High-Dimensional Single-Cell Cartography Reveals Novel Skeletal Muscle-Resident Cell Populations.

Adult tissue repair and regeneration require stem-progenitor cells that can self-renew and generate differentiated progeny. Skeletal muscle regenerative capacity relies on muscle satellite cells (MuSCs) and their interplay with different cell types within the niche. However, our understanding of skeletal muscle tissue cellular composition is limited. Here, using a combined approach of single-cell RNA sequencing and mass cytometry, we precisely mapped 10 different mononuclear cell types in adult mouse muscle. We also characterized gene signatures and determined key discriminating markers of each cell type. We identified two previously understudied cell populations in the interstitial compartment. One expresses the transcription factor scleraxis and generated tenocytes in vitro. The second expresses markers of smooth muscle and mesenchymal cells (SMMCs) and, while distinct from MuSCs, exhibited myogenic potential and promoted MuSC engraftment following transplantation. The blueprint presented here yields crucial insights into muscle-resident cell-type identities and can be exploited to study muscle diseases.

Molecular regulation of stem cell quiescence.

Subsets of mammalian adult stem cells reside in the quiescent state for prolonged periods of time. This state, which is reversible, has long been viewed as dormant and with minimal basal activity. Recent advances in adult stem cell isolation have provided insights into the epigenetic, transcriptional and post-transcriptional control of quiescence and suggest that quiescence is an actively maintained state in which signalling pathways are involved in maintaining a poised state that allows rapid activation. Deciphering the molecular mechanisms regulating adult stem cell quiescence will increase our understanding of tissue regeneration mechanisms and how they are dysregulated in pathological conditions and in ageing.

Stem cell quiescence: the challenging path to activation.

Quiescence is a cellular state in which a cell remains out of the cell cycle but retains the capacity to divide. The unique ability of adult stem cells to maintain quiescence is crucial for life-long tissue homeostasis and regenerative capacity. Quiescence has long been viewed as an inactive state but recent studies have shown that it is in fact an actively regulated process and that adult stem cells are highly reactive to extrinsic stimuli. This has fuelled hopes of boosting the reactivation potential of adult stem cells to improve tissue function during ageing. In this Review, we provide a perspective of the quiescent state and discuss how quiescent adult stem cells transition into the cell cycle. We also discuss current challenges in the field, highlighting recent technical advances that could help overcome some of these challenges.

Nuclear receptors NHR-49 and NHR-79 promote peroxisome proliferation to compensate for aldehyde dehydrogenase deficiency in C. elegans.

The intracellular level of fatty aldehydes is tightly regulated by aldehyde dehydrogenases to minimize the formation of toxic lipid and protein adducts. Importantly, the dysregulation of aldehyde dehydrogenases has been implicated in neurologic disorder and cancer in humans. However, cellular responses to unresolved, elevated fatty aldehyde levels are poorly understood. Here, we report that ALH-4 is a C. elegans aldehyde dehydrogenase that specifically associates with the endoplasmic reticulum, mitochondria and peroxisomes. Based on lipidomic and imaging analysis, we show that the loss of ALH-4 increases fatty aldehyde levels and reduces fat storage. ALH-4 deficiency in the intestine, cell-nonautonomously induces NHR-49/NHR-79-dependent hypodermal peroxisome proliferation. This is accompanied by the upregulation of catalases and fatty acid catabolic enzymes, as indicated by RNA sequencing. Such a response is required to counteract ALH-4 deficiency since alh-4; nhr-49 double mutant animals are sterile. Our work reveals unexpected inter-tissue communication of fatty aldehyde levels and suggests pharmacological modulation of peroxisome proliferation as a therapeutic strategy to tackle pathology related to excess fatty aldehydes.

YY1 regulates skeletal muscle regeneration through controlling metabolic reprogramming of satellite cells.

Skeletal muscle satellite cells (SCs) are adult muscle stem cells responsible for muscle regeneration after acute or chronic injuries. The lineage progression of quiescent SC toward activation, proliferation, and differentiation during the regeneration is orchestrated by cascades of transcription factors (TFs). Here, we elucidate the function of TF Yin Yang1 (YY1) in muscle regeneration. Muscle-specific deletion of YY1 in embryonic muscle progenitors leads to severe deformity of diaphragm muscle formation, thus neonatal death. Inducible deletion of YY1 in SC almost completely blocks the acute damage-induced muscle repair and exacerbates the chronic injury-induced dystrophic phenotype. Examination of SC revealed that YY1 loss results in cell-autonomous defect in activation and proliferation. Mechanistic search revealed that YY1 binds and represses mitochondrial gene expression. Simultaneously, it also stabilizes Hif1α protein and activates Hif1α-mediated glycolytic genes to facilitate a metabolic reprogramming toward glycolysis which is needed for SC proliferation. Altogether, our findings have identified YY1 as a key regulator of SC metabolic reprogramming through its dual roles in modulating both mitochondrial and glycolytic pathways.

A long noncoding RNA, LncMyoD, modulates chromatin accessibility to regulate muscle stem cell myogenic lineage progression.

Epigenetics regulation plays a critical role in determining cell identity by controlling the accessibility of lineage-specific regulatory regions. In muscle stem cells, epigenetic mechanisms of how chromatin accessibility is modulated during cell fate determination are not fully understood. Here, we identified a long noncoding RNA, LncMyoD, that functions as a chromatin modulator for myogenic lineage determination and progression. The depletion of LncMyoD in muscle stem cells led to the down-regulation of myogenic genes and defects in myogenic differentiation. LncMyoD exclusively binds with MyoD and not with other myogenic regulatory factors and promotes transactivation of target genes. The mechanistic study revealed that loss of LncMyoD prevents the establishment of a permissive chromatin environment at myogenic E-box-containing regions, therefore restricting the binding of MyoD. Furthermore, the depletion of LncMyoD strongly impairs the reprogramming of fibroblasts into the myogenic lineage. Taken together, our study shows that LncMyoD associates with MyoD and promotes myogenic gene expression through modulating MyoD accessibility to chromatin, thereby regulating myogenic lineage determination and progression.

p110α of PI3K is necessary and sufficient for quiescence exit in adult muscle satellite cells.

Adult mouse muscle satellite cells (MuSCs) are quiescent in uninjured muscles. Upon injury, MuSCs exit quiescence in vivo to become activated, re-enter the cell cycle to proliferate, and differentiate to repair the damaged muscles. It remains unclear which extrinsic cues and intrinsic signaling pathways regulate quiescence exit during MuSC activation. Here, we demonstrated that inducible MuSC-specific deletion of p110α, a catalytic subunit of phosphatidylinositol 3-kinase (PI3K), rendered MuSCs unable to exit quiescence, resulting in severely impaired MuSC proliferation and muscle regeneration. Genetic reactivation of mTORC1, or knockdown of FoxOs, in p110α-null MuSCs partially rescued the above defects, making them key effectors downstream of PI3K in regulating quiescence exit. c-Jun was found to be a key transcriptional target of the PI3K/mTORC1 signaling axis essential for MuSC quiescence exit. Moreover, induction of a constitutively active PI3K in quiescent MuSCs resulted in spontaneous MuSC activation in uninjured muscles and subsequent depletion of the MuSC pool. Thus, PI3K-p110α is both necessary and sufficient for MuSCs to exit quiescence in response to activating signals.

Intronic polyadenylation of PDGFRα in resident stem cells attenuates muscle fibrosis.

Platelet-derived growth factor receptor α (PDGFRα) exhibits divergent effects in skeletal muscle. At physiological levels, signalling through this receptor promotes muscle development in growing embryos and angiogenesis in regenerating adult muscle. However, both increased PDGF ligand abundance and enhanced PDGFRα pathway activity cause pathological fibrosis. This excessive collagen deposition, which is seen in aged and diseased muscle, interferes with muscle function and limits the effectiveness of gene- and cell-based therapies for muscle disorders. Although compelling evidence exists for the role of PDGFRα in fibrosis, little is known about the cells through which this pathway acts. Here we show in mice that PDGFRα signalling regulates a population of muscle-resident fibro/adipogenic progenitors (FAPs) that play a supportive role in muscle regeneration but may also cause fibrosis when aberrantly regulated. We found that FAPs produce multiple transcriptional variants of Pdgfra with different polyadenylation sites, including an intronic variant that codes for a protein isoform containing a truncated kinase domain. This variant, upregulated during regeneration, acts as a decoy to inhibit PDGF signalling and to prevent FAP over-activation. Moreover, increasing the expression of this isoform limits fibrosis in vivo in mice, suggesting both biological relevance and therapeutic potential of modulating polyadenylation patterns in stem-cell populations.

Pre-operative cognitive burden as predictor of motor outcome following bilateral subthalamic nucleus deep brain stimulation in Parkinson's disease.

INTRODUCTION: The interrelationship between neurocognitive impairments and motor functions was observed in patients with advanced Parkinson's disease (PD). This study was conducted to identify pre-operative neurocognitive and clinical predictors of short-term motor outcome following subthalamic nucleus deep brain stimulation (STN-DBS). METHODS: All consecutive PD patients who were eligible for bilateral STN-DBS from 2009 to 2019 were evaluated before and at 1 year following surgery. Standard motor evaluation and neurocognitive tests including global cognition, memory, executive functions (attention and category fluency), confrontational speech, visuospatial abilities, and mood were conducted at baseline. The post-operative STN-DBS effects were assessed at 1 year following the surgery. Multiple regression analysis was applied to identify baseline independent predictors of post-operative STN-DBS effect. RESULTS: A total of 82 patients were analyzed. It was found that younger age at operation, higher levodopa responsiveness at baseline based on UPDRS-III total score, and better baseline verbal delayed memory and category fluency predicted post-operative motor outcome at 1 year following STN-DBS (F = 9.639, p < 0.001, R2 = .340). CONCLUSION: Our findings demonstrated the role of baseline cognitive burden, especially cognitive processes related to frontostriatal circuits, was significant clinical predictors of short-term motor outcomes following STN-DBS. Profile analysis of neurocognitive functions at baseline is recommended.

α2-Chimaerin is essential for neural stem cell homeostasis in mouse adult neurogenesis.

Adult hippocampal neurogenesis involves the lifelong generation of neurons. The process depends on the homeostasis of the production of neurons and maintenance of the adult neural stem cell (NSC) pool. Here, we report that α2-chimaerin, a Rho GTPase-activating protein, is essential for NSC homeostasis in adult hippocampal neurogenesis. Conditional deletion of α2-chimaerin in adult NSCs resulted in the premature differentiation of NSCs into intermediate progenitor cells (IPCs), which ultimately depleted the NSC pool and impaired neuron generation. Single-cell RNA sequencing and pseudotime analyses revealed that α2-chimaerin-conditional knockout (α2-CKO) mice lacked a unique NSC subpopulation, termed Klotho-expressing NSCs, during the transition of NSCs to IPCs. Furthermore, α2-CKO led to defects in hippocampal synaptic plasticity and anxiety/depression-like behaviors in mice. Our findings collectively demonstrate that α2-chimaerin plays an essential role in adult hippocampal NSC homeostasis to maintain proper brain function.

IL-33 ameliorates Alzheimer's disease-like pathology and cognitive decline.

Alzheimer's disease (AD) is a devastating condition with no known effective treatment. AD is characterized by memory loss as well as impaired locomotor ability, reasoning, and judgment. Emerging evidence suggests that the innate immune response plays a major role in the pathogenesis of AD. In AD, the accumulation of ß-amyloid (Aß) in the brain perturbs physiological functions of the brain, including synaptic and neuronal dysfunction, microglial activation, and neuronal loss. Serum levels of soluble ST2 (sST2), a decoy receptor for interleukin (IL)-33, increase in patients with mild cognitive impairment, suggesting that impaired IL-33/ST2 signaling may contribute to the pathogenesis of AD. Therefore, we investigated the potential therapeutic role of IL-33 in AD, using transgenic mouse models. Here we report that IL-33 administration reverses synaptic plasticity impairment and memory deficits in APP/PS1 mice. IL-33 administration reduces soluble Aß levels and amyloid plaque deposition by promoting the recruitment and Aß phagocytic activity of microglia; this is mediated by ST2/p38 signaling activation. Furthermore, IL-33 injection modulates the innate immune response by polarizing microglia/macrophages toward an antiinflammatory phenotype and reducing the expression of proinflammatory genes, including IL-1ß, IL-6, and NLRP3, in the cortices of APP/PS1 mice. Collectively, our results demonstrate a potential therapeutic role for IL-33 in AD.

Maintenance of muscle stem-cell quiescence by microRNA-489.

Among the key properties that distinguish adult mammalian stem cells from their more differentiated progeny is the ability of stem cells to remain in a quiescent state for prolonged periods of time. However, the molecular pathways for the maintenance of stem-cell quiescence remain elusive. Here we use adult mouse muscle stem cells (satellite cells) as a model system and show that the microRNA (miRNA) pathway is essential for the maintenance of the quiescent state. Satellite cells that lack a functional miRNA pathway spontaneously exit quiescence and enter the cell cycle. We identified quiescence-specific miRNAs in the satellite-cell lineage by microarray analysis. Among these, miRNA-489 (miR-489) is highly expressed in quiescent satellite cells and is quickly downregulated during satellite-cell activation. Further analysis revealed that miR-489 functions as a regulator of satellite-cell quiescence, as it post-transcriptionally suppresses the oncogene Dek, the protein product of which localizes to the more differentiated daughter cell during asymmetric division of satellite cells and promotes the transient proliferative expansion of myogenic progenitors. Our results provide evidence of the miRNA pathway in general, and of a specific miRNA, miR-489, in actively maintaining the quiescent state of an adult stem-cell population.

A sequential comparison on the risk of haemorrhage with different sizes of biopsy needles for stereotactic brain biopsy.

AIM: To compare the risk of postoperative haemorrhage with different sizes of brain biopsy needles. PATIENTS AND METHOD: A cohort of patients using a 2.5-mm outer diameter side-cutting biopsy needle was compared to a subsequent cohort using a 1.8-mm needle of the same type. All data were collected prospectively. A CT scan was done within 12 h after surgery. Any visible haemorrhage at the operated site was documented. RESULTS: From 2007 to 2013, 54 stereotactic brain biopsies (all frameless except for one frame-based) were performed. The 2.5-mm group comprised 29 procedures from 2007 to 2009. The 1.8-mm group comprised the subsequent 25 procedures. The diagnostic yields were 90 and 96% in the 2.5- and the 1.8-mm group, respectively (p = 0.615). Comparing the 2.5- and the 1.8-mm group, haemorrhage was significantly reduced: incidence (72 vs. 40%, p = 0.016); size of haemorrhage (mean 7.2 vs. 2.6 mm, p = 0.002); proportion of haemorrhage size >10 mm (34.5 vs. 4%, p = 0.006). Symptomatic haemorrhage rates were 3.4 and 0.0% in the 2.5- and the 1.8-mm group, respectively (p = 1.00). CONCLUSION: The 1.8-mm outer diameter needle carried a lower risk of postoperative haemorrhage than the 2.5-mm one, without compromising the diagnostic yield.

Middle cerebral artery infarction in a cancer patient: a fatal case of Trousseau's syndrome.

Trousseau's syndrome is defined as any unexplained thrombotic event that precedes the diagnosis of an occult visceral malignancy or appears concomitantly with a tumour. This report describes a young, previously healthy man diagnosed to have an acute middle cerebral arterial ischaemic stroke and lower-limb deep vein thrombosis, who subsequently succumbed to pulmonary arterial embolism. During the course of his illness, he was diagnosed to have a malignant pleural effusion secondary to an occult adenocarcinoma. This report highlights the need for a high degree of suspicion for occult malignancy and non-bacterial thrombotic endocarditis in young (<60 years old) ischaemic stroke patients with no identifiable conventional cardiovascular risks. In selected patients, transoesophageal echocardiography is the diagnostic investigation of choice, since transthoracic imaging is not sensitive. Screening tests for serum tumour markers and prompt heparinisation of these patients are suggested whenever ischaemic stroke secondary to malignancy-induced systemic hypercoagulability is suspected.

Spontaneous intracranial hypotension: improving recognition and treatment strategies in the local setting.

We report a case of spontaneous intracranial hypotension with classic symptoms of orthostatic headache and acute presentation of subdural haematoma on computed tomographic scan. Conventional approach with conservative treatment was initially adopted. The patient's condition, however, deteriorated after 2 weeks, requiring surgical evacuation of the intracranial haemorrhage. We reviewed the clinical features of this disease and the correlated magnetic resonance imaging findings with the pathophysiological mechanisms, and described treatment strategies in the local setting. Subtle findings on initial computed tomographic scan are also reported which might improve pathology recognition. Spontaneous intracranial hypotension is not uncommonly encountered in Hong Kong, and physicians must adopt a high level of clinical suspicion to facilitate early diagnosis and appropriate management. In addition, novel therapeutic approaches may be required in those with recurrent symptoms or who are refractory to current treatment strategies.

Muscle stem cell niche dynamics during muscle homeostasis and regeneration.

The process of skeletal muscle regeneration involves a coordinated interplay of specific cellular and molecular interactions within the injury site. This review provides an overview of the cellular and molecular components in regenerating skeletal muscle, focusing on how these cells or molecules in the niche regulate muscle stem cell functions. Dysfunctions of muscle stem cell-to-niche cell communications during aging and disease will also be discussed. A better understanding of how niche cells coordinate with muscle stem cells for muscle repair will greatly aid the development of therapeutic strategies for treating muscle-related disorders.

Diagnostic Accuracy and Field for Improvement of Frameless Stereotactic Brain Biopsy: A Focus on Nondiagnostic Cases.

BACKGROUND: The diagnostic accuracy of frameless stereotactic brain biopsy has been reported, but there is limited literature focusing on the reasons for nondiagnostic cases. In this study, we evaluate the diagnostic accuracy of frameless stereotactic brain biopsy, compare it with the current international standard, and review the field for improvement. METHODS: This is a retrospective analysis of consecutive, prospectively collected frameless stereotactic brain biopsies from 2007 to 2020. We evaluated the diagnostic accuracy of the frameless stereotactic brain biopsies using defined criteria. The biopsy result was classified as conclusive, inconclusive, or negative, based on the pathologic, radiologic, and clinical diagnosis concordance. For inconclusive or negative results, we further evaluated the preoperative planning and postoperative imaging to review the errors. A literature review for the diagnostic accuracy of frameless stereotactic biopsy was performed for the validity of our results. RESULTS: There were 106 patients with 109 biopsies performed from 2007 to 2020. The conclusive diagnosis was reached in 103 (94.5%) procedures. An inconclusive diagnosis was noted in four (3.7%) procedures and the biopsy was negative in two (1.9%) procedures. Symptomatic hemorrhage occurred in one patient (0.9%). There was no mortality in our series. Registration error (RE) and inaccurate targeting occurred in three trigonal lesions (2.8%), sampling of the nonrepresentative part of the lesion occurred in two cases (1.8%), and one biopsy (0.9%) for lymphoma was negative due to steroid treatment. The literature review suggested that our diagnostic accuracy was comparable with the published literature. CONCLUSION: The frameless stereotactic biopsy is a safe procedure with high diagnostic accuracy only if meticulous preoperative planning and careful intraoperative registration is performed. The common pitfalls precluding a conclusive diagnosis are RE and biopsies at nonrepresentative sites.

Niche inflammatory signals control oscillating mammary regeneration and protect stem cells from cytotoxic stress.

Stem cells are known for their resilience and enhanced activity post-stress. The mammary gland undergoes frequent remodeling and is subjected to recurring stress during the estrus cycle, but it remains unclear how mammary stem cells (MaSCs) respond to the stress and contribute to regeneration. We discovered that cytotoxic stress-induced activation of CD11c+ ductal macrophages aids stem cell survival and prevents differentiation. These macrophages boost Procr+ MaSC activity through IL1ß-IL1R1-NF-κB signaling during the estrus cycle in an oscillating manner. Deleting IL1R1 in MaSCs results in stem cell loss and skewed luminal differentiation. Moreover, under cytotoxic stress from the chemotherapy agent paclitaxel, ductal macrophages secrete higher IL1ß levels, promoting MaSC survival and preventing differentiation. Inhibiting IL1R1 sensitizes MaSCs to paclitaxel. Our findings reveal a recurring inflammatory process that regulates regeneration, providing insights into stress-induced inflammation and its impact on stem cell survival, potentially affecting cancer therapy efficacy.

Setdb1 protects genome integrity in murine muscle stem cells to allow for regenerative myogenesis and inflammation.

The histone H3 lysine 9 methyltransferase SETDB1 controls transcriptional repression to direct stem cell fate. Here, we show that Setdb1 expression by adult muscle stem cells (MuSCs) is required for skeletal muscle regeneration. We find that SETDB1 represses the expression of endogenous retroviruses (ERVs) in MuSCs. ERV de-repression in Setdb1-null MuSCs prevents their amplification following exit from quiescence and promotes cell death. Multi-omics profiling shows that chromatin decompaction at ERV loci activates the DNA-sensing cGAS-STING pathway, entailing cytokine expression by Setdb1-null MuSCs. This is followed by aberrant infiltration of inflammatory cells, including pathological macrophages. The ensuing histiocytosis is accompanied by myofiber necrosis, which, in addition to progressive MuSCs depletion, completely abolishes tissue repair. In contrast, loss of Setdb1 in fibro-adipogenic progenitors (FAPs) does not impact immune cells. In conclusion, genome maintenance by SETDB1 in an adult somatic stem cell is necessary for both its regenerative potential and adequate reparative inflammation.

Notch signaling is necessary to maintain quiescence in adult muscle stem cells.

Satellite cells (SCs) are myogenic stem cells found in skeletal muscle that function to repair tissue damaged by injury or disease. SCs are quiescent at rest, although the signaling pathways required to maintain quiescence are unknown. Using a transgenic Notch reporter mouse and quantitative reverse-transcription polymerase chain reaction analysis of Notch target genes, we determined that Notch signaling is active in quiescent SCs. SC-specific deletion of recombining binding protein-Jκ (RBP-Jκ), a nuclear factor required for Notch signaling, resulted in the depletion of the SC pool and muscles that lacked any ability to regenerate in response to injury. SC depletion was not due to apoptosis. Rather, RBP-Jκ-deficient SCs spontaneously activate, fail to self-renew, and undergo terminal differentiation. Intriguingly, most of the cells differentiate without first dividing. They then fuse with adjacent myofibers, leading to the gradual disappearance of SCs from the muscle. These results demonstrate the requirement of Notch signaling for the maintenance of the quiescent state and for muscle stem cell homeostasis by the regulation of self-renewal and differentiation, processes that are all critical for normal postnatal myogenesis.