Early-Derived Murine Macrophages Temporarily Renounce Tissue Identity during Acute Systemic Inflammation.

In mice, a subset of cardiac macrophages and Kupffer cells derive from fetal precursors, seed the developing tissues, self-renew locally, and persist into adulthood. In this study we investigated how these cells survive acute systemic inflammation. In both tissues, early-derived subsets rapidly responded to acute systemic inflammation by assuming a temporary nonclassical activation state featuring upregulation of both proinflammatory (Il1b, Tnf, Nfkb1), and anti-inflammatory (Il10, Il4ra, Nfkbiz) genes. During this process, transcription factor genes associated with myeloid identity (Spi1, Zeb2) were upregulated, whereas those associated with tissue specificity (Nr1h3 for Kupffer cells and Nfatc2 and Irf4 for cardiac macrophages) were downregulated, suggesting that the cells reasserted their myeloid identity but renounced their tissue identity. Most of these changes in gene expression reverted to steady-state levels postresolution. We conclude that these early-derived macrophage subsets are resilient in the face of acute stress by temporary loss of adaptation to local tissue-specific niches while reasserting their generic myeloid identity.

Fetal origin confers radioresistance on liver macrophages via p21cip1/WAF1.

BACKGROUND & AIMS: Cells of hematopoietic origin, including macrophages, are generally radiation sensitive, but a subset of Kupffer cells (KCs) is relatively radioresistant. Here, we focused on the identity of the radioresistant KCs in unmanipulated mice and the mechanism of radioresistance. METHODS: We employed Emr1- and inducible CX3Cr1-based fate-mapping strategies combined with the RiboTag reporter to identify the total KCs and the embryo-derived KCs, respectively. The KC compartment was reconstituted with adult bone-marrow-derived KCs (bm-KCs) using clodronate depletion. Mice were lethally irradiated and transplanted with donor bone marrow, and the radioresistance of bone-marrow- or embryo-derived KCs was studied. Gene expression was analyzed using in situ mRNA isolation via RiboTag reporter mice, and the translatomes were compared among subsets. RESULTS: Here, we identified the radioresistant KCs as the long-lived subset that is derived from CX3CR1-expressing progenitor cells in fetal life, while adult bm-KCs do not resist irradiation. While both subsets upregulated the Cdkn1a gene, encoding p21-cip1/WAF1 protein, radioresistant embryo-derived KCs showed a greater increase in response to irradiation. In the absence of this molecule, the radioresistance of KCs was compromised. Replacement KCs, derived from adult hematopoietic stem cells, differed from radioresistant KCs in their expression of genes related to immunity and phagocytosis. CONCLUSIONS: Here, we show that, in the murine liver, a subset of KCs of embryonic origin resists lethal irradiation through Cdkn1a upregulation and is maintained for a long period, while bm-KCs do not survive lethal irradiation. LAY SUMMARY: Kupffer cells (KCs) are the tissue-resident macrophages of the liver. KCs can be originated from fetal precursors and from monocytes during the fetal stage and post-birth, respectively. Most immune cells in mice are sensitive to lethal-irradiation-induced death, while a subset of KCs resists radiation-induced death. These radioresistant KCs continue to live in the irradiated mice. We discovered that this relatively radioresistant KC subset are the fetal-derived KCs, and they achieve this through cell-cycle arrest. Understanding the radiobiology of KCs will provide valuable insights into the mechanisms that elicit radiation-induced liver disease.

Precision-cut human liver slice cultures as an immunological platform.

The liver is the central metabolic organ in the human body, and also plays an essential role in innate and adaptive immunity. While mouse models offer significant insights into immune-inflammatory liver disease, human immunology differs in important respects. It is not easy to address those differences experimentally. Therefore, to improve the understanding of human liver immunobiology and pathology, we have established precision-cut human liver slices to study innate immunity in human tissue. Human liver slices collected from resected livers could be maintained in ex vivo culture over a two-week period. Although an acute inflammatory response accompanied by signs of tissue repair was observed in liver tissue following slicing, the expression of many immune genes stabilized after day 4 and remained stable until day 15. Remarkably, histological evidence of pre-existing liver diseases was preserved in the slices for up to 7 days. Following 7 days of culture, exposure of liver slices to the toll-like receptor (TLR) ligands, TLR3 ligand Poly-I:C and TLR4 ligand LPS, resulted in a robust activation of acute inflammation and cytokine genes. Moreover, Poly-I:C treatment induced a marked antiviral response including increases of interferons IFNB, IL-28B and a group of interferon-stimulated genes. Therefore, precision-cut liver slices emerge as a valuable tool to study human innate immunity.

Central role of the TIR-domain-containing adaptor-inducing interferon-ß (TRIF) adaptor protein in murine sterile liver injury.

Multiple pathways drive the sterile injury response in the liver; however, it is unclear how the type of cells injured or the mechanism of injury activates these pathways. Here, we use a model of selective hepatocyte death to investigate sterile liver injury. In this model, the TIR-domain-containing adaptor-inducing interferon-ß (TRIF) was a central mediator of the resulting intrahepatic inflammatory response that was independent of both upstream Toll-like receptor (TLR) 4 signaling and downstream type I interferon (IFN) signaling. TRIF was required for induction of interleukin (IL)-10, IL-6, and IL-1ß cytokines. Conversely, although induction of C-C motif chemokine ligand (CCL) 2 and C-X-C motif chemokine ligand (CXCL) 1 chemokines and up-regulation of chemokine (Ccl2, Ccl7, Cxcl1, Cxcl2, and Cxcl10) and cell-adhesion (intracellular adhesion molecule 1 and vascular cell adhesion molecule 1) genes involved in myeloid cell recruitment was reduced in a majority of TRIF-/- mice, a subset of TRIF-/- mice showed breakthrough inflammation and the ability to induce these genes and proteins, indicating that redundant pathways exist to respond to hepatocyte death. Furthermore, we found that hepatocytes themselves were the main responders to hepatocyte death, increasing transcription of genes involved in myeloid cell recruitment more than either liver sinusoidal endothelial cells or Kupffer cells. CONCLUSION: Our studies define a TRIF-dependent, TLR4- and type I IFN-independent pathway of sterile liver injury in which hepatocytes are both the targets of damage and the principal responding cell type. (Hepatology 2017;65:1336-1351).

Role of Smad3 in platelet-derived growth factor-C-induced liver fibrosis.

Chronic liver injury leads to fibrosis and cirrhosis. Cirrhosis, the end stage of chronic liver disease, is a leading cause of death worldwide and increases the risk of developing hepatocellular carcinoma. Currently, there is a lack of effective antifibrotic therapies to treat fibrosis and cirrhosis. Development of antifibrotic therapies requires an in-depth understanding of the cellular and molecular mechanisms involved in inflammation and fibrosis after hepatic injury. Two growth factor signaling pathways that regulate liver fibrosis are transforming growth factor-ß (TGFß) and platelet-derived growth factor (PDGF). However, their specific contributions to fibrogenesis are not well understood. Using a genetic model of liver fibrosis, we investigated whether the canonical TGFß signaling pathway was necessary for fibrogenesis. PDGF-C transgenic (PDGF-C Tg) mice were intercrossed with mice that lack Smad3, and molecular and histological fibrosis was analyzed. PDGF-C Tg mice that also lacked Smad3 had less fibrosis and improved liver lobule architecture. Loss of Smad3 also reduced expression of collagen genes, which were induced by PDGF-C, but not the expression of genes frequently associated with hepatic stellate cell (HSC) activation. In vitro HSCs isolated from Smad3-null mice proliferated more slowly than cells from wild-type mice. Taken together, these findings indicate that PDGF-C activates TGFß/Smad3 signaling pathways to regulate HSC proliferation, collagen production and ultimately fibrosis. In summary, these results suggest that inhibition of both PDGF and TGFß signaling pathways may be required to effectively attenuate fibrogenesis in patients with chronic liver disease.

An interprofessional pediatric prescribing workshop.

OBJECTIVE: To design, implement, and evaluate an interprofessional learning workshop on pediatric prescribing. DESIGN: An interactive workshop on pediatric prescribing was designed and delivered by pediatricians and pharmacists to fourth-year medical and pharmacy students on 3 university campus settings. Students were assigned to either interprofessional workshop groups (pharmacy and medical students) or non-interprofessional workshop groups (medical students only). ASSESSMENT: Two hundred thirty students completed the workshops and 92% returned both the pre- and post-workshop questionnaires. Attitudes toward interprofessional learning significantly improved among students in the interprofessional workshop groups (p< 0.001) and confidence in prescribing for pediatric patients significantly improved among all students (p< 0.001). CONCLUSIONS: The workshop improved medical and pharmacy students' knowledge and confidence in pediatric prescribing and significantly improved their attitudes toward working and learning with other professionals.

Inactivation of p38 MAPK during liver regeneration.

There is increasing evidence that p38 MAPK, which is classified as a stress-activated kinase, also participates in cell cycle regulation, functioning as a suppressor of cell proliferation and tumorigenesis. We conducted a study of p38 MAPK phosphorylation during liver regeneration in mice to determine whether p38 MAPK activation or inactivation may correlate with events that lead to DNA replication after partial hepatectomy (PH), and whether p38 MAPK activation may be required for hepatocyte DNA replication in vivo and in culture. We report that active p38 (Pi-p38 MAPK) is present in normal liver, is rapidly inactivated starting 30 min after PH, and is re-activated by 12h. Although levels of Pi-MKK 3/6, the upstream kinases that activate p38 MAPK increase after PH, the expression of the dual protein phosphatase 1 is also elevated, and may be responsible for Pi-p38 MAPK dephosphorylation after PH. Inactivation and re-activation of p38 MAPK inversely correlates with the stimulation of protein synthesis and translation pathways, as indicated by activation of p70S6 kinase, increases in the phosphorylation of initiation factor elF-4E and translational repressor, 4E-BP. The activity of a p38 MAPK downstream substrate, MAPKAPK2 (MK2), did not reflect the changing levels of Pi-p38 MAPK during liver regeneration. Pi-p38 MAPK may be involved in TNF-stimulated DNA replication of murine hepatocytes in culture, but is not necessary for hepatocyte DNA replication after PH. Our results suggest that p38 MAPK inactivation plays a permissible role in DNA replication during liver regeneration and is consistent with a role for p38 MAPK in the maintenance of hepatocyte cell cycle arrest in adult liver.

Using care bundles to prevent infection in neonatal and paediatric ICUs.

PURPOSE OF REVIEW: Quality and safety of care are national priorities. Healthcare-acquired infections are now considered preventable and unacceptable. Care bundles are used to prevent and treat health-care acquired infections in adults. This paper considers the evidence and context for their use in children. RECENT FINDINGS: There is evidence that care bundles are effective in the adult literature. There have been few reports in the paediatric literature on the implementation of care bundles in children. Paediatric reports focus on the impact of interventions to reduce ventilator-associated pneumonia and central line infections. Recent articles suggest that care bundles are beneficial as part of a comprehensive improvement programme in the ICU. Other papers confirm that ventilator-associated pneumonia bundles can be translated from adults to children, supporting the business case for improving quality. SUMMARY: The adult and paediatric literature agree that care bundles are valuable tools for ensuring that evidence-based medicine is delivered reliably. Care bundles should be adopted in paediatric and neonatal units. In particular, if applied correctly, they are likely to significantly reduce certain health-care acquired infections. Further research is needed to refine the individual elements of the bundles and to evaluate new applications for them.