Proliferation Of Monocytes Research Articles

The effect of halofugınone and pirfenidone on wound healing in experimental glaucoma filtration surgery

To investigate the effects of halofuginone and pirfenidone on wound healing in experimental glaucoma filtration surgery (GFS). Animal experimentation. A total of 42 male New Zealand albino rabbits were separated into 6 equal groups. A limbal-based trabeculectomy was performed on 5 groups, and Group I (control group) underwent no surgery and received no postoperative medication. For Group II (sham group), 1 drop 0.9% NaCl was instilled qid for 14 days. For Group III, 1% topical corticosteroid (prednisolone acetate) was instilled 1 drop qid for 14 days. For Group IV, 0.4mg/mL mitomycin-C (MMC) was applied intraoperatively to the region of the scleral flap. For Group V, 0.5% pirfenidone was instilled 1 drop qid for 14 days postoperatively. For Group VI, a sponge soaked in 10ng/mL halofuginone was applied to the surgical region for 3 mins. In addition, 1% topical corticosteroid was instilled ×1 drop qid for 14 days postoperatively for Groups IV, V and VI. After 14 days, sections prepared from the bleb regions of the enucleated eyes were evaluated histopathologically and immunohistochemically. Statistical analyses of the study were performed with Kruskal-Wallis variance analysis and the Mann-Whitney U test. With regard to fibroblasts, suppression of the number of mononuclear cells and immunohistochemical staining intensity of transforming growth factor-b (TGF-β), fibroblast growth factor-b (FGF-β) and platelet derived growth factor (PDGF), the corticosteroid, MMC, pirfenidone and halofuginone groups were seen to exhibit more effect than the sham group (P<0.05). Compared to the pirfenidone and fuginone groups, inhibition of fibroblast and monocyte proliferation was determined to be lower in the MMC group (P<0.05). The intensity of TGF-β and FGF-β staining was seen to be lower in these two treatment groups than in the MMC group (P<0.05). Halofuginone and pirfenidone may be used as effective alternative agents in delaying wound healing in glaucoma filtration surgery.

Type I interferon upregulation and deregulation of genes involved in monopoiesis in chronic myelomonocytic leukemia

Chronic myelomonocytic leukemia (CMML) is characterized by myelomonocytic bias and monocytic proliferation. Whether cell-intrinsic innate immune or inflammatory upregulation mediate disease pathogenesis and phenotype or whether the degree of aberrant monocytic differentiation influences outcomes remains unclear. We compared the transcriptomic features of bone marrow CD34+ cells from 19 patients with CMML and compared to healthy individuals. A total of 1495 genes had significantly differential expression in CMML (q<0.05, fold change>2), including 1271 genes that were significantly upregulated and 224 that were significantly downregulated in CMML. Top upregulated genes were associated with interferon (IFN) alpha and beta signaling, chemokine receptors, IFN gamma, G protein-coupled receptor ligand signaling, and genes involved in immunomodulatory interactions between lymphoid and non-lymphoid cells. Additionally, 6 gene sets were differentially upregulated and 139 were significantly downregulated in patients with myeloproliferative compared to myelodysplastic CMML. A total of 23 genes involved in regulation of monopoiesis were upregulated in CMML compared to healthy controls. We developed a prediction model using Cox regression including 3 of these genes, which differentiated patients into two prognostic subsets with distinct survival outcomes. This data warrants further evaluation of the roles and therapeutic potential of type I IFN signaling and monopoiesis in CMML.

Modulation of proliferation, apoptosis and inflammation of Caco-2 epithelial cells and THP-1 macrophage-like monocytes in LPS stimulated co-culture model.

Epithelial cells and macrophages play major roles in modulating the state of inlammatory response regulated by intracellular signaling pathways. The aim of this study was to characterize changes in cell proliferation, apoptosis and inflammation related intracellular signalling pathways MAPKs and NF-κB in Caco-2 epithelial cells and THP-1 macrophage-like monocytes contacted and filter-seperated co-cultures in the presence of LPS stimulation. We assessed the apoptosis and inflammation by measuring caspase-3 activity, TNF-α and IL-10 cytokines, total and phosphorylated forms of intracellular signalling pathway molecules p53, JNK, Jun, ERK, p38, NF-κB p65 and IkB. The contacted co-culture of Caco-2 and THP-1 cells represented higher levels of JNK, jun and p38 MAPK pathway proteins associated with cell proliferation, whereas apoptosis related molecule caspase-3 and p53 increased in the filter-separated co-culture condition. Also, the contacted co-culture condition led to proinflammatory changes in NF-κB signalling pathways and cytokine responses with high phosphorylated NF-κB p65 and TNF-α, and low I-κB and IL-10 levels. Epithelia - monocytes co-culture stimulated with LPS regulated cell proliferation and inflammation in a contact dependent manner, whereas apoptosis was associated with non-contacted cell culture condition. Thus, co-culture models are important models for explaining the immunopathologies in the mucosal areas (Fig. 5, Ref. 30).

Antioxidant and Biological Activities of Hydroxytyrosol and Homovanillic Alcohol Obtained from Olive Mill Wastewaters of Extra-Virgin Olive Oil Production.

Some constituents of the Mediterranean diet, such as extra-virgin olive oil (EVOO) contain substances such as hydroxytyrosol (HT) and its metabolite homovanillic alcohol (HA). HT has aroused much interest due to its antioxidant activity as a radical scavenger, whereas only a few studies have been made on the HA molecule. Both chemical synthesis and extraction techniques have been developed to obtain these molecules, with each method having its advantages and drawbacks. In this study, we report the use of tyrosol from olive mill wastewaters as a starting molecule to synthesize HT and HA, using a sustainable procedure characterized by high efficiency and low cost. The effects of HT and HA were evaluated on two cell lines, THP-1 human leukemic monocytes and L-6 myoblasts from rat skeletal muscle, after treating the cells with a radical generator. Both HT and HA efficiently inhibited ROS production. In particular, HT inhibited the proliferation of the THP-1 leukemic monocytes, while HA protected L-6 myoblasts from cytotoxicity.

Iguratimod inhibits osteoclastogenesis by modulating the RANKL and TNF-α signaling pathways

BackgroundIguratimod, a small molecular drug, has been proven to have effective bone protection for treatment of patients with bone loss-related diseases, such as rheumatoid arthritis (RA). However, the exact bone protective mechanism of iguratimod remains to be determined. The purpose of this study was to better explore the underlying mechanism of bone protection of iguratimod. MethodsBone marrow monocytes from C57/BL6 mice were stimulated with either RANKL or TNF-α plus M-CSF. The effects of iguratimod on morphology and function of osteoclasts were confirmed by TRAP staining and bone resorption assay, respectively. The expression of osteoclast related genes was detected by RT-PCR and the activation of signal pathway was detected by Western blotting. We used rodent models of osteoporosis (ovariectomy) and of arthritis (modified TNF-α-induced osteoclastogenesis) to evaluate the osteoprotective effect of iguratimod in vivo. ResultsIguratimod potently inhibited osteoclast formation in a dose-dependent manner at the early stage of RANKL-induced osteoclastogenesis, whereas iguratimod had no effect on M-CSF-induced proliferation and RANK expression in bone marrow monocytes. Bone resorption was significantly reduced by both early and late addition of iguratimod. Administration of iguratimod prevented bone loss in ovariectomized mice. The blockage of osteoclastogenesis elicited by iguratimod results from abrogation of the p38、ERK and NF-κB pathways induced by RANKL. Importantly, Iguratimod also dampened TNF-α-induced osteoclastogenesis in vitro and attenuated osteoclasts generation in vivo through disrupting NF-κB late nuclear translocation without interfering with IκBα degradation. ConclusionsIguratimod not only suppresses osteoclastogenesis by interfering with RANKL and TNF-α signals, but also inhibits the bone resorption of mature osteoclasts. These results provided promising evidence for the therapeutic application of iguratimod as a unique treatment option against RA and especially in prevention of bone loss.

Leukocyte immunoglobulin-like receptor A3 is increased in IBD patients and functions as an anti-inflammatory modulator.

Growing evidence shows that a homozygous 6·7-kb deletion of the novel anti-inflammatory molecule leukocyte immunoglobulin-like receptor A3 (LILRA3) is associated with many autoimmune disorders. However, its effects on pathogenesis of inflammatory bowel disease (IBD) have yet not been clarified. LILRA3 is mainly expressed in monocytes, whereas its effects on biological behaviors of monocytes have not been systematically reported. In our study, to investigate the association between LILRA3 polymorphism and IBD susceptibility, LILRA3 polymorphism was assessed in 378 IBD patients and 509 healthy controls. Quantitative real time PCR (qRT-PCR), Western blot and immunohistochemistry (IHC) were employed to detect the LILRA3 expression in IBD patient blood and intestinal samples. The human U937 monocyte cell line was employed to establish LILRA3 over-expressing cells and the effects of LILRA3 on the biological behaviors of U937 cells were systematically explored. Although no association of the polymorphism with IBD development was found, LILRA3 expression was markedly increased in IBD patients compared with healthy controls. Over-expression of LILRA3 in monocytes led to significant decreases in secretion of interferon (IFN)-γ, tumor necrosis factor (TNF)-α and interleukin (IL)-6. Additionally, LILRA3 abated monocyte migration by reducing the expression of several chemokines and enhanced monocyte phagocytosis by increasing CD36 expression. Furthermore, LILRA3 promoted monocyte proliferation through a combination of Akt and extracellular receptor kinase/mitogen-activated protein kinase (Erk/MEK) signaling pathways. We report for the first time, to our knowledge, that LILRA3 is related to IBD and functions as an anti-inflammatory modulator in U937 cells.

Phase 1 Study of Axatilimab (SNDX-6352), a CSF-1R Humanized Antibody, for Chronic Graft-Versus-Host Disease after 2 or More Lines of Systemic Treatment

Phase 1 Study of Axatilimab (SNDX-6352), a CSF-1R Humanized Antibody, for Chronic Graft-Versus-Host Disease after 2 or More Lines of Systemic Treatment

Anti-Myeloma Drug Melflufen Inhibits RANKL-Stimulated Osteoclastogenesis By Suppressing Proliferation of CD14+ Precursor Cells

Background Melflufen (melphalan flufenamide) is a first-in-class peptide-drug conjugate (PDC) that targets aminopeptidases and rapidly releases alkylating agents into tumor cells. Osteolytic lesions are a hallmark of multiple myeloma and occur in almost all MM patients, causing severe bone pain and pathological fractures. Previous reports indicate that melphalan enhances osteoclastogenesis. On other other hand, melflufen is able to suppress proliferation of malignant myeloid cells including osteoclast precursors, monocytes. In the present study we therefore evaluated the direct effects of melflufen and melphalan on human osteoclastogenesis using cultured peripheral blood CD14+ monocytes. Methods CD14+ osteoclast precursors were isolated from peripheral blood. Osteoclastogenesis was stimulated by adding RANKL and M-CSF to CD14+ osteoclast precursors and assessed by tartrate-resistant acid phosphatase (TRAP) staining using para-nitrophenyl phosphate (PNPP). Osteoclastic activity was assessed by two assays. First, TRAP activity was measured by quantification of resorption pits on osteo assay synthetic surface made of crystalline calcium phosphate coating mimicking living bone material. Second, collagen degradation was quantified by release of C-terminal telopeptide of type I collagen (CTx) from mineralized bone slices. The proliferation of mature osteoclasts and their precursors in the presence of melflufen and melphalan was assessed using cytotoxicity assays. RNA isolated from osteoclast precursors and mature osteoclasts was subject to gene expression analysis by microarray. Results Melflufen inhibited RANKL-induced osteoclast formation at 1 µM concentration, while 25 µM melphalan was needed to initiate osteoclastogenesis inhibition. However, only extremely high concentrations of melflufen (10 µM) and melphalan (100 µM) affected mature osteoclasts and their TRAP activity. Cytotoxicity assays have demonstrated that melflufen, but not melphalan, can inhibit proliferation of monocytes, osteoclast progenitor cells, at clinically relevant concentrations. When added during osteoclastogenesis, melflufen has significantly inhibited TRAP phosphatase activity on crystalline calcium phosphate-coated surface at 1 µM, whereas 25 µM of melphalan was required to achieve the same effect. As for collagen degradation, the addition of 1 µM melflufen completely suppressed the CTx fragment release. In a parallel assay, 10 µM of melphalan was necessary to produce the same effect. Noteworthy, we did not observe any positive effect of melphalan on osteoclastogenesis or proliferation of osteoclasts. Gene expression analysis showed that mature osteoclasts express higher levels of bone lytic genes ACP5/TRAP, CTSK, MMP2, MMP7, and MMP9. In addition, CD276 and PDCD1LG2 mRNAs encoding B7 family immune checkpoint molecules B7-H3 and PD-L2 are also strongly up-regulated upon osteoclastogenesis. Discussion Myeloma patients often suffer from severe bone pain associated with osteolytic bone lesions. These lesions are the result of excessive osteoclastogenesis induced by multiple myeloma in the bone marrow microenvironment. Moreover, this process creates an immunosuppressive microenvironment since osteoclasts express immune checkpoint molecules known to negatively control activity of T lymphocytes (PD-L2, B7-H3) and NK cells (B7-H3). In this study, we have shown that a novel anti-myeloma PDC melflufen efficiently inhibits osteoclastogenesis and associated collagen degradation and bone resorption when used at clinically relevant concentrations. Melphalan could also induce a reduction of bone resorption and collagen release, however high and clinically irrelevant concentrations were necessary to obtain these effects. Given these results, it is plausible that melflufen-treated MM patients might achieve a reduction in bone pain, and this should be assessed in more detail in the follow-up studies. Also, elimination of immunosuppressive osteoclasts by melflufen may provide a synergistic effect between melflufen and immunotherapy while treating MM patients. This hypothesis is currently being tested in clinical trials. Disclosures Byrgazov: Oncopeptides AB: Current Employment. Gullbo:Theradex: Current Employment; Oncopeptides AB: Consultancy, Current Employment, Current equity holder in publicly-traded company. Slipicevic:Oncopeptides AB: Current Employment. Lehmann:Oncopeptides AB: Current Employment, Current equity holder in publicly-traded company. Larsson:Oncopeptides AB: Current equity holder in publicly-traded company. Fryknäs:Oncopeptides AB: Research Funding.

Central and local controls of monocytopoiesis influence the outcome of Leishmania infection

Leishmaniases represent a complex of tropical and subtropical diseases caused by an intracellular protozoon of the genus Leishmania. The principal cells controlling the interaction between the host and the parasite Leishmania are monocytes and macrophages, as these cells play a decisive role in establishing the pathogenesis or cure. These cells are involved in controlling the growth of Leishmania and in modulating the adaptive immune responses. The heterogeneity and extensive plasticity of monocytes allow these cells to adjust their functional phenotypes in response to the pathogen-directed immunological cues. In Leishmania-infected host, the rate of myelopoiesis is augmented by enhanced monocytic lineage commitment and proliferation of myeloid progenitor cells both in the BM and at the site of infection. These newly generated monocytes play as “safe haven” for the parasite and also as the antigen-presenting cells for T cells to cause deregulated cytokine production. This altered monocytopoiesis is characterized by tissue-specific immune responses, spatiotemporal dynamics of immunoregulation and functional heterogeneity. In the presence of Th1 cytokines, monocytes exhibit a pro-inflammatory phenotype that protects the host from Leishmania. By contrast, in an environment of Th2 cytokines, monocytes display anti-inflammatory phenotype with pro-parasitic functions. In this review, we summarize the involvement of cytokines in the regulation of monocytopoiesis and differentiation of macrophages during leishmanial infection. Understanding the role of cytokines in regulating interactions between Leishmania and the host monocytes is key to developing new therapeutic interventions against leishmaniases.

Fibroblast growth factor 1 ameliorates adipose tissue inflammation and systemic insulin resistance via enhancing adipocyte mTORC2/Rictor signal.

Obesity‐induced activation and proliferation of resident macrophages and infiltration of circulating monocytes in adipose tissues contribute to adipose tissue inflammation and insulin resistance. These effects further promote the development of metabolic syndromes, such as type 2 diabetes, which is one of the most prevalent health conditions severely threatening human health worldwide. Our study examined the potential molecular mechanism employed by fibroblast growth factor 1 (FGF1) to improve insulin sensitivity. The leptin receptor‐deficient obese mice (db/db) served as an insulin‐resistant model. Our results demonstrated that FGF1‐induced amelioration of insulin resistance in obese mice was related to the decreased levels of pro‐inflammatory adipose tissue macrophages (ATMs) and plasma inflammatory factors. We found that FGF1 enhanced the adipocyte mTORC2/Rictor signalling pathway to inhibit C‐C chemokine ligand 2 (CCL2) production, the major cause of circulating monocytes infiltration, activation and proliferation of resident macrophages in adipose tissues. Conversely, these alleviating effects of FGF1 were substantially abrogated in adipocytes with reduced expression of mTORC2/rictor. Furthermore, a model of adipocyte‐specific mTORC2/Rictor‐knockout (AdRiKO) obese mice was developed to further understand the in vitro result. Altogether, these results demonstrated adipocyte mTORC2/Rictor was a crucial target for FGF1 function on adipose tissue inflammation and insulin sensitivity.

The marine compound and elongation factor 1A1 inhibitor, didemnin B, provides benefit in western diet-induced non-alcoholic fatty liver disease

Inhibition of eukaryotic elongation factor 1A1 (EEF1A1) with the marine compound didemnin B decreases lipotoxic HepG2 cell death in vitro and improves early stage non-alcoholic fatty liver disease (NAFLD) in young genetically obese mice. However, the effects of didemnin B on NAFLD in a model of long-term diet-induced obesity are not known. We investigated the effects of didemnin B on NAFLD severity and metabolic parameters in western diet-induced obese mice, and on the cell types that contribute to liver inflammation and fibrosis in vitro. Male 129S6 mice were fed either standard chow or western diet for 26 weeks, followed by intervention with didemnin B (50 μg/kg) or vehicle by intraperitoneal (i.p.) injection once every 3 days for 14 days. Didemnin B decreased liver and plasma triglycerides, improved oral glucose tolerance, and decreased NAFLD severity. Moreover, didemnin B moderately increased hepatic expression of genes involved in ER stress response (Perk, Chop), and fatty acid oxidation (Fgf21, Cpt1a). In vitro, didemnin B decreased THP-1 monocyte proliferation, disrupted THP-1 monocyte-macrophage differentiation, decreased THP-1 macrophage IL-1β secretion, and decreased hepatic stellate cell (HSteC) proliferation and collagen secretion under both basal and lipotoxic (high fatty acid) conditions. Thus, didemnin B improves hepatic steatosis, glucose tolerance, and blood lipids in obesity, in association with moderate, possibly hormetic, upregulation of pathways involved in cell stress response and energy balance in the liver. Furthermore, it decreases the activity of the cell types implicated in liver inflammation and fibrosis in vitro. These findings highlight the therapeutic potential of partial protein synthesis inhibition in the treatment of NAFLD.

Abstract CT149: SNDX-6352-0502 - A phase 1, open-label, dose escalation trial to investigate the safety, tolerability, pharmacokinetics and pharmacodynamic activity of SNDX-6352 monotherapy in patients with unresectable, recurrent, locally-advanced, or metastatic solid tumors

Abstract Background: Immunosuppressive tumor associated macrophages (TAMs) have been shown to correlate with poor prognosis. SNDX-6352 is a humanized, high-affinity monoclonal IgG4 antibody that blocks dual ligand activation of colony stimulating factor- 1 receptor (CSF-1R), which regulates monocyte migration, proliferation and differentiation into TAMs. By inhibiting and reducing TAM numbers, SNDX-6352 may slow tumor growth and enhance anti-tumor immunotherapeutic approaches. Methods: SNDX-6352-0502 was a multi-center Phase 1 study consisting of Phase 1a (monotherapy) and Phase 1b (combination with durvalumab) in patients with pretreated, advanced solid tumors. The primary objective was determining the MTD and RP2D of SNDX-6352 alone and in combination with durvalumab. Dose regimens of 1, 2, 3 and 6 mg/kg bi-weekly (q2wks) and 6 mg/kg once monthly (q4wks) were administered in 1a, and 1, 2, and 3 mg/kg bi-weekly in 1b. Drug pharmacokinetics (PK) and pharmacodynamic (PD) biomarkers were measured. Results: Thirty-two patients with advanced solid tumors were enrolled with a median age of 61. The median number of prior therapies was 5 (range 2-12) and 72% of patients had ECOG performance status of 1. Median exposure in terms of cycles was 2, maximum was 10. Three patients had SD greater than 4 months. Most common (&amp;gt; 10%) treatment-related AEs were fatigue (31%), periorbital edema (31%) aspartate aminotransferase increased (22%), blood creatinine phosphokinase increased (22%) nausea (13%) and decreased appetite (13%). Grade 3/4 related AEs were observed in 12 patients (38%) with ≥ 2 events in CPK (5), amylase (3), AST (3) and lipase (2). One possibly related SAE of pneumonitis at 6 mg/kg q2wks was reported. DLTs reported in two subjects were G3 fatigue at 2 mg/kg and G3 pneumonitis at 6 mg/kg q2wks. Laboratory abnormalities included increases in blood creatinine phosphokinase, ALT, AST, amylase, and lipase. These abnormalities were all asymptomatic and are consistent with the known class effect of CSF-1Ri. There was no evidence of hepatocellular toxicity. Plasma concentrations of SNDX-6352 increased in a dose-proportional manner with drug accumulation observed at &amp;gt; 1 mg/kg. Plasma CSF1 and IL-34 concentrations increased with treatment and remained elevated at all doses &amp;gt; 1 mg/kg. CSF1 receptor occupancy was saturated at 4 hours post-dose in all treatment cohorts. Circulating non-classical monocytes (CD14+CD16hi) were ablated at all dose levels within 24 hrs and remained suppressed at all doses &amp;gt;1 mg/kg. Conclusion: SNDX-6352 demonstrates tolerability at the highest dose level evaluated (6 mg/kg) with robust PD biomarker modulation at doses as low as 1 mg/kg. A RP2D of 6mg/kg q4wks will be explored in future solid tumor studies. Citation Format: Nilo Azad, Drew Rasco, Sunil Sharma, Matthew Taylor, Christine Quaranto, David L. Tamang, Robert Nordness, Michael L. Meyers, Serap Sankoh, Peter Ordentlich, Anthony W. Tolcher. SNDX-6352-0502 - A phase 1, open-label, dose escalation trial to investigate the safety, tolerability, pharmacokinetics and pharmacodynamic activity of SNDX-6352 monotherapy in patients with unresectable, recurrent, locally-advanced, or metastatic solid tumors [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr CT149.

3148 – SRSF2P95H AND TET2 LOSS CO-OPERATE TO PROMOTE CHRONIC MYELOMONOCYTIC LEUKAEMIA IN VIVO

Sequencing of >3000 MDS and related myeloid neoplasms has provided a detailed picture of the genetic landscape of these cancers. Most harbour mutations in two pathways: the RNA spliceosome (eg. SRSF2, SF3B1, U2AF1) and epigenetic regulators (eg. DMNT3, TET2). Co-mutation of SRSF2 and TET2 are profoundly enriched in MDS and MDS/MPN (p=2.63 × 10-14; q=1.21 × 10-12). In CMML and the related chronic neutrophilic leukaemia and atypical chronic myeloid leukaemia, there is a positive association between mutation of SRSF2 and TET2 (p=0.002). This association is strongest in CMML (p=0.004), where SRSF2 mutations occur in ∼50% of patients and TET2 mutations in ∼60%. SRSF2/TET2 co-mutation influences clinical presentation, with co-mutated CMML having higher leukocyte counts yet less pronounced monocytosis compared to those with wild-type SRSF2/TET2. Based on the human clinical genetics, we crossed the conditional Srsf2P95H/+ mutants with Tet2fl/fl mice to establish animals where we could mutate both concomitantly. At 52 weeks post gene mutation there is a hypocellular bone marrow with increased myeloid bias at the expense of B lymphoid and erythroid lineages in the Srsf2/Tet2 double mutants. Under conditions of native haematopoiesis with aging, a distinct myeloid bias and monocytosis develop in the Srsf2/Tet2 mutants which is not seen in either single mutant or controls. The compound Srsf2/Tet2 mutants display increased granulocytic and monocytic proliferation (myelo-monocytic hyperplasia) with increased immature promonocytes and monoblasts (∼10-15% total nucleated cells) and evidence of binucleate promonocytes. These are features of CMML (CMML-2). Upon secondary transplant, recipients developed leukocytosis, monocytosis and splenomegaly. This demonstrates that our model is able to reproduce SRSF2/TET2 co-operativity in vivo, yielding a disease with core characteristics of CMML, unlike single Srsf2 or Tet2 mutation.

Examination of ex-vivo viability of human adipose tissue slice culture.

Obesity is associated with significantly higher mortality rates, and excess adipose tissue is involved in respective pathologies. Here we established a human adipose tissue slice cultures (HATSC) model ex vivo. HATSC match the in vivo cell composition of human adipose tissue with, among others, mature adipocytes, mesenchymal stem cells as well as stroma tissue and immune cells. This is a new method, optimized for live imaging, to study adipose tissue and cell-based mechanisms of obesity in particular. HATSC survival was tested by means of conventional and immunofluorescence histological techniques, functional analyses and live imaging. Surgery-derived tissue was cut with a tissue chopper in 500 μm sections and transferred onto membranes building an air-liquid interface. HATSC were cultured in six-well plates filled with Dulbecco’s Modified Eagle’s Medium (DMEM), insulin, transferrin, and selenium, both with and without serum. After 0, 1, 7 and 14 days in vitro, slices were fixated and analyzed by morphology and Perilipin A for tissue viability. Immunofluorescent staining against IBA1, CD68 and Ki67 was performed to determine macrophage survival and proliferation. These experiments showed preservation of adipose tissue as well as survival and proliferation of monocytes and stroma tissue for at least 14 days in vitro even in the absence of serum. The physiological capabilities of adipocytes were functionally tested by insulin stimulation and measurement of Phospho-Akt on day 7 and 14 in vitro. Viability was further confirmed by live imaging using Calcein-AM (viable cells) and propidium iodide (apoptosis/necrosis). In conclusion, HATSC have been successfully established by preserving the monovacuolar form of adipocytes and surrounding macrophages and connective tissue. This model allows further analysis of mature human adipose tissue biology ex vivo.

Immunologic timeline of Ebola virus disease and recovery in humans.

A complete understanding of human immune responses to Ebola virus infection is limited by the availability of specimens and the requirement for biosafety level 4 (BSL-4) containment. In an effort to bridge this gap, we evaluated cryopreserved PBMCs from 4 patients who survived Ebola virus disease (EVD) using an established mass cytometry antibody panel to characterize various cell populations during both the acute and convalescent phases. Acute loss of nonclassical monocytes and myeloid DCs, especially CD1c+ DCs, was noted. Classical monocyte proliferation and CD38 upregulation on plasmacytoid DCs coincided with declining viral load. Unsupervised analysis of cell abundance demonstrated acute declines in monocytic, NK, and T cell populations, but some populations, many of myeloid origin, increased in abundance during the acute phase, suggesting emergency hematopoiesis. Despite cell losses during the acute phase, upregulation of Ki-67 correlated with recovery of cell populations over time. These data provide insights into the human immune response during EVD.

Pexidartinib, a Novel Small Molecule CSF-1R Inhibitor in Use for Tenosynovial Giant Cell Tumor: A Systematic Review of Pre-Clinical and Clinical Development.

Tenosynovial giant cell tumor (TGCT) is a rare benign tumor that involves the synovium, bursa, and tendon sheath, resulting in reduced mobility of the affected joint or limb. The current standard of care for TGCT is surgical resection. However, some patients have tumor recurrence, present with unresectable tumors, or have tumors that are in locations where resection could result in amputations or significant debility. Therefore, the development of systemic agents with activity against TGCT to expand treatment options is a highly unmet medical need. Pathologically, TGCT is characterized by the overexpression of colony-stimulating factor 1 (CSF-1), which leads to the recruitment of colony-stimulating factor-1 receptor (CSF-1R) expressing macrophages that make up the primary cell type within these giant cell tumors. The binding of CSF-1 and CSF-1R controls cell survival and proliferation of monocytes and the switch from a monocytic to macrophage phenotype contributing to the growth and inflammation within these tumors. Therefore, molecules that target CSF-1/CSF-1R have emerged as potential systemic agents for the treatment of TGCT. Given the role of macrophages in regulating tumorigenesis, CSF1/CSF1R-targeting agents have emerged as attractive therapeutic targets for solid tumors. Pexidartinib is an orally bioavailable and potent inhibitor of CSF-1R which is one of the most clinically used agents. In this review, we discuss the biology of TGCT and review the pre-clinical and clinical development of pexidartinib which ultimately led to the FDA approval of this agent for the treatment of TGCT as well as ongoing clinical studies utilizing pexidartinib in the setting of cancer.

Surface modification of titanium manufactured through selective laser melting inhibited osteoclast differentiation through mitogen-activated protein kinase signaling pathway.

Selective laser melting used in manufacturing custom-made titanium implants becomes more popular. In view of the important role played by osteoclasts in peri-implant bone resorption and osseointegration, we modified selective laser melting-manufactured titanium surfaces using sandblasting/alkali-heating and sandblasting/acid-etching, and investigated their effect on osteoclast differentiation as well as their underlying mechanisms. The properties of the surfaces, including elements, roughness, wettability and topography, were analyzed. We evaluated the proliferation and morphology of primary mouse bone marrow-derived monocytes, as well as induced osteoclasts derived from bone marrow-derived monocytes, on samples. Then, osteoclast differentiation was determined by the tartrate-resistant acid phosphatase activity assay, calcitonin receptors immunofluorescence staining and the expression of osteoclast-related genes. The results showed that sandblasting/alkali-heating established nanonet structure with the lowest water contact angle, and both sandblasting/alkali-heating and sandblasting/acid-etching significantly decreased surface roughness and heterogeneity compared with selective laser melting. Surface modifications of selective laser melting-produced titanium altered bone marrow-derived monocyte morphology and suppressed bone marrow-derived monocyte proliferation and osteoclastogenesis in vitro (sandblasting/alkali-heating>sandblasting/acid-etching>selective laser melting). These surface modifications reduced the activation of extracellular signal-regulated kinase and c-Jun N-terminal kinases compared to native-selective laser melting. Sandblasting/alkali-heating additionally blocked tumor necrosis factor receptor-associated factor 6 recruitment. The results suggested that sandblasting/alkali-heating and sandblasting/acid-etching modifications on selective laser melting titanium could inhibit osteoclast differentiation through suppressing extracellular signal-regulated kinase and c-Jun N-terminal kinase phosphorylation in mitogen-activated protein kinase signaling pathway and provide a promising technique which might reduce peri-implant bone resorption for optimizing native-selective laser melting implants.

Vitamin E

Vitamin E, the collective name for a group of fat-soluble compounds with distinctive antioxidant activities, is found naturally in some foods, added to others, and available as a dietary supplement. Naturally occurring vitamin E exists in eight chemical forms (alpha-, beta-, gamma-, and delta-tocopherol and alpha-, beta, gamma-, and delta-tocotrienol), but alpha-tocopherol is the only form that meets human dietary requirements. Vitamin E functions as an antioxidant, protecting cells from the damaging effects of free radicals that may contribute to the development of cardiovascular disease and cancer. In addition, vitamin E is involved in immune function, cell signaling, regulation of gene expression, and other metabolic processes. Alpha-tocopherol also inhibits the activity of protein kinase C, an enzyme involved in cell proliferation and differentiation in smooth muscle cells, platelets, and monocytes that helps blood vessels resist components that may adhere to their surface. Vitamin E also increases the expression of two enzymes that suppress arachidonic acid metabolism and may inhibit platelet aggregation. Numerous foods provide vitamin E, with nuts, seeds, and vegetable oils among the best sources of alpha-tocopherol. Significant amounts of alpha-tocoperol are also available in green leafy vegetables and fortified cereals. Most vitamin E in American diets is in the form of gamma-tocopherol from soybean, canola, corn, and other vegetable oils and food products. Naturally sourced vitamin E is referred to as RRR-alpha-tocopherol (or d-alpha-tocopherol); the synthetically produced form is all rac-alpha-tocopherol (or dl-alpha-tocopherol). The RDA for vitamin E varies by age, ranging from 4 mg for young infants (<6 mo) to 15 mg for those 14 years or older. One mg vitamin E (alpha-tocopherol) is equivalent to 1 mg RRR-alpha-tocopherol or 2 mg all rac-alpha-tocopherol. Vitamin E is listed on the new Nutrition Facts and Supplement Facts labels in milligrams rather than in the traditional international units (IUs). FDA required manufacturers to use the new labels starting in January 2020, but companies with annual sales of less than $10 million may continue to use the old labels until January 2021. One IU of natural RRR-alpha-tocopherol is equivalent to 0.67 mg of alpha-tocopherol; 1 IU of synthetic all rac-alpha-tocopherol is equivalent to 0.45 mg alpha-tocopherol. Supplements typically provide only alpha-tocopherol, although “mixed” products containing other tocopherols and tocotrienols are also available. A given amount of synthetic alpha-tocopherol (all rac-alpha-tocopherol, often labeled as “DL” or “dl”) is only half as active as the same amount (by weight in mg) of the natural form (RRR-alpha-tocopherol; often labeled as “D” or “d”). Most vitamin E–only supplements provide 67 mg or greater (100 IU of natural vitamin E), which is higher than the RDA. In addition, alpha-tocopherol in dietary supplements and fortified foods is often esterified to prolong shelf life. These esters (alpha-tocopheryl acetate and succinate) are hydrolyzed and absorbed by the body as efficiently as alpha-tocopherol. The Food and Nutrition Board at the Institute of Medicine of The National Academies suggests that mean intakes of vitamin E among healthy adults are probably higher than the RDA but cautions that low-fat diets might provide insufficient amounts unless people make their food choices carefully by, for example, increasing their intakes of nuts, seeds, fruits, and vegetables. Vitamin E deficiency is rare, and overt deficiency symptoms have not been found in healthy people who obtain little vitamin E from their diets. Because fat is required in the digestive tract to absorb vitamin E, people with fat-malabsorption disorders are more likely to become deficient. Symptoms include peripheral neuropathy, ataxia, skeletal myopathy, retinopathy, and impairment of the immune response. Patients with Crohn’s disease, cystic fibrosis, or an inability to secrete bile from the liver into the digestive tract sometimes require water-soluble forms of vitamin E, such as tocopheryl polyethylene glycol-1000 succinate. Ensure that patients understand that high doses of alpha-tocopherol supplements can cause hemorrhage and interrupt blood coagulation, as well as inhibit platelet aggregation. Doses of up to 1,000 mg/d (1,500 IU/d of the natural form or 1,100 IU/d of the synthetic form) in adults appear to be safe, although data are limited. Because vitamin E may interact with several types of medications, including anticoagulant and antiplatelet meds, simvastatin and niacin, and chemotherapy and radiotherapy drugs, patients should consult with their physician before taking vitamin E supplements. Claims have also been made about vitamin E’s potential to prevent and treat disease, including heart disease, cancer, eye disorders, and cognitive decline. Ensure patients understand that further work is needed to confirm these claims and the safety of vitamin E supplementation.

In This Issue

HomeCirculation ResearchVol. 126, No. 1In This Issue Free AccessIn BriefPDF/EPUBAboutView PDFView EPUBSections ToolsAdd to favoritesDownload citationsTrack citationsPermissions ShareShare onFacebookTwitterLinked InMendeleyReddit Jump toFree AccessIn BriefPDF/EPUBIn This Issue Ruth Williams Ruth WilliamsRuth Williams Search for more papers by this author Originally published2 Jan 2020https://doi.org/10.1161/RES.0000000000000319Circulation Research. 2020;126:2is related toTriglyceride Paradox Is Related to Lipoprotein Size, Visceral Adiposity and Stearoyl-CoA Desaturase Activity in Black Versus White WomenRapamycin-Loaded Biomimetic Nanoparticles Reverse Vascular Inflammationis related toEnriched Marine Oil Supplements Increase Peripheral Blood Specialized Pro-Resolving Mediators Concentrations and Reprogram Host Immune ResponsesBiomimetic Particles Reduce Vascular Inflammation (p 25)Can Atherosclerosis be Reversed? Rapamycin-loaded nanoparticles curb vascular inflammation without toxicity, report Boada et al.Download figureDownload PowerPointIn addition to excessive lipids in the blood, inflammation is a key driver of atherosclerosis. Indeed, plaques grow as a result of fatty deposits and the accumulation and proliferation of monocytes and macrophages in the vessel walls. In mice, such proliferation can be inhibited by the systemic administration of the immunosuppressive compound rapamycin. However, because such treatment can cause toxic side effects, researchers are investigating tissue-localized delivery methods. One such approach suggested by Boada and colleagues is to use rapamycin-loaded leukosomes—nanoparticles encapsulated in a lipid coating containing leukocyte membrane proteins. The coating enables the particles to mimic leukocyte homing such that they accumulate at atherosclerotic plaques. And now the team has shown that with a rapamycin payload, the particles reduce plaque macrophage proliferation better than systemic administration of the drug. Following 7 days of once-daily rapamycin-leukosome administration, plaque burden in atherosclerosis model mice was significantly lower than in control animals. Importantly, no toxicity was detected. These proof-of-principle results now set the stage for further testing in larger, clinically relevant animals, say the authors.Upregulation of Plasma SPM by Enriched Marine Oils (p 75)Can fish oil help human immunity? SPMs indicate the clinical efficacy of fatty acid supplementation, say Souza et al.Download figureDownload PowerPointIn the body, essential fatty acids, such as those found in fish oils, are converted into specialized proresolving mediators (SPMs)—molecules that promote phagocytosis of dead cells and reduce inflammation. Whether taking such oils reduces vascular inflammation in people, however, is unclear. Some studies show the oils reduce inflammation, others show no effect. One problem is that there is no impartial measure of the clinical efficacy of such supplementation. Souza and colleagues, therefore, performed a double-blind, placebo-controlled study to determine the effects of marine oil supplementation on both SPM levels and immune cell function. Twenty-two volunteers were randomly assigned to receive placebo or 1 of 3 doses of marine oil. Blood samples from the subjects revealed a dose-dependent increase in SPMs that was significant in the 2 high-dose groups and peaked a few hours after dosing. The team went on to show that, in the high-dose blood samples, monocytes and neutrophils had increased phagocytic activity while leukocyte activation was decreased. These effects persisted after SPM levels returned to baseline. The results suggest SPMs are not only likely mediators of fatty acid-induced immune effects but that they could be useful efficacy indicators for future trials.Triglyceride Lipoproteins and Insulin Resistance (p 94)Chung et al tackle the triglyceride paradox and discover size of the lipoproteins matters.Download figureDownload PowerPointA high level of triglycerides in the blood is a marker of cardiometabolic disease—a collection of symptoms including insulin resistance, obesity, dyslipidemia, and hypertension. However, this correlation does not occur in people of African ancestry. Indeed, black individuals may have apparently low triglyceride levels even when symptoms of cardiometabolic disease are present. To get to the bottom of this paradox, Chung and colleagues examined triglycerides in blood samples from black and white women to see if a specific subfraction of the lipoproteins—based on particle size—might correlate with symptoms. Women (122) of similar ages, body mass indices, and backgrounds had blood samples collected for insulin sensitivity tests and triglyceride measurements. The team found that while levels of large, medium, and small triglycerides were lower in blacks than whites regardless of insulin sensitivity, levels of very small triglycerides, which are known to be proatherogenic, did correlate with insulin sensitivity in both racial groups. The findings could not only form the basis of more accurate triglyceride tests for assessing cardiovascular disease severity in all races, but also highlight the potential importance of very small triglycerides in contributing to the progression of the condition. Previous Back to top Next FiguresReferencesRelatedDetailsRelated articlesTriglyceride Paradox Is Related to Lipoprotein Size, Visceral Adiposity and Stearoyl-CoA Desaturase Activity in Black Versus White WomenStephanie T. Chung, et al. Circulation Research. 2020;126:94-108Rapamycin-Loaded Biomimetic Nanoparticles Reverse Vascular InflammationChristian Boada, et al. Circulation Research. 2020;126:25-37Enriched Marine Oil Supplements Increase Peripheral Blood Specialized Pro-Resolving Mediators Concentrations and Reprogram Host Immune ResponsesPatricia R. Souza, et al. Circulation Research. 2020;126:75-90 January 3, 2020Vol 126, Issue 1 Advertisement Article InformationMetrics © 2019 American Heart Association, Inc.https://doi.org/10.1161/RES.0000000000000319 Originally publishedJanuary 2, 2020 PDF download Advertisement

Efficacy of a synthetic biomimetic skin substitute of PLLA/gelatin nanofiber membrane in facilitating chronic cutaneous wound healing

ABSTRACT This research was set up to determine the efficacy of a synthetic biomimetic skin substitute of poly-L-lactide (PLLA)/gelatin nanofiber membrane in facilitating chronic cutaneous wound healing. The material physical properties and biological characteristics were evaluated. The safety and effectiveness were further confirmed by clinical applications. Results indicated that the average diameter of the nano pores in the biomimetic substitute was 0.65 μm. The mean contact angle was 80.39°, and the average tensile strength was 4.42 ± 0.43 MPa. The proliferation of fibroblasts and monocytes was significantly improved at day 7 cultured on the substitute. Moreover, the bacterial adhesion on the substitute was decreased. In the clinical case study, the percentage of complete wound healing was 93.3%, and the rate of reoperation was 6.7%. Thus, this technology presents an intriguing possible therapeutic modality that provides pain relief and diminishes the burden of reoperation in the setting of chronic wound management.