture and linear sequence order. The variation of the chloroplast genome in cultivated tea was mainly manifested in the nucleotide polymorphisms and sequence insertions. These results provided evidence regarding the influence of human activities on tea.

As a potentially unlimited autologous cell source, induced pluripotent stem cells (iPSCs) provide a needed option for the application of iPSC-derived neural progenitor cells (NPCs) for regenerative medicine for the treatment of stroke. To enable the application of iPSC technology, it is essential to develop a practical approach to generate iPSC cells under a non-viral, non-integration, feeder-free condition from the most optimal somatic cell type. In this study, we differentiated NPCs from a urine-derived iPSC line (UC-05) which was generated with optimized episomal vectors in a feeder-free culture system. UC-05 can be induced into NPCs efficiently in monolayer cultures using dual SMAD inhibitions, and have the ability to differentiate further into astrocytes and functional neurons in vitro. We then characterized UC-05-derived NPCs upon transplantation into the striatum of adult male rats subjected to transient middle cerebral artery occlusion (tMCAO) reperfusion. While NPCs were grafted into rats 7 days before the MCAO surgery, cells were found to migrate from the grafted side to the lesion side of the brain via corpus callosum 14 days after tMCAO. UC05-derived NPCs were grafted into the striatum 7 days after tMCAO, grafted cells can survive and differentiate into neurons and astrocytes 35 days after transplantation, and synaptic protein SYNAPSIN 1 could also be detected around the grafted human cells. tMCAO rats with NPC engraftment showed better behavior improvement in both postural reflex test and cylinder test compared to control rats engrafted with the cell medium only. Our data indicate that NPCs differentiated from urine-derived iPSCs could act similarly to endogenous neural progenitors in vitro and in vivo. Urine-derived iPSCs could be a potential candidate for cell transplantation therapy in stroke.

[Image: see text] Microglial dysfunction is involved in the pathological cascade of Alzheimer’s disease (AD). The regulation of microglial function may be a novel strategy for AD therapy. We previously reported the discovery of AD16, an antineuroinflammatory molecule that could improve learning and memory in the AD model. Here, we studied its properties of microglial modification in the AD mice model. In this study, AD16 reduced interleukin-1β (IL-1β) expression in the lipopolysaccharide-induced IL-1β-Luc transgenic mice model. Compared with mice receiving placebo, the group treated with AD16 manifested a significant reduction of microglial activation, plaque deposition, and peri-plaques microgliosis, but without alteration of the number of microglia surrounding the plaque. We also found that AD16 decreased senescent microglial cells marked with SA-β-gal staining. Furthermore, altered lysosomal positioning, enhanced Lysosomal Associated Membrane Protein 1 (LAMP1) expression, and elevated adenosine triphosphate (ATP) concentration were found with AD16 treatment in lipopolysaccharide-stimulated BV2 microglial cells. The underlying mechanisms of AD16 might include regulating the microglial activation/senescence and recovery of its physiological function via the improvement of lysosomal function. Our findings provide new insights into the AD therapeutic approach through the regulation of microglial function and a promising lead compound for further study.

Mutations in the Leucine rich repeat kinase 2 (LRRK2) gene are found in both familial and sporadic Parkinson's disease (PD), and are also associated with immune-related disorders including Crohn's disease (CD) and leprosy. We have generated two homozygous LRRK2 knockout human induced pluripotent stem cell (iPSC) lines using CRISPR-Cas9 in a well-characterized human iPSC clone. The LRRK2 knockout cell lines retained normal morphology, gene expression, and the capacity to differentiate into cell types of the three germ layers. These cell lines are valuable for elucidating the role of LRRK2 in innate immunity and PD.

Background: The cultivated tea is one of the most important economic and ecological trees distributing worldwide. The cultivated tea suffered from long-term targeted selection of traits and overexploitation of habitats by human beings, which may change the genetic structure. Chloroplast is an organelle with a conserved cyclic structure, and can help us better understand the evolutionary relationship of Camellia plants.Results: The study conducted comparative analysis and evolution analysis between cultivated tea and wild tea, and detected the evolution characteristic in cultivated tea. Chloroplast genome sizes of cultivated tea were slightly different, ranged from 157,025 bp to 157,085 bp. These cultivated species were more conservative than wild species, in terms of the genome length, genes number, genes arrangement and GC contents. However, the IRs length of cultivated species was about 20 bp shorter than that of C. sinensis var. sisnensis. We also found that the nucleotide diversity of 14 sequences in cultivated tea was higher than that of wild tea. These results provided the evidence on the variation of chloroplast genomes of cultivated tea. Detail analysis on the chloroplast genome variation and evolution of cultivated tea showed that 67 SNPs and 46 indels and 16 protein coding genes had nucleotide substitutions. The most comment variation gene was ycf1. It has the largest number of nucleotide substitutions. At the same time, in ycf1, five amino acid sites were exhibiting site-specific selection, and a 9 bp sequence insertion was found in the C. sinensis cultivar. Anhua. The phylogenetic tree constructed by ycf1 sequence shows that two cultivated tea were not completely clustered, and the evolutionary relationship between C. sinensis var. sisnensis and C. sinensis cultivar. Longjing is closer than that of C. sinensis cultivar. Anhua.Conclusions: The cultivated species were more conservative than wild species in terms of architecture and linear sequence order. The variation of chloroplast genome of cultivated tea was mainly manifested in the nucleotide polymorphism of some sequences. The ycf1 gene played an important role in the adaptive evolution of cultivated tea. These results provided evidence regarding the influence of human activities on tea.

Loss of function mutations in PARK2 (encoding PARKIN) cause autosomal recessive Parkinson's disease (PD), which often manifests at a juvenile age. Molecular and biochemical studies show that PARKIN functions as an E3 ubiquitin ligase controlling mitochondrial homeostasis. Yet, the exact mechanisms are unclear due to the use of sub-optimal models including cancer cells and fibroblasts. We have generated a PARK2 knockout (KO) isogenic cell line using a well-characterized induced pluripotent stem cell (iPSC) clone with good differentiation potential. This cell line lacks the expression of all PARKIN isoforms and is valuable for elucidating the role of PARK2 mutations in PD.

Somatic cell reprogramming by exogenous factors requires cooperation with transcriptional co-activators and co-repressors to effectively remodel the epigenetic environment. How this interplay is regulated remains poorly understood. Here, we demonstrate that NCoR/SMRT co-repressors bind to pluripotency loci to create a barrier to reprogramming with the four Yamanaka factors (OCT4, SOX2, KLF4 and c-MYC), and consequently, suppressing NCoR/SMRT significantly enhances reprogramming efficiency and kinetics. The core epigenetic subunit of the NCoR/SMRT complex, histone deacetylase 3 (HDAC3), contributes to the effects of NCoR/SMRT by inducing histone deacetylation at pluripotency loci. Among the Yamanaka factors, recruitment of NCoR/SMRT–HDAC3 to genomic loci is mostly facilitated by c-MYC. Hence, we describe how c-MYC is beneficial for the early phase of reprogramming but deleterious later. Overall, we uncover a role for NCoR/SMRT co-repressors in reprogramming and propose a dual function for c-MYC in this process. Zhuang et al. demonstrate that suppression of NCoR/SMRT enhances OSKM reprogramming efficiency, and that the barrier mechanism depends on the recruitment of HDAC3 to pluripotency loci by c-MYC.

Polycomb repressive complex 2 and the epigenetic mark that it deposits, H3K27me3, are evolutionarily conserved and play critical roles in development and cancer. However, their roles in cell fate decisions in early embryonic development remain poorly understood. Here we report that knockout of polycomb repressive complex 2 genes in human embryonic stem cells causes pluripotency loss and spontaneous differentiation toward a meso-endoderm fate, owing to de-repression of BMP signalling. Moreover, human embryonic stem cells with deletion of EZH1 or EZH2 fail to differentiate into ectoderm lineages. We further show that polycomb repressive complex 2-deficient mouse embryonic stem cells also release Bmp4 but retain their pluripotency. However, when converted into a primed state, they undergo spontaneous differentiation similar to that of hESCs. In contrast, polycomb repressive complex 2 is dispensable for pluripotency when human embryonic stem cells are converted into the naive state. Our studies reveal both lineage- and pluripotent state-specific roles of polycomb repressive complex 2 in cell fate decisions., Polycomb repressive complex 2 (PRC2) plays an essential role in development by modifying chromatin but what this means at a cellular level is unclear. Here, the authors show that ablation of PRC2 genes in human embryonic stem cells and in mice results in changes in pluripotency and the primed state of cells.

Summary Familial hypercholesterolemia (FH) causes elevation of low-density lipoprotein cholesterol (LDL-C) in blood and carries an increased risk of early-onset cardiovascular disease. A caveat for exploration of new therapies for FH is the lack of adequate experimental models. We have created a comprehensive FH stem cell model with differentiated hepatocytes (iHeps) from human induced pluripotent stem cells (iPSCs), including genetically engineered iPSCs, for testing therapies for FH. We used FH iHeps to assess the effect of simvastatin and proprotein convertase subtilisin/kexin type 9 (PCSK9) antibodies on LDL-C uptake and cholesterol lowering in vitro. In addition, we engrafted FH iHeps into the liver of Ldlr−/−/Rag2−/−/Il2rg−/− mice, and assessed the effect of these same medications on LDL-C clearance and endothelium-dependent vasodilation in vivo. Our iHep models recapitulate clinical observations of higher potency of PCSK9 antibodies compared with statins for reversing the consequences of FH, demonstrating the utility for preclinical testing of new therapies for FH patients., Graphical Abstract, Highlights • Generation of a comprehensive stem cell model for familial hypercholesterolemia • Generation of chimeric mice with engrafted human iPSC-derived iHeps • FH iHeps respond to LDL-C-lowering medications in vitro and in vivo • Our model can be used for preclinical testing of novel LDL-C-lowering medications, In this paper, Tse, Esteban and colleagues have generated chimeric mice with human hepatocytes derived from familial hypercholesterolemia (FH) induced pluripotent stem cells. These chimeric mice resemble FH patients in the response to clinical-grade PCSK9 antibodies and statins. These results indicate the utility of this stem cell model for preclinical testing of new LDL-C-lowering therapies for FH patients.

Death-associated protein kinase 1 (DAPK1) is a Ca2+/calmodulin regulated Ser/Thr kinase involved in various cellular processes including cell death, autophagy and inflammation. Its dysregulation has been linked to tumour metastasis, anti-viral responses, Alzheimer's disease and other neurological disorders. To further investigate the role of DAPK1 in these processes, we generated a DAPK1 knockout first (conditional ready) human embryonic stem (hES) cell line in which the endogenous DAPK1 can be easily restored with expression of FLPe. This cell line provides an ideal model to study the role of DAPK1 in human development and various pathologies related to DAPK1 dysregulation in vitro.

Familial Parkinson's disease (PD) can be caused by deleterious mutations in PINK1 (encoding PINK1) in an autosomal recessive manner. Functional studies suggest that PINK1 works as a regulator of mitochondrial homeostasis. However, how loss of PINK1 induces dopaminergic neuron degeneration is still unclear. Here, we have generated a patient-derived induced pluripotent stem cell (iPSC) line with mutant PINK1 (p. I368N). This cell line will facilitate PD disease modeling in vitro and can be used for generating isogenic cell lines through gene correction.

Highly expressed in the mammalian brain and widely distributed across the genome, MeCP2 is a key player in recognizing modified DNA and interpreting the epigenetic information encoded in different DNA methylation/hydroxymethylation patterns. Alterations in sequence or copy number of the X-linked human MECP2 gene cause either Rett syndrome (RTT) or MECP2 duplication syndrome. Alterations in MECP2 levels have also been identified in patients with autism. To fully understand the significant role of MECP2 in regulating the development and function of the nervous system, it is important to study all aspects of MeCP2 function. Stimulus-induced MeCP2 phosphorylation has been shown to influence the proliferation and differentiation of neural progenitor cells, synaptic scaling, excitatory synaptogenesis, and animal behavior. However, all of the previous functional evidence is from studying phospho-dead mutations. In addition, the relationship between phosphorylation events at multiple sites on the MeCP2 protein is not well understood. Here, we report the generation of a phospho-mimic knockin Mecp2 mouse line. At the synaptic and behavioral levels, the phospho-mimic Mecp2 mice show phenotypes opposite to those observed in phospho-dead mutation at the same phosphorylation site. Moreover, we report opposite phenotypes between phospho-mutants of two sites on the MeCP2 protein. Our new data further confirm the functional significance of specific MeCP2 phosphorylation event and support the opposing regulatory role between different MeCP2 phosphorylation events.

During neurogenesis, neural patterning is a critical step during which neural progenitor cells differentiate into neurons with distinct functions. However, the molecular determinants that regulate neural patterning remain poorly understood. Here we optimized the "dual SMAD inhibition" method to specifically promote differentiation of human pluripotent stem cells (hPSCs) into forebrain and hindbrain neural progenitor cells along the rostral-caudal axis. We report that neural patterning determination occurs at the very early stage in this differentiation. Undifferentiated hPSCs expressed basal levels of the transcription factor orthodenticle homeobox 2 (OTX2) that dominantly drove hPSCs into the "default" rostral fate at the beginning of differentiation. Inhibition of glycogen synthase kinase 3β (GSK3β) through CHIR99021 application sustained transient expression of the transcription factor NANOG at early differentiation stages through Wnt signaling. Wnt signaling and NANOG antagonized OTX2 and, in the later stages of differentiation, switched the default rostral cell fate to the caudal one. Our findings have uncovered a mutual antagonism between NANOG and OTX2 underlying cell fate decisions during neural patterning, critical for the regulation of early neural development in humans.

The disease mechanism of Rett syndrome (RTT) is not well understood. Studies in RTT mouse models have suggested a non-cell-autonomous role for astrocytes in RTT pathogenesis. However, it is not clear whether this is also true for human RTT astrocytes. To establish an in vitro human RTT model, we previously generated isogenic induced pluripotent stem cell (iPSC) lines from several RTT patients carrying different disease-causing mutations. Here, we show that these RTT iPSC lines can be efficiently differentiated into astroglial progenitors and glial fibrillary acidic protein-expressing (GFAP(+)) astrocytes that maintain isogenic status, that mutant RTT astrocytes carrying three different RTT mutations and their conditioned media have adverse effects on the morphology and function of wild-type neurons and that the glial effect on neuronal morphology is independent of the intrinsic neuronal deficit in mutant neurons. Moreover, we show that both insulin-like growth factor 1 (IGF-1) and GPE (a peptide containing the first 3 amino acids of IGF-1) are able to partially rescue the neuronal deficits caused by mutant RTT astrocytes. Our findings confirm the critical glial contribution to RTT pathology, reveal potential cellular targets of IGF-1 therapy and further validate patient-specific iPSCs and their derivatives as valuable tools to study RTT disease mechanism.

Chloroplast genomes are widely used in genetic engineering, molecular marker development, and phylogeny. In order to analyze the complete chloroplast genome of Dianthus caryophyllus, the complete c...

DNA methylation-dependent epigenetic mechanisms underlie the development and function of the mammalian brain. MeCP2 is highly expressed in neurons and functions as a molecular linker between DNA methylation, chromatin remodeling and transcription regulation. Previous in vitro studies have shown that neuronal activity-induced phosphorylation (NAIP) of methyl CpG-binding protein 2 (MeCP2) precedes its release from the Bdnf promoter and the ensuing Bdnf transcription. However, the in vivo function of this phosphorylation event remains elusive. We generated knock-in mice that lack NAIP of MeCP2 and found that they performed better in hippocampus-dependent memory tests, presented enhanced long-term potentiation at two synapses in the hippocampus and showed increased excitatory synaptogenesis. At the molecular level, the phospho-mutant MeCP2 protein bound more tightly to several MeCP2 target gene promoters and altered the expression of these genes. Our results suggest that NAIP of MeCP2 is required for modulating dynamic functions of the adult mouse brain.

In the version of this Article originally published, in Fig. 2c, the '+' sign and 'OSKM' were superimposed in the label '+OSKM'. In Fig. 4e, in the labels, all instances of 'Ant' should have been 'Anti-'. And, in Fig. 7a, the label '0.0' was misplaced; it should have been on the colour scale bar. These figures have now been corrected in the online versions.