Molecular Insights Into the Effects of PLLA-SCA on Gene Expression and Collagen Synthesis in Human 3D Skin Models Containing Macrophages.

Injectable poly-L-lactic acid (PLLA-SCA) is used for the correction of shallow to deep nasolabial fold contour deficiencies, cheek wrinkles, and other facial wrinkles. In contrast to hyaluronan (HA) fillers, PLLA-SCA has a biostimulatory effect by activating resident fibroblasts to produce collagen, but the mechanisms are not known in detail at the molecular level. Therefore, our aim was to investigate the molecular effects of PLLA-SCA in a comprehensive in vitro study. Since PLLA-SCA-dependent collagen production in fibroblasts depends on the interaction with macrophages, we generated novel macrophage-containing 3D skin models. According to the clinical application, PLLA-SCA was injected once into the dermal equivalent of the 3D skin model. Histological analysis showed a significant increase in epidermal thickness in these models after 5 and 14 days. Gene expression profiling revealed an upregulation of integrins and laminins (e.g., LAMA3, ITGA6), which are essential components of the dermal-epidermal junction. In addition, we found an upregulation of cytokines and chemokines (TGFB2, CXCL6, IL1B) at day 14 after PLLA-SCA injection. Interestingly, immunohistochemical analyses exhibited a significantly stimulated collagen I production in our models. These effects might be attributed, at least in part, to the upregulation of IL1B and subsequently CXCL6, which stimulates collagen I synthesis in human dermal fibroblasts as we could demonstrate. Taken together, our data provide for the first time molecular insights into the biostimulatory effects of PLLA-SCA on collagen I production in novel human 3D skin models comprising macrophages. J Drugs Dermatol. 2024;23(4):7791.    doi:10.36849/JDD.7791.

Macrophage-like rapid uptake and toxicity of tattoo ink in human monocytes.

Macrophages play a critical role for the persistence of tattoo ink in human skin. However, a comparison to other skin-resident and blood circulating immune cells and a profound analysis of REACH-compliant tattoo ink are unmet medical needs. We hence characterized the size distribution of ink particles using physicochemical methods. We studied the uptake of tattoo ink by key human skin cells and blood-derived immune cells using optical and electron microscopy as well as flow cytometry. Scanning electron microscopy of ink revealed its crystalline structure, and a tendency towards aggregations was indicated by size changes upon diluting it. Flow cytometric analyses of skin and immune cells after incubation with tattoo ink demonstrated an increase in cellular granularity upon uptake and red ink additionally evoked fluorescent signals. Human macrophages were most potent in internalizing ink in full thickness 3D skin models. Macrophage cultures demonstrated that the ink did not lead to elevated inflammatory mediators, and showed no indications for toxicity, even after nice days. Strikingly, monocytes were most efficient in ink uptake, but displayed reduced viability, whereas granulocytes and lymphocytes showed only temporary ink uptake with flow cytometric signals declining after 1 day. Mechanistic studies on ink retention by corticosteroids or dexpanthenol in macrophage cultures demonstrated that these compounds do not lead to ink excretion, but even slightly increase the ink load in macrophages. The highly motile monocytes, precursors of macrophages, may play an underrated role for tattoo ink translocation from dermal blood vessels into internal organs.

CCL2 chemokine inhibition primes the tumor vasculature for improved nanomedicine delivery and efficacy.

Blood vessel functionality is crucial for efficient tumor-targeted drug delivery. Heterogeneous distribution and perfusion of angiogenic blood vessels contribute to suboptimal accumulation of (nano-) therapeutics in tumors and metastases. To attenuate pathological angiogenesis, an L-RNA aptamer inhibiting the CC motif chemokine ligand 2 (CCL2) was administered to mice bearing orthotopic 4T1 triple-negative breast cancer tumors. The effect of CCL2 inhibition on tumor blood vessel functionality and tumor-targeted drug delivery was evaluated via multimodal and multiscale optical imaging, employing fluorophore-labeled polymeric (10 nm) and liposomal (100 nm) nanocarriers. Anti-CCL2 treatment induced a dose-dependent anti-angiogenic effect, reflected by a decreased relative blood volume, increased blood vessel maturity and functionality, and reduced macrophage infiltration, accompanied by a shift in the polarization of tumor-associated macrophages (TAM) towards a less M2-like and more M1-like phenotype. In line with this, CCL2 inhibitor treatment improved the delivery of polymers and liposomes to tumors, and enhanced the antitumor efficacy of free and liposomal doxorubicin. Together, these findings demonstrate that blocking the CCL2-CCR2 axis modulates TAM infiltration and polarization, resulting in vascular normalization and improved tumor-targeted drug delivery.

JNKs protect from cholestatic liver disease progression by modulating Apelin signalling.

Background & Aims: Cholestatic liver injury is associated with c-Jun N-terminal kinases (JNK) activation in distinct cell types. Its hepatocyte-specific function during cholestasis, however, has not yet been established. Therefore, in our present study, we investigated the role of JNK1/2 during cholestasis and dissected its hepatocyte-specific function. Methods: A cohort of patients with primary biliary cholangitis (n = 29) and primary sclerosing cholangitis (n = 37) was examined. Wild-type, hepatocyte-specific knockout mice for Jnk2 (Jnk2Δhepa) or Jnk1 and Jnk2 (Jnk1Δhepa/2Δhepa) were generated. Mice were subjected to bile duct ligation (BDL) or carbon tetrachloride (CCl4) treatment. Finally, Apelin signalling was blocked using a specific inhibitor. As an interventional approach, Jnk1/2 were silenced in wild-type mice using lipid nanoparticles for small interfering RNA delivery. Results: JNK activation was increased in liver specimens from patients with chronic cholestasis (primary biliary cholangitis and primary sclerosing cholangitis) and in livers of Mdr2-/- and BDL-treated animals. In Jnk1Δhepa/2Δhepa animals, serum transaminases increased after BDL, and liver histology demonstrated enhanced cell death, compensatory proliferation, hepatic fibrogenesis, and inflammation. Furthermore, microarray analysis revealed that hepatocytic Jnk1/2 ablation induces JNK-target genes involved in oxidative stress and Apelin signalling after BDL. Consequently, blocking Apelin signalling attenuated BDL-induced liver injury and fibrosis in Jnk1Δhepa/2Δhepa mice. Finally, we established an interventional small interfering RNA approach of selective Jnk1/2 targeting in hepatocytes in vivo, further demonstrating the essential protective role of Jnk1/2 during cholestasis. Conclusions: Jnk1 and Jnk2 work together to protect hepatocytes from cholestatic liver disease by controlling Apelin signalling. Dual modification of JNK signalling in hepatocytes is feasible, and enhancing its expression might be an attractive therapeutic approach for cholestatic liver disease. Impact and Implications: The cell-specific function of Jnk genes during cholestasis has not been explicitly explored. In this study, we showed that combined Jnk1/2, but not Jnk2 deficiency, in hepatocytes exacerbates liver damage and fibrosis by enhancing Apelin signalling, which contributes to cholestasis progression. Combined cell-specific Jnk targeting may be a new molecular strategy for treating cholestatic liver disease.

Expression of Cyclin E1 in hepatic stellate cells is critical for the induction and progression of liver fibrosis and hepatocellular carcinoma in mice.

Hepatocellular carcinoma (HCC) is one of the most severe malignancies with increasing incidence and limited treatment options. Typically, HCC develops during a multistep process involving chronic liver inflammation and liver fibrosis. The latter is characterized by the accumulation of extracellular matrix produced by Hepatic Stellate Cells (HSCs). This process involves cell cycle re-entry and proliferation of normally quiescent HSCs in an ordered sequence that is highly regulated by cyclins and associated cyclin-dependent kinases (CDKs) such as the Cyclin E1 (CCNE1)/CDK2 kinase complex. In the present study, we examined the role of Cyclin E1 (Ccne1) and Cdk2 genes in HSCs for liver fibrogenesis and hepatocarcinogenesis. To this end, we generated conditional knockout mice lacking Ccne1 or Cdk2 specifically in HSCs (Ccne1∆HSC or Cdk2∆HSC). Ccne1∆HSC mice showed significantly reduced liver fibrosis formation and attenuated HSC activation in the carbon tetrachloride (CCl4) model. In a combined model of fibrosis-driven hepatocarcinogenesis, Ccne1∆HSC mice revealed decreased HSC activation even after long-term observation and substantially reduced tumor load in the liver when compared to wild-type controls. Importantly, the deletion of Cdk2 in HSCs also resulted in attenuated liver fibrosis after chronic CCl4 treatment. Single-cell RNA sequencing revealed that only a small fraction of HSCs expressed Ccne1/Cdk2 at a distinct time point after CCl4 treatment. In summary, we provide evidence that Ccne1 expression in a small population of HSCs is sufficient to trigger extensive liver fibrosis and hepatocarcinogenesis in a Cdk2-dependent manner. Thus, HSC-specific targeting of Ccne1 or Cdk2 in patients with liver fibrosis and high risk for HCC development could be therapeutically beneficial.

Microgels as Platforms for Antibody-Mediated Cytokine Scavenging.

Therapeutic antibodies are the key treatment option for various cytokine-mediated diseases, such as rheumatoid arthritis, psoriasis, and inflammatory bowel disease. However, systemic injection of these antibodies can cause side effects and suppress the immune system. Moreover, clearance of therapeutic antibodies from the blood is limiting their efficacy. Here, water-swollen microgels are produced with a size of 25 µm using droplet-based microfluidics. The microgels are functionalized with TNFα antibodies to locally scavenge the pro-inflammatory cytokine TNFα. Homogeneous distribution of TNFα-antibodies is shown throughout the microgel network and demonstrates specific antibody-antigen binding using confocal microscopy and FLIM-FRET measurements. Due to the large internal accessibility of the microgel network, its capacity to bind TNFα is extremely high. At a TNFα concentration of 2.5 µg mL-1 , the microgels are able to scavenge 88% of the cytokine. Cell culture experiments reveal the therapeutic potential of these microgels by protecting HT29 colorectal adenocarcinoma cells from TNFα toxicity and resulting in a significant reduction of COX II and IL8 production of the cells. When the microgels are incubated with stimulated human macrophages, to mimic the in vivo situation of inflammatory bowel disease, the microgels scavenge almost all TNFα that is produced by the cells.

Targeting Ligand Independent Tropism of siRNA-LNP by Small Molecules for Directed Therapy of Liver or Myeloid Immune Cells.

Hepatic clearance of lipid nanoparticles (LNP) with encapsulated nucleic acids restricts their therapeutic applicability. Therefore, tools for regulating hepatic clearance are of high interest for nucleic acid delivery. To this end, this work employs wild-type (WT) and low-density lipoprotein receptor (Ldlr)-/- mice pretreated with either a leukotriene B4 receptor inhibitor (BLT1i) or a high-density lipoprotein receptor inhibitor (HDLRi) prior to the injection of siRNA-LNP. This work is able to demonstrate significantly increased hepatic uptake of siRNA-LNP by the BLT1i in Ldlr-/- mice by in vivo imaging and discover an induction of specific uptake-related proteins. Irrespective of the inhibitors and Ldlr deficiency, the siRNA-LNP induced RNA-binding and transport-related proteins in liver, including haptoglobin (HP) that is also identified as most upregulated serum protein. This work observes a downregulation of proteins functioning in hepatic detoxification and of serum opsonins. Most strikingly, the HDLRi reduces hepatic uptake and increases siRNA accumulation in spleen and myeloid immune cells of blood and liver. RNA sequencing demonstrates leukocyte recruitment by the siRNA-LNP and the HDLRi through induction of chemokine ligands in liver tissue. The data provide insights into key mechanisms of siRNA-LNP biodistribution and indicate that the HDLRi has potential for extrahepatic and leukocyte targeting.

Profiling target engagement and cellular uptake of cRGD-decorated clinical-stage core-crosslinked polymeric micelles.

Polymeric micelles are increasingly explored for tumor-targeted drug delivery. CriPec® technology enables the generation of core-crosslinked polymeric micelles (CCPMs) based on thermosensitive (mPEG-b-pHPMAmLacn) block copolymers, with high drug loading capacity, tailorable size, and controlled drug release kinetics. In this study, we decorated clinical-stage CCPM with the αvß3 integrin-targeted cyclic arginine-glycine-aspartic acid (cRGD) peptide, which is one of the most well-known active targeting ligands evaluated preclinically and clinically. Using a panel of cell lines with different expression levels of the αvß3 integrin receptor and exploring both static and dynamic incubation conditions, we studied the benefit of decorating CCPM with different densities of cRGD. We show that incubation time and temperature, as well as the expression levels of αvß3 integrin by target cells, positively influence cRGD-CCPM uptake, as demonstated by immunofluorescence staining and fluorescence microscopy. We demonstrate that even very low decoration densities (i.e., 1 mol % cRGD) result in increased engagement and uptake by target cells as compared to peptide-free control CCPM, and that high cRGD decoration densities do not result in a proportional increase in internalization. In this context, it should be kept in mind that a more extensive presence of targeting ligands on the surface of nanomedicines may affect their pharmacokinetic and biodistribution profile. Thus, we suggest a relatively low cRGD decoration density as most suitable for in vivo application.

Hepatocyte-Directed Delivery of Lipid-Encapsulated Small Interfering RNA.

Lipid formulations for cell transfection are among the most efficient systems for nucleic acid delivery. During the COVID-19 pandemic, lipid-encapsulated RNA (lipid nanoparticles, LNP) has succeeded as a superior vaccine. Moreover, other similar lipid nanocarriers for siRNA are approved and many are on the pipelines. While lipid encapsulation required several devices for the mixing of components, lipoplex technology allows to rapidly mix nucleic acids and positively charged lipids for cell transfection. In vivo, hepatocytes are important target cells of lipid formulated RNAi. This chapter describes the state-of-the-art lipoplex and LPN manufacturing for treating primary hepatocytes with lipid formulations. Furthermore, protocols for isolating murine hepatocytes and for transfecting these cells with pharmaceutically relevant lipid formulations are provided and discussed.

Physiologically Based Pharmacokinetic (PBPK) Model of Gold Nanoparticle-Based Drug Delivery System for Stavudine Biodistribution.

Computational modelling has gained attention for evaluating nanoparticle-based drug delivery systems. Physiologically based pharmacokinetic (PBPK) modelling provides a mechanistic approach for evaluating drug biodistribution. The aim of this work is to develop a specific PBPK model to simulate stavudine biodistribution after the administration of a 40 nm gold nanoparticle-based drug delivery system in rats. The model parameters used have been obtained from literature, in vitro and in vivo studies, and computer optimization. Based on these, the PBPK model was built, and the compartments included were considered as permeability rate-limited tissues. In comparison with stavudine solution, a higher biodistribution of stavudine into HIV reservoirs and the modification of pharmacokinetic parameters such as the mean residence time (MRT) have been observed. These changes are particularly noteworthy in the liver, which presents a higher partition coefficient (from 0.27 to 0.55) and higher MRT (from 1.28 to 5.67 h). Simulated stavudine concentrations successfully describe these changes in the in vivo study results. The average fold error of predicted concentrations after the administration of stavudine-gold nanoparticles was within the 0.5-2-fold error in all of the tissues. Thus, this PBPK model approach may help with the pre-clinical extrapolation to other administration routes or the species of stavudine gold nanoparticles.

Immunomodulatory Nanomedicine for the Treatment of Atherosclerosis.

Atherosclerosis is the main underlying cause of cardiovascular diseases (CVDs), which remain the number one contributor to mortality worldwide. Although current therapies can slow down disease progression, no treatment is available that can fully cure or reverse atherosclerosis. Nanomedicine, which is the application of nanotechnology in medicine, is an emerging field in the treatment of many pathologies, including CVDs. It enables the production of drugs that interact with cellular receptors, and allows for controlling cellular processes after entering these cells. Nanomedicine aims to repair, control and monitor biological and physiological systems via nanoparticles (NPs), which have been shown to be efficient drug carriers. In this review we will, after a general introduction, highlight the advantages and limitations of the use of such nano-based medicine, the potential applications and targeting strategies via NPs. For example, we will provide a detailed discussion on NPs that can target relevant cellular receptors, such as integrins, or cellular processes related to atherogenesis, such as vascular smooth muscle cell proliferation. Furthermore, we will underline the (ongoing) clinical trials focusing on NPs in CVDs, which might bring new insights into this research field.

Lipid nanoparticle formulations for targeting leukocytes with therapeutic RNA in liver fibrosis.

Obesity and low-grade inflammation are promoters of a multitude of diseases including liver fibrosis. Activation of the mobile leukocytes has a major impact on the outcome of inflammatory disease and can hence foster or mitigate liver fibrosis. This renders immunological targets valuable for directed interventions using nanomedicines. Particularly, RNA-based drugs formulated as lipid nanoparticles (LNP) can open new avenues for the personalized treatment of liver fibrosis both through specific interference and via the induction of the expression of functional and therapeutic proteins. Using microfluidics technology, all components, including lipid-anchored targeting ligands, are assembled in a single-step mixing process. A highlight is set to immunologically relevant liver cell types that are most vulnerable for being reached by LNP. A selection of LNP from other therapeutic fields applicable for reaching these cells in liver fbrosis is summarized. Furthermore, recent proceedings and major obstacles in the field of these targeted LNP are presented.

Roles of CCR2 and CCR5 for Hepatic Macrophage Polarization in Mice With Liver Parenchymal Cell-Specific NEMO Deletion.

BACKGROUND & AIMS: Macrophages are key regulators of inflammation and cancer promotion in the liver, and their recruitment and activation is linked to chemokine receptor signaling. However, the exact roles of the chemokine receptors CCR2 and CCR5 for macrophage functions in the liver is obscure. METHODS: To study CCR2 and CCR5 in inflammatory liver injury, we used mice with a hepatocyte-specific knock-out of the nuclear factor κB (NF-κB) essential modulator (NEMO), termed NEMOLPC-KO mice, and generated NEMOLPC-KOCcr2-/- and NEMOLPC-KOCcr5-/- mice. NEMOLPC-KO mice develop hepatitis and fibrosis after two and liver tumors after six months. RESULTS: We found that both CCR2 and CCR5 deficiency led to reduced fibrosis, while CCR5 deficiency increased steatosis and tumor burden in NEMOLPC-KO mice. CCR2 was required for recruitment of hepatic macrophages, whereas CCR5 promoted stellate cell activation. The reduction of monocytes and macrophages by either anti-Gr1 antibody or clodronate-loaded liposomes (CLL), but not of CD8+ T cells or NK cells, significantly aggravated liver injury in NEMOLPC-KO mice and was further increased in NEMOLPC-KOCcr5-/- mice. CLL-induced liver injury was dampened by the adoptive transfer of ex vivo generated macrophages, whereas the adoptive transfer of control CD115+ immature monocytes or B cells did not reduce liver injury. CONCLUSIONS: Although CCR2 and CCR5 principally promote liver fibrosis, they exert differential functions on hepatic macrophages during liver disease progression in NEMOLPC-KO mice. While CCR2 controls the recruitment of monocytes to injured livers, CCR5-dependent functions of liver macrophages limit hepatic injury, thereby reducing steatosis and hepatocarcinogenesis.

Optical imaging of the whole-body to cellular biodistribution of clinical-stage PEG-b-pHPMA-based core-crosslinked polymeric micelles.

Core-crosslinked polymeric micelles (CCPM) based on PEG-b-pHPMA-lactate are clinically evaluated for the treatment of cancer. We macroscopically and microscopically investigated the biodistribution and target site accumulation of CCPM. To this end, fluorophore-labeled CCPM were intravenously injected in mice bearing 4T1 triple-negative breast cancer (TNBC) tumors, and their localization at the whole-body, tissue and cellular level was analyzed using multimodal and multiscale optical imaging. At the organism level, we performed non-invasive 3D micro-computed tomography-fluorescence tomography (µCT-FLT) and 2D fluorescence reflectance imaging (FRI). At the tissue and cellular level, we performed extensive immunohistochemistry, focusing primarily on cancer, endothelial and phagocytic immune cells. The CCPM achieved highly efficient tumor targeting in the 4T1 TNBC mouse model (18.6 %ID/g), with values twice as high as those in liver and spleen (9.1 and 8.9 %ID/g, respectively). Microscopic analysis of tissue slices revealed that at 48 h post injection, 67% of intratumoral CCPM were localized extracellularly. Phenotypic analyses on the remaining 33% of intracellularly accumulated CCPM showed that predominantly F4/80+ phagocytes had taken up the nanocarrier formulation. Similar uptake patterns were observed for liver and spleen. The propensity of CCPM to primarily accumulate in the extracellular space in tumors suggests that the anticancer efficacy of the formulation mainly results from sustained release of the chemotherapeutic payload in the tumor microenvironment. In addition, their high uptake by phagocytic immune cells encourages potential use for immunomodulatory anticancer therapy. Altogether, the beneficial biodistribution, efficient tumor targeting and prominent engagement of PEG-b-pHPMA-lactate-based CCPM with key cell populations underline the clinical versatility of this clinical-stage nanocarrier formulation.

Lipid-encapsulated siRNA for hepatocyte-directed treatment of advanced liver disease.

Lipid-based RNA nanocarriers have been recently accepted as a novel therapeutic option in humans, thus increasing the therapeutic options for patients. Tailored nanomedicines will enable to treat chronic liver disease (CLD) and end-stage liver cancer, disorders with high mortality and few treatment options. Here, we investigated the curative potential of gene therapy of a key molecule in CLD, the c-Jun N-terminal kinase-2 (Jnk2). Delivery to hepatocytes was achieved using a lipid-based clinically employable siRNA formulation that includes a cationic aminolipid to knockdown Jnk2 (named siJnk2). After assessing the therapeutic potential of siJnk2 treatment, non-invasive imaging demonstrated reduced apoptotic cell death and improved hepatocarcinogenesis was evidenced by improved liver parenchyma as well as ameliorated markers of hepatic damage, reduced fibrogenesis in 1-year-old mice. Strikingly, chronic siJnk2 treatment reduced premalignant nodules, indicative of tumor initiation. Furthermore, siJnk2 treatment led to a significant activation of the immune cell compartment. In conclusion, Jnk2 knockdown in hepatocytes ameliorated hepatitis, fibrogenesis, and initiation of hepatocellular carcinoma (HCC), and hence might be a suitable therapeutic option, to define novel molecular targets for precision medicine in CLD.

Risk of falls in postmenopausal women treated with romosozumab: Preliminary indices from a meta-analysis of randomized, controlled trials.

OBJECTIVES: Falls are the well-known risk factor for osteoporotic fractures and some medications can increase the risk of falls. Therefore, the aim of our study is to evaluate the effect of romosozumab on risk of falls in postmenopausal women. METHODS: Studies were searched on PubMed, Cochrane Central Register of Controlled Trials, and ClinicalTrials.gov using the search term "romosozumab." Randomized, clinical trials with romosozumab in postmenopausal women, which met the inclusion criteria and in particular reported on falls in safety or efficacy data, were included into the meta-analysis. Risk ratios (RRs) and corresponding 95% confidence intervals (CIs) were calculated using a binary effects model. RESULTS: A total of four studies with overall 12,128 postmenopausal women with osteoporosis were included into the meta-analysis. Twelve-months treatment with romosozumab reduced the risk of falls nonsignificantly by 16% (RR, 0.84; 95% CI, 0.67-1.04; P = 0.10; n = 11,829). A subgroup analysis with double-blind studies indicated a statistically significant reduction in risk of falls by 20% (RR, 0.80; 95% CI, 0.71-0.92; P ≤ 0.01; n = 11,211). A sequential treatment of romosozumab followed by an antiresorptive medication resulted in a 12% (RR, 0.88; 95% CI, 0.80-0.96; P ≤ 0.01; n = 11,211) reduction of falls in the romosozumab group compared to the control group. CONCLUSIONS: This analysis indicates that romosozumab has a tendency to reduce risk of falls in postmenopausal women with osteoporosis. Nevertheless, our findings are preliminary results with a low significance and to confirm these findings more data and analyses are needed.

Differential Gene Expression in Circulating CD14+ Monocytes Indicates the Prognosis of Critically Ill Patients with Sepsis.

Critical illness and sepsis are characterized by drastic changes in the systemic innate immune response, particularly involving monocytes. The exact monocyte activation profile during sepsis, however, has remained obscure. Therefore, we prospectively analyzed the gene expression profile of circulating CD14+ monocytes from healthy volunteers (n = 54) and intensive care unit (ICU) patients (n = 76), of which n = 36 had sepsis. RNA sequencing of selected samples revealed that monocytes from septic ICU patients display a peculiar activation pattern, which resembles characteristic functional stages of monocyte-derived macrophages and is distinct from controls or non-sepsis ICU patients. Focusing on 55 highly variable genes selected for further investigation, arachidonate 5-lipoxygenase-activating protein (ALOX5AP) was highly upregulated in monocytes of ICU patients and only normalized during 7 days in the ICU in non-sepsis patients. Strikingly, low monocytic guanine nucleotide exchange factor 10-like protein (ARHGEF10L) mRNA expression was associated with the disease severity and mortality of ICU patients. Collectively, our comprehensive analysis of circulating monocytes in critically ill patients revealed a distinct activation pattern, particularly in ICU patients with sepsis. The association with disease severity, the longitudinal recovery or lack thereof during the ICU stay, and the association with prognosis indicate the clinical relevance of monocytic gene expression profiles during sepsis.

Therapeutic Targeting of Neutrophil Granulocytes in Inflammatory Liver Disease.

Neutrophil granulocytes are the most numerous type of leukocyte in humans bearing an enormous, yet largely unexplored therapeutic potential. Scientists have very recently increased their efforts to study and understand these cells which contribute to various types of inflammatory diseases and cancer. The mechanisms that regulate neutrophil recruitment to inflamed tissues and neutrophil cytotoxic activities against host tissues and pathogens require more attention. The reactive oxygen species (ROS) are a popular source of cellular stress and organ injury, and are critically expressed by neutrophils. By combating pathogens using molecular combat factors such as neutrophil extracellular traps (NETs), these are immobilized and killed i.e., by ROS. NETs and ROS are essential for the immune defense, but upon excessive activation, may also harm healthy tissue. Thus, exploring new routes for modulating their migration and activation is highly desired for creating novel anti-inflammatory treatment options. Leukocyte transmigration represents a key process for inflammatory cell infiltration to injury sites. In this review, we briefly summarize the differentiation and roles of neutrophils, with a spotlight on intravital imaging. We further discuss the potential of nanomedicines, i.e., selectin mimetics to target cell migration and influence liver disease outcome in animal models. Novel perspectives further arise from formulations of the wide array of options of small non-coding RNA such as small interfering RNA (siRNA) and micro-RNA (miR) which exhibit enzymatic functions: while siRNA binds and degrades a single mRNA based on full complementarity of binding, miR can up and down-regulate multiple targets in gene transcription and translation, mediated by partial complementarity of binding. Notably, miR is known to regulate at least 60% of the protein-coding genes and thus includes a potent strategy for a large number of targets in neutrophils. Nanomedicines can combine properties of different drugs in a single formulation, i.e., combining surface functionalization with ligands and drug delivery. Inevitably, nanomedicines accumulate in other phagocytes, a fact that should be controlled for every novel formulation to restrain activation of macrophages or modifications of the immunological synapse. Controlled drug release enabled by nanotechnological delivery systems may advance the options of modulating neutrophil activation and migration.

Magnetic-Assisted Treatment of Liver Fibrosis.

Chronic liver injury can be induced by viruses, toxins, cellular activation, and metabolic dysregulation and can lead to liver fibrosis. Hepatic fibrosis still remains a major burden on the global health systems. Nonalcoholic fatty liver disease (NAFLD) and nonalcoholic steatohepatitis (NASH) are considered the main cause of liver fibrosis. Hepatic stellate cells are key targets in antifibrotic treatment, but selective engagement of these cells is an unresolved issue. Current strategies for antifibrotic drugs, which are at the critical stage 3 clinical trials, target metabolic regulation, immune cell activation, and cell death. Here, we report on the critical factors for liver fibrosis, and on prospective novel drugs, which might soon enter the market. Apart from the current clinical trials, novel perspectives for anti-fibrotic treatment may arise from magnetic particles and controlled magnetic forces in various different fields. Magnetic-assisted techniques can, for instance, enable cell engineering and cell therapy to fight cancer, might enable to control the shape or orientation of single cells or tissues mechanically. Furthermore, magnetic forces may improve localized drug delivery mediated by magnetism-induced conformational changes, and they may also enhance non-invasive imaging applications.

Erratum: Hohlstein, P. et al. Prognostic Relevance of Altered Lymphocyte Subpopulations in Critical Illness and Sepsis. Journal of Clinical Medicine 2019, 8, 353.

The authors wish to make the following corrections to this paper [...].