Reprogramming roadmap reveals route to human induced trophoblast stem cells.

The reprogramming of human somatic cells to primed or naive induced pluripotent stem cells recapitulates the stages of early embryonic development1-6. The molecular mechanism that underpins these reprogramming processes remains largely unexplored, which impedes our understanding and limits rational improvements to reprogramming protocols. Here, to address these issues, we reconstruct molecular reprogramming trajectories of human dermal fibroblasts using single-cell transcriptomics. This revealed that reprogramming into primed and naive pluripotency follows diverging and distinct trajectories. Moreover, genome-wide analyses of accessible chromatin showed key changes in the regulatory elements of core pluripotency genes, and orchestrated global changes in chromatin accessibility over time. Integrated analysis of these datasets revealed a role for transcription factors associated with the trophectoderm lineage, and the existence of a subpopulation of cells that enter a trophectoderm-like state during reprogramming. Furthermore, this trophectoderm-like state could be captured, which enabled the derivation of induced trophoblast stem cells. Induced trophoblast stem cells are molecularly and functionally similar to trophoblast stem cells derived from human blastocysts or first-trimester placentas7. Our results provide a high-resolution roadmap for the transcription-factor-mediated reprogramming of human somatic cells, indicate a role for the trophectoderm-lineage-specific regulatory program during this process, and facilitate the direct reprogramming of somatic cells into induced trophoblast stem cells.

Alzheimer's disease-induced phagocytic microglia express a specific profile of coding and non-coding RNAs.

INTRODUCTION: Alzheimer's disease (AD) is a neurodegenerative disease and the main cause of dementia in the elderly. AD pathology is characterized by accumulation of microglia around the beta-amyloid (Aß) plaques which assumes disease-specific transcriptional signatures, as for the disease-associated microglia (DAM). However, the regulators of microglial phagocytosis are still unknown. METHODS: We isolated Aß-laden microglia from the brain of 5xFAD mice for RNA sequencing to characterize the transcriptional signature in phagocytic microglia and to identify the key non-coding RNAs capable of regulating microglial phagocytosis. Through spatial sequencing, we show the transcriptional changes of microglia in the AD mouse brain in relation to Aß proximity. RESULTS: Finally, we show that phagocytic messenger RNAs are regulated by miR-7a-5p, miR-29a-3p and miR-146a-5p microRNAs and segregate the DAM population into phagocytic and non-phagocytic states. DISCUSSION: Our study pinpoints key regulators of microglial Aß clearing capacity suggesting new targets for future therapeutic approaches.

Caecal bascule - A variant of caecal volvulus: Review of diagnostic challenges and approaches.

Caecal volvulus is a rare cause of intestinal obstruction, of which the caecal bascule is the most uncommonly encountered subtype. Definitive radiological diagnosis of a caecal bascule can be challenging. Lack of familiarity with this rare condition can contribute to delayed diagnosis and treatment, which may result in unnecessary morbidity. This article highlights the case of a patient who presented with a diagnostic challenge, and also discusses the pathogenesis and diagnostic features of caecal bascules. Various options in the surgical management of caecal bascules are reviewed, including the feasibility of laparoscopic-assisted approaches in the emergency setting.

Novel Anti-Neuroinflammatory Properties of a Thiosemicarbazone-Pyridylhydrazone Copper(II) Complex.

Neuroinflammation has a major role in several brain disorders including Alzheimer's disease (AD), yet at present there are no effective anti-neuroinflammatory therapeutics available. Copper(II) complexes of bis(thiosemicarbazones) (CuII(gtsm) and CuII(atsm)) have broad therapeutic actions in preclinical models of neurodegeneration, with CuII(atsm) demonstrating beneficial outcomes on neuroinflammatory markers in vitro and in vivo. These findings suggest that copper(II) complexes could be harnessed as a new approach to modulate immune function in neurodegenerative diseases. In this study, we examined the anti-neuroinflammatory action of several low-molecular-weight, charge-neutral and lipophilic copper(II) complexes. Our analysis revealed that one compound, a thiosemicarbazone-pyridylhydrazone copper(II) complex (CuL5), delivered copper into cells in vitro and increased the concentration of copper in the brain in vivo. In a primary murine microglia culture, CuL5 was shown to decrease secretion of pro-inflammatory cytokine macrophage chemoattractant protein 1 (MCP-1) and expression of tumor necrosis factor alpha (Tnf), increase expression of metallothionein (Mt1), and modulate expression of Alzheimer's disease-associated risk genes, Trem2 and Cd33. CuL5 also improved the phagocytic function of microglia in vitro. In 5xFAD model AD mice, treatment with CuL5 led to an improved performance in a spatial working memory test, while, interestingly, increased accumulation of amyloid plaques in treated mice. These findings demonstrate that CuL5 can induce anti-neuroinflammatory effects in vitro and provide selective benefit in vivo. The outcomes provide further support for the development of copper-based compounds to modulate neuroinflammation in brain diseases.

Midbrain organoids-development and applications in Parkinson's disease.

Human brain development is spatially and temporally complex. Insufficient access to human brain tissue and inadequacy of animal models has limited the study of brain development and neurodegenerative diseases. Recent advancements of brain organoid technology have created novel opportunities to model human-specific neurodevelopment and brain diseases. In this review, we discuss the use of brain organoids to model the midbrain and Parkinson's disease. We critically evaluate the extent of recapitulation of PD pathology by organoids and discuss areas of future development that may lead to the model to become a next-generation, personalized therapeutic strategy for PD and beyond.

Evaluating Capture Sequence Performance for Single-Cell CRISPR Activation Experiments.

The combination of single-cell RNA sequencing with CRISPR inhibition/activation provides a high-throughput approach to simultaneously study the effects of hundreds if not thousands of gene perturbations in a single experiment. One recent development in CRISPR-based single-cell techniques introduces a feature barcoding technology that allows for the simultaneous capture of mRNA and guide RNA (gRNA) from the same cell. This is achieved by introducing a capture sequence, whose complement can be incorporated into each gRNA and that can be used to amplify these features prior to sequencing. However, because the technology is in its infancy, there is little information available on how such experimental parameters can be optimized. To overcome this, we varied the capture sequence, capture sequence position, and gRNA backbone to identify an optimal gRNA scaffold for CRISPR activation gene perturbation studies. We provide a report on our screening approach along with our observations and recommendations for future use.

Transcriptional signature in microglia associated with Aß plaque phagocytosis.

The role of microglia cells in Alzheimer's disease (AD) is well recognized, however their molecular and functional diversity remain unclear. Here, we isolated amyloid plaque-containing (using labelling with methoxy-XO4, XO4+) and non-containing (XO4-) microglia from an AD mouse model. Transcriptomics analysis identified different transcriptional trajectories in ageing and AD mice. XO4+ microglial transcriptomes demonstrated dysregulated expression of genes associated with late onset AD. We further showed that the transcriptional program associated with XO4+ microglia from mice is present in a subset of human microglia isolated from brains of individuals with AD. XO4- microglia displayed transcriptional signatures associated with accelerated ageing and contained more intracellular post-synaptic material than XO4+ microglia, despite reduced active synaptosome phagocytosis. We identified HIF1α as potentially regulating synaptosome phagocytosis in vitro using primary human microglia, and BV2 mouse microglial cells. Together, these findings provide insight into molecular mechanisms underpinning the functional diversity of microglia in AD.

A single-cell atlas of entorhinal cortex from individuals with Alzheimer's disease reveals cell-type-specific gene expression regulation.

There is currently little information available about how individual cell types contribute to Alzheimer's disease. Here we applied single-nucleus RNA sequencing to entorhinal cortex samples from control and Alzheimer's disease brains (n = 6 per group), yielding a total of 13,214 high-quality nuclei. We detail cell-type-specific gene expression patterns, unveiling how transcriptional changes in specific cell subpopulations are associated with Alzheimer's disease. We report that the Alzheimer's disease risk gene APOE is specifically repressed in Alzheimer's disease oligodendrocyte progenitor cells and astrocyte subpopulations and upregulated in an Alzheimer's disease-specific microglial subopulation. Integrating transcription factor regulatory modules with Alzheimer's disease risk loci revealed drivers of cell-type-specific state transitions towards Alzheimer's disease. For example, transcription factor EB, a master regulator of lysosomal function, regulates multiple disease genes in a specific Alzheimer's disease astrocyte subpopulation. These results provide insights into the coordinated control of Alzheimer's disease risk genes and their cell-type-specific contribution to disease susceptibility. These results are available at http://adsn.ddnetbio.com.

CuII(atsm) Attenuates Neuroinflammation.

Background: Neuroinflammation and biometal dyshomeostasis are key pathological features of several neurodegenerative diseases, including Alzheimer's disease (AD). Inflammation and biometals are linked at the molecular level through regulation of metal buffering proteins such as the metallothioneins. Even though the molecular connections between metals and inflammation have been demonstrated, little information exists on the effect of copper modulation on brain inflammation. Methods: We demonstrate the immunomodulatory potential of the copper bis(thiosemicarbazone) complex CuII(atsm) in an neuroinflammatory model in vivo and describe its anti-inflammatory effects on microglia and astrocytes in vitro. Results: By using a sophisticated in vivo magnetic resonance imaging (MRI) approach, we report the efficacy of CuII(atsm) in reducing acute cerebrovascular inflammation caused by peripheral administration of bacterial lipopolysaccharide (LPS). CuII(atsm) also induced anti-inflammatory outcomes in primary microglia [significant reductions in nitric oxide (NO), monocyte chemoattractant protein 1 (MCP-1), and tumor necrosis factor (TNF)] and astrocytes [significantly reduced NO, MCP-1, and interleukin 6 (IL-6)] in vitro. These anti-inflammatory actions were associated with increased cellular copper levels and increased the neuroprotective protein metallothionein-1 (MT1) in microglia and astrocytes. Conclusion: The beneficial effects of CuII(atsm) on the neuroimmune system suggest copper complexes are potential therapeutics for the treatment of neuroinflammatory conditions.

Neuroinflammation and copper in Alzheimer's disease.

Inflammation is the innate immune response to infection or tissue damage. Initiation of proinflammatory cascades in the central nervous system (CNS) occurs through recognition of danger associated molecular patterns by cognate immune receptors expressed on inflammatory cells and leads to rapid responses to remove the danger stimulus. The presence of activated microglia and astrocytes in the vicinity of amyloid plaques in the brains of Alzheimer's disease (AD) patients and mouse models implicates inflammation as a contributor to AD pathogenesis. Activated microglia play a critical role in amyloid clearance, but chronic deregulation of CNS inflammatory pathways results in secretion of neurotoxic mediators that ultimately contribute to neurodegeneration in AD. Copper (Cu) homeostasis is profoundly affected in AD, and accumulated extracellular Cu drives A ß aggregation, while intracellular Cu deficiency limits bioavailable Cu required for CNS functions. This review presents an overview of inflammatory events that occur in AD in response to A ß and highlights recent advances on the role of Cu in modulation of beneficial and detrimental inflammatory responses in AD.