The Complement System: A Powerful Modulator and Effector of Astrocyte Function in the Healthy and Diseased Central Nervous System

The complement system, an effector arm of the innate immune system that plays a critical role in tissue inflammation, the elimination of pathogens and the clearance of dead cells and cell debris, has emerged as a regulator of many processes in the central nervous system, including neural cell genesis and migration, control of synapse number and function, and modulation of glial cell responses. Complement dysfunction has also been put forward as a major contributor to neurological disease. Astrocytes are neuroectoderm-derived glial cells that maintain water and ionic homeostasis, and control cerebral blood flow and multiple aspects of neuronal functioning. By virtue of their expression of soluble as well as membrane-bound complement proteins and receptors, astrocytes are able to both send and receive complement-related signals. Here we review the current understanding of the multiple functions of the complement system in the central nervous system as they pertain to the modulation of astrocyte activity, and how astrocytes use the complement system to affect their environment in the healthy brain and in the context of neurological disease.

TheAtoh7remote enhancer provides transcriptional robustness during retinal ganglion cell development

The retinal ganglion cell (RGC) competence factor ATOH7 is dynamically expressed during retinal histogenesis.ATOH7transcription is controlled by a promoter-adjacent primary enhancer and a remote shadow enhancer (SE). Deletion of theATOH7human SE causes nonsyndromic congenital retinal nonattachment (NCRNA) disease, characterized by optic nerve aplasia and total blindness. We used genome editing to model NCRNA in mice. Deletion of the murine SE reducesAtoh7messenger RNA (mRNA) fivefold but does not recapitulate optic nerve loss; however, SEdel/knockout (KO)transheterozygotes have thin optic nerves. By analyzingAtoh7mRNA and protein levels, RGC development and survival, and chromatin landscape effects, we show that the SE ensures robustAtoh7transcriptional output. Combining SE deletion and KO and wild-type alleles in a genotypic series, we determined the amount ofAtoh7needed to produce a normal complement of adult RGCs, and the secondary consequences of graded reductions inAtoh7dosage. Together, these data reveal the workings of an evolutionary fail-safe, a duplicate enhancer mechanism that is hard-wired in the machinery of vertebrate retinal ganglion cell genesis.

Age-Associated B Cells

The age-associated B cell subset has been the focus of increasing interest over the last decade. These cells have a unique cell surface phenotype and transcriptional signature, and they rely on TLR7 or TLR9 signals in the context of Th1 cytokines for their formation and activation. Most are antigen-experienced memory B cells that arise during responses to microbial infections and are key to pathogen clearance and control. Their increasing prevalence with age contributes to several well-established features of immunosenescence, including reduced B cell genesis and damped immune responses. In addition, they are elevated in autoimmune and autoinflammatory diseases, and in these settings they are enriched for characteristic autoantibody specificities. Together, these features identify age-associated B cells as a subset with pivotal roles in immunological health, disease, and aging. Accordingly, a detailed understanding of their origins, functions, and physiology should make them tractable translational targets in each of these settings.

Bromodeoxyuridine (BrdU) marker for proliferating cells in living tissues: Detection in adult avian brain

Knowing how to perform cell proliferation assays is an undeniably essential skill to detect the effect of external or internal stimuli on cell genesis. Immunohistochemical staining using Bromodeoxyuridine (BrdU), enables quantification of proliferative fraction of cells in sectioned tissues. BrdU, an artificial nucleoside that substitutes for thymidine. It is currently quite popular research tool of choice for new cell genesis studies. It incorporates into the DNA during S-phase of the cell cycle that’s why used for birth dating and observing cell proliferation. BrdU immunohistochemistry is currently the most used technique for studying adult cell genesis in situ. In this article we provide a step-by step protocol for immunodetection of BrdU by light microscopy using avian species (Ploceus philippinus); starting from BrdU administration to the final analysis.

Sufu- and Spop-mediated downregulation of Hedgehog signaling promotes beta cell differentiation through organ-specific niche signals

Human embryonic stem cell-derived beta cells offer a promising cell-based therapy for diabetes. However, efficient stem cell to beta cell differentiation has proven difficult, possibly due to the lack of cross-talk with the appropriate mesenchymal niche. To define organ-specific niche signals, we isolated pancreatic and gastrointestinal stromal cells, and analyzed their gene expression during development. Our genetic studies reveal the importance of tightly regulated Hedgehog signaling in the pancreatic mesenchyme: inactivation of mesenchymal signaling leads to annular pancreas, whereas stroma-specific activation of signaling via loss of Hedgehog regulators, Sufu and Spop, impairs pancreatic growth and beta cell genesis. Genetic rescue and transcriptome analyses show that these Sufu and Spop knockout defects occur through Gli2-mediated activation of gastrointestinal stromal signals such as Wnt ligands. Importantly, inhibition of Wnt signaling in organoid and human stem cell cultures significantly promotes insulin-producing cell generation, altogether revealing the requirement for organ-specific regulation of stromal niche signals.

Elevated Levels of Alpha Cells Emanating from the Pancreatic Ducts of a Patient with a Low BMI and Chronic Pancreatitis

Chronic pancreatitis (CP) is an inflammatory disease that causes progressive damage to the pancreatic parenchyma with irreversible morphological changes and fibrotic replacement of the gland. The risk factors associated with developing CP have been described as toxic (e.g., alcohol and tobacco); idiopathic (e.g., unknown); genetic, autoimmune, recurrent acute pancreatitis, and obstructive (the TIGAR-O system). Upon histological screening of the pancreata from a cohort of CP patients who had undergone pancreatectomy for the treatment of intractable pain in Leicester, UK, one sample showed a striking change in the morphological balance toward an endocrine phenotype, most notably there was evidence of substantial α cell genesis enveloping entire cross sections of ductal epithelium and the presence of α cells within the ductal lumens. This patient had previously undergone a partial pancreatectomy, had severe sclerosing CP, an exceptionally low body mass index (15.2), and diabetes at the time the pancreas was removed, and although these factors have been shown to induce tissue remodeling, such high levels of α cells was an unusual finding within our series of patients. Due to the fact that α cells have been shown to be the first endocrine cell type that emerges during islet neogenesis, future research profiling the factors that caused such marked α cell genesis may prove useful in the field of islet transplantation.