Effect of 1-MHz ultrasound on the proinflammatory interleukin-6 secretion in human keratinocytes

Keratinocytes, the main cell type of the skin, are one of the most exposed cells to environmental factors, providing a first defence barrier for the host and actively participating in immune response. In fact, keratinocytes express pattern recognition receptors that interact with pathogen associated molecular patterns and damage associated molecular patterns, leading to the production of cytokines and chemokines, including interleukin (IL)-6. Herein, we investigated whether mechanical energy transported by low intensity ultrasound (US) could generate a mechanical stress able to induce the release of inflammatory cytokine such IL-6 in the human keratinocyte cell line, HaCaT. The extensive clinical application of US in both diagnosis and therapy suggests the need to better understand the related biological effects. Our results point out that US promotes the overexpression and secretion of IL-6, associated with the activation of nuclear factor-κB (NF-κB). Furthermore, we observed a reduced cell viability dependent on exposure parameters together with alterations in membrane permeability, paving the way for further investigating the molecular mechanisms related to US exposure.

Beyond the Extracellular Vesicles: Technical Hurdles, Achieved Goals and Current Challenges When Working on Adipose Cells

Adipose tissue and its crosstalk with other organs plays an essential role in the metabolic homeostasis of the entire body. Alteration of this communication (i.e., due to obesity) is related to the development of several comorbidities including type 2 diabetes, cardiovascular diseases, or cancer. Within the adipose depot, adipocytes are the main cell type and thus the main source of secreted molecules, which exert modulating effects not only at a local but also at a systemic level. Extracellular vesicles (EVs) have recently emerged as important mediators in cell–cell communication and account for part of the cellular secretome. In recent years, there has been a growing body of research on adipocyte-derived extracellular vesicles (Ad-EVs). However, there is still a lack of standardized methodological approaches, especially regarding primary adipocytes. In this review, we will provide an outline of crucial aspects when working on adipose-derived material, with a special focus on primary adipocytes. In parallel, we will point out current methodological challenges in the EV field and how they impact the transcriptomic, proteomic and functional evaluations of Ad-EVs.

Case Report: Microglia Composition and Immune Response in an Immunocompetent Patient With an Intracranial Syphilitic Gumma

The pathogenesis of intracranial syphilitic gummas remains poorly understood. Microglia are generally considered to be the main cell type of the innate immune system in the brain. Determination of the composition of infiltrating microglia of patients with typical intracranial syphilitic gummas may contribute to the understanding of the pathological process. We report a case of an intracranial syphilitic gumma who presented with right upper limb weakness. The histological analysis showed the presence of Treponema pallidum and infiltration with histiocytes. Immunostaining indicated that cells were predominantly the M2a and M2c, which were Arg-1+ and IL-10+. These findings suggest that there is an increased number of M2a/M2c microglia in intracranial syphilitic gummas, which may be part of the immune escape mechanisms triggered by Treponema pallidum.

Differential susceptibility of retinal ganglion cell subtypes in acute and chronic models of injury and disease

Retinal ganglion cells (RGCs) are a heterogeneous population of neurons, comprised of numerous subtypes that work synchronously to transmit visual information to the brain. In blinding disorders such as glaucoma, RGCs are the main cell type to degenerate and lead to loss of vision. Previous studies have identified and characterized a variety of RGC subtypes in animal models, although only a handful of studies demonstrate the differential loss of these RGC subtypes in response to disease or injury. Thus, efforts of the current study utilized both chronic (bead occlusion) and acute (optic nerve crush, ONC) rat models to characterize disease response and differential loss of RGC subtypes. Bead occlusion and ONC retinas demonstrated significant RGC loss, glial reactivity and apoptosis compared to control retinas. Importantly, bead occlusion and ONC retinas resulted in differential subtype-specific loss of RGCs, with a high susceptibility for alpha- and direction selective-RGCs and preferential survival of ipRGCs. Results of this study serve as an important foundation for future experiments focused on the mechanisms resulting in the loss of RGCs in optic neuropathies, as well as the development of targeted therapeutics for RGC subtype-specific neuroprotection.

Effects of BDNF and PEC Nanoparticles on Osteocytes

Bone substitute materials loaded with mediators that stimulate fracture healing are demanded in the clinical treatment in trauma surgery and orthopedics. Brain-derived neurotrophic factor (BDNF) enhances the proliferation and differentiation of mesenchymal stem cells into osteoblast. To load the implants with BDNF, a drug delivery system that allows the release of BDNF under spatiotemporal control would improve functionality. Polyelectrolyte complex nanoparticles (PECNP) have been reported as a suitable drug delivery system. The suitability of PECNP in contact with osteocytes as the main cell type of bone is not known so far. Thus, we aimed to verify that BDNF and PECNP loaded with BDNF (PECNP+BDNF) as well as pure PECNP have no negative effects on osteocytes in vitro. Therefore, the murine osteocyte cell line MLO-Y4 was treated with BDNF and PECNP+BDNF. The effects on proliferation were analyzed by the BrdU test (n = 5). The results demonstrated a significant increase in proliferation 24 h after BDNF application, whereas PECNP+BDNF did not lead to significant changes. Thus, we conclude that BDNF is an appropriate mediator to stimulate osteocytes. Since the addition of PECNP did not affect the viability of osteocytes, we conclude that PECNP are a suitable drug delivery system for bone implants.

VEGF-A in Cardiomyocytes and Heart Diseases

The vascular endothelial growth factor (VEGF), a homodimeric vasoactive glycoprotein, is the key mediator of angiogenesis. Angiogenesis, the formation of new blood vessels, is responsible for a wide variety of physio/pathological processes, including cardiovascular diseases (CVD). Cardiomyocytes (CM), the main cell type present in the heart, are the source and target of VEGF-A and express its receptors, VEGFR1 and VEGFR2, on their cell surface. The relationship between VEGF-A and the heart is double-sided. On the one hand, VEGF-A activates CM, inducing morphogenesis, contractility and wound healing. On the other hand, VEGF-A is produced by CM during inflammation, mechanical stress and cytokine stimulation. Moreover, high concentrations of VEGF-A have been found in patients affected by different CVD, and are often correlated with an unfavorable prognosis and disease severity. In this review, we summarized the current knowledge about the expression and effects of VEGF-A on CM and the role of VEGF-A in CVD, which are the most important cause of disability and premature death worldwide. Based on clinical studies on angiogenesis therapy conducted to date, it is possible to think that the control of angiogenesis and VEGF-A can lead to better quality and span of life of patients with heart disease.

Smooth Muscle Cell-Proteoglycan-Lipoprotein Interactions as Drivers of Atherosclerosis

In humans, smooth muscle cells (SMCs) are the main cell type in the artery medial layer, in pre-atherosclerotic diffuse thickening of the intima, and in all stages of atherosclerotic lesion development. SMCs secrete the proteoglycans responsible for the initial binding and retention of atherogenic lipoproteins in the artery intima, with this retention driving foam cell formation and subsequent stages of atherosclerosis. In this chapter we review current knowledge of the extracellular matrix generated by SMCs in medial and intimal arterial layers, their relationship to atherosclerotic lesion development and stabilization, how these findings correlate with mouse models of atherosclerosis, and potential therapies aimed at targeting the SMC matrix-lipoprotein interaction for atherosclerosis prevention.

PTX3 Regulates miR-21 Expression and Secretion in Brown Adipocytes During Lipopolysaccharide-induced Inflammation (P21-072-19)

Objectives Metabolic endotoxemia is known to induce systemic inflammation and adipose tissue dysfunction during obesity. Pentraxin 3 (PTX3), a member of the pentraxin family can be induced by inflammatory stimuli in adipocytes. However, how PTX3 regulates inflammation in adipocytes is not fully understood. MicroRNAs (miRNAs) are key regulators of inflammatory gene expression. Specifically, miR-21 plays a role in the resolution of inflammation through inhibiting Toll-like receptor 4 (TLR4) signaling pathway. This study aimed to investigate how PTX3 regulates inflammatory homeostasis involving miR-21 during lipopolysaccharide (LPS) stimulation in brown adipocytes. Methods Using PTX3 knockout (KO) mouse and primary stromal-vascular (SV) cell culture models, we determined the effect of PTX3 deficiency on miR-21 expression, secretion, and inflammatory response in brown adipocytes, as well as the rescue effect of recombinant PTX3 on LPS-induced inflammation. Results Brown adipose tissue (BAT) had significantly higher levels of Dicer protein expression than inguinal and epididymal adipose depots; fully differentiated brown adipocytes expressed higher levels of Dicer than SV cells. These results suggest that BAT is the major site of miRNA production, and brown adipocyte is the main cell type that produces miRNAs. Moreover, we found brown adipocytes but not SV cells are the LPS responsive cells in miR-21 expression and secretion. In WT brown adipocytes, LPS stimulation significantly up-regulated cellular levels of miR-21. Interestingly, PTX3 deficiency led to a significant increase in the basal levels of both cellular and secreted miR21 compared to WT cells. Unlike in WT cells, LPS treatment suppressed cellular expression as well as secretion of miR-21 in PTX3 KO brown adipocytes. Treatment of recombinant PTX3 slightly up-regulated cellular expression of miR-21, but significantly increased the secretion of miR-21 in PTX3 KO brown adipocytes. Moreover, Treatment of recombinant PTX3 significantly attenuated LPS-stimulated increase in the expression of TNFα and MCP1 genes in PTX3 KO adipocytes. Conclusions We conclude that PTX3 plays an anti-inflammatory role in part via regulating the expression and secretion of miR-21. Funding Sources American Diabetes Association.

Liver Regeneration: Different Sub-Populations of Parenchymal Cells at Play Choreographed by an Injury-Specific Microenvironment

Liver regeneration is crucial for the maintenance of liver functional mass during homeostasis and diseases. In a disease context-dependent manner, liver regeneration is contributed to by hepatocytes or progenitor cells. As long as they are replicatively competent, hepatocytes are the main cell type responsible for supporting liver size homeostasisand regeneration. The concept that all hepatocytes within the lobule have the same proliferative capacity but are differentially recruited according to the localization of the wound, or whether a yet to be defined sub-population of hepatocytes supports regeneration is still debated. In a chronically or severely injured liver, hepatocytes may enter a state of replicative senescence. In such conditions, small biliary cells activate and expand, a process called ductular reaction (DR). Work in the last few decades has demonstrated that DR cells can differentiate into hepatocytes and thereby contribute to parenchymal reconstitution. In this study we will review the molecular mechanisms supporting these two processes to determine potential targets that would be amenable for therapeutic manipulation to enhance liver regeneration.

Differential Susceptibility of Rat Retinal Ganglion Cells Following Optic Nerve Crush

Retinal ganglion cells (RGCs) are a heterogeneous group of cells, comprised of numerous subpopulations, that work together to send visual information to the brain. In numerous blinding disorders termed optic neuropathies, RGCs are the main cell type affected leading to degeneration of these cells and eventual loss of vision. Previous studies have identified and characterized RGC subtypes in numerous animal systems, with only a handful of studies demonstrating their differential loss in response to disease and injury. Thus, efforts of the current study utilized an optic nerve crush (ONC) model to characterize the loss of RGCs and disease phenotypes associated with this injury. Additionally, the loss of RGC subtypes including direction selective-, alpha-, and ip-RGCs following ONC was explored. Results of this study demonstrated the differential loss of RGC subtypes with a high susceptibility for loss of alpha- and direction selective-RGCs and the preferential survival of ip-RGCs following ONC and allows for the establishment of additional studies focused on mechanisms and loss of these cells in optic neuropathies. Additionally, these results put important emphasis on the development of therapeutics targeted at the loss of specific subtypes as well as cellular replacement following injury and disease.