Ratio Of M2 Research Articles

Modeling of tumor response to macrophage and T lymphocyte interactions in the liver metastatic microenvironment.

The dynamic interactions between macrophages and T-lymphocytes in the tumor microenvironment exert both antagonistic and synergistic functions affecting tumor growth. Extensive experimental effort has been expended to investigate immunotherapeutic strategies targeting macrophage polarization as well as T-cell activation with the goal to promote tumor cell killing and cancer elimination. However, these interactions remain poorly understood, and cancer immunotherapeutic strategies are often disappointing. The complex system encompassing innate and adaptive immune cell activity in response to tumor growth could benefit from a systems perspective built upon mathematical modeling. This study develops a modeling system to help evaluate the effects of macrophage and T-lymphocyte interactions on tumor growth. The system enables simulating the combined cytotoxic and tumor-promoting interactions of these two immune cell populations in a vascularized organ microenvironment, such as in liver metastases. A hypothetical immunotherapeutic strategy is simulated to increase the number of tumor-suppressive (M1-phenotype) vs. tumor-promoting (M2-phenotype) macrophages to gauge their effects on CD8+ T-cells and CD4+ T-helper cells, which in turn affect the macrophage functions. The results highlight the dynamic interactions between macrophages and T-lymphocytes in the tumor microenvironment and show that with the chosen set of parameter values, the overall cytotoxic effect from macrophages and T-lymphocytes obtained by driving the M1:M2 ratio higher could saturate and fail to achieve tumor regression. Further expansion of this modeling platform to include additional tumor-immune cell interactions, coupled with parameters representing particular tumor characteristics, could enable systematic evaluation of immunotherapeutic strategies tailored to patient-tumor specific conditions, including metastatic disease.

Inhibition of Colony-Stimulating Factor-1 Receptor Enhances the Efficacy of Radiotherapy and Reduces Immune Suppression in Glioblastoma

Glioblastoma (GBM) remains an invariably fatal tumor with high rates of recurrence and progression after radiotherapy (RT). Recent evidence suggests M2 tumor associated macrophages (TAMs) play a contributory role towards the immune suppressive microenvironment of GBM and may mitigate RT-induced anti-tumor immunity. We investigated the inhibition of colony-stimulating factor-1 receptor (CSF-1R) as a potential strategy to target TAMs and improve the efficacy of RT in an immunocompetent pre-clinical GBM model. We used the CSF-1R inhibitor BLZ-945, which is currently under investigation in clinical trials as a mono-therapy for multiple solid tumors. The CSF-1R inhibitor BLZ-945 was used to examine the impact of CSF-1R inhibition on M2 polarization using in vitro assays. Then, using an orthotopic, immunocompetent murine GBM-model, mice were treated with vehicle control, RT, BLZ-945, or RT concurrently with BLZ-945. Survival and changes in the immune landscape were compared between treatment arms. BLZ-945 significantly reduced M2 polarization from bone marrow-derived monocytes in vitro. BLZ-945 alone did not result in improvement in median overall survival (mOS = 29 days) when compared to control mice (mOS = 27 days). RT led to a significant improvement in survival (mOS = 45 days; p = 0.02), while the combination of RT + BLZ-945 led to a further improvement in survival when compared to RT alone (mOS not reached; p = 0.005) with >70% of mice achieving long-term survival. Correlative studies performed on tumors resected from these mice identified a relatively large population of M2 TAMs in GBM at baseline that was further increased in response to RT. BLZ-945 mitigated the increase in M2 infiltration in response to RT. CD8 T-cells also increased significantly in response to RT. The ratio of M2 to CD8 cells was significantly lower in the BLZ-945 treated mice. Inhibition of CSF-1R with BLZ-945 improved the efficacy of RT in the treatment of GBM in a mouse model and may represent a promising strategy to improve RT-induced antitumor immune responses.

Schisandra Inhibit Bleomycin-Induced Idiopathic Pulmonary Fibrosis in Rats via Suppressing M2 Macrophage Polarization.

Idiopathic pulmonary fibrosis (IPF) is defined as a specific form of chronic, progressive fibrosing interstitial pneumonia of unknown cause and limited to the lungs. Schisandrae chinensis fructus (Wuweizi, Schisandra) is commonly used traditional Chinese medicines (TCM) for the treatment of pulmonary fibrosis, bronchitis, and other lung diseases in China. In this study, we investigated the therapeutic effect of Schisandra on IPF which is induced by bleomycin (BLM) in rats and the inhibition of alternatively activated macrophage (M2) polarization. Bleomycin-induced pulmonary fibrosis was used as a model for IPF, and rats were given drug interventions for 7 and 28 days to evaluate the role of Schisandra in the early oxidative phase and late fibrotic phases of BLM-induced pulmonary injury. The data showed that Schisandra exerted protective effects on BLM-induced pulmonary injury in two phases, which were improving inflammatory cell infiltration and severe damages of lung architectures and decreasing markers of M2 subtype. In order to prove the inhibitory effect of Schisandra on M2 polarization, in vitro experiments, we found that Schisandra downregulated the M2 ratio, which confirmed that the polarization of M2 was suppressed. Moreover, Schisandra blocked TGF-β1 signaling in AMs by reducing the levels of Smad3 and Smad4; meanwhile, the upregulation of Smad7 by Schisandra also promoted the effect of inhibition on the TGF-β1/Smad pathway. These results demonstrate that suppression of M2 polarization by Schisandra is associated with the development of IPF in rats.

Abstract 3970: Tumor-associated macrophage, angiogenesis and lymphangiogenesis predicts prognosis in patients with non-small cell lung cancer

Abstract Background: The tumor microenvironment (TME) is a critical player in tumor progression, metastasis and therapy outcome. Tumor-associated macrophages (TAMs) are a well-recognized core element of the TME, and are generally characterized as M2-like macrophages. TAMs are believed to contribute to tumor progression, but the mechanism behind this remains unclear. We aimed to investigate the clinical value, angiogenic and lymphangiogenic significance of TAMs in non-small cell lung cancer (NSCLC). Methods: We assessed CD68, CD163, VEGF-A, and VEGF-C expression in 349 patients with NSCLC by combined immunohistochemistry and digital image analysis. Subsequently, the potential association between M2 TAMs and angiogenetic VEGF-A and/or lymphangiogenetic VEGF-C was evaluated for its prognostic value. Furthermore, the effects of M2 TAMs on angiogenesis and lymphangiogenesis were explored in vitro co-culture system. Results: Elevated M2 (CD163+/CD68+) ratio was significantly associated with poor overall survival (p=0.023). Dual high expression of M2 ratio and VEGF-C was correlated with worse overall survival (p=0.033). Furthermore, CD68 and CD163 expression are positively associated with VEGF-A and/or VEGF-C expression. Multivariate analysis revealed that M2 ratio was an independent predictor of poor survival [HR (95% CI) = 1.53 (1.01-2.33), p=0.046]. Combined M2+/VEGF-C+ expression was also an independent predictor of poor survival [HR (95% CI) = 2.01 (1.28-3.16), p=0.003]. Notably, we confirmed that M2 macrophages enhanced the protein and mRNA expression of both VEGF-A and VEGF-C significantly, while M1 macrophages induced only mRNA expression of VEGF-A significantly in A549 cells. Conclusions: This study suggests that TAMs are significantly associated with angiogenesis or lymphangiogenesis in the progression of NSCLC. Furthermore, elevated M2 ratio and combined M2 and VEGF-C high expression are strong indicators of poor prognosis in patients with NSCLC. Citation Format: Ilseon Hwang, Keun Uk Park, Eun Hee Jung, Hun-Mo Ryoo, Kris Ylaya, Joon-Yong Chung, Stephen M. Hewitt. Tumor-associated macrophage, angiogenesis and lymphangiogenesis predicts prognosis in patients with non-small cell lung cancer [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 3970.

The In Vitro M1/M2 Polarization of Macrophages of BCG-Infected Mice.

Cultured peritoneal macrophages from intact (control) and BCG-infected (experiment) male BALB/c mice were studied 90 days after infection. Polarization of macrophages by M1 (expression of GM-CSF, IFNγ, and CD16/32) and M2 (expression of bFGF and CD36) differentiation pathways was studied with consideration for their the nuclearity class. Mononuclear cells predominated (90% and higher) in macrophage cultures of both groups and presumably, were presented by mainly epithelioid cells. The results indicated polarization of mononuclear and multinuclear macrophages in the M2 direction under conditions of BCG granulomatosis and a higher initial M2 polarization of binuclear macrophages. In control cultures, the ratio of M2 to M1 macrophages was 0.57, in experimental cultures this ratio was 1.6. It seems that long persistence of Mycobacterium tuberculosis in macrophages served as a factor stimulating the plastic processes and transformation of macrophages into epithelioid cells that form the "core" of granulomas and their enlargement upon incorporation of macrophages.

Fas mutation reduces obesity by increasing IL-4 and IL-10 expression and promoting white adipose tissue browning

Brown adipose tissue generates heat via the mitochondrial uncoupling protein UCP1 to protect against obesity and hypothermia. Fas mutant MRL/lpr mice exhibit a significantly leaner phenotype compared to wild type MRL/MpJ mice. In this study, we evaluated the inflammatory cell population in the adipose tissue of MRL/lpr mice, which could potentially influence their lean phenotype. Furthermore, we compared beige fat activity between the MRL/MpJ and MRL/lpr mice. Fas mutation resulted in high body temperature, improved glucose tolerance, and decreased fat mass and adipocyte size. Fas mutation prevented high-fat diet-induced obesity and decreased the white adipose tissue M1:M2 ratio. When mice were fed a high-fat diet, UCP1, IL-4, IL-10, and tyrosine hydroxylase genes had significantly higher expression in Fas-mutant mice than in wild type mice. After a cold challenge, UCP1 expression and browning were also significantly higher in the Fas-mutant mice. In summary, Fas-mutant mice are resistant to high-fat diet-induced obesity due to increased IL-4 and IL-10 levels and the promotion of thermogenic protein activity and browning in their adipose tissues. STAT6 activation might contribute to M2 polarisation by increasing IL-4 and IL-10 levels while increases in M2 and tyrosine hydroxylase levels promote browning in response to Fas mutation.

Inhibition of phosphodiesterase IV enzyme improves locomotor and sensory complications of spinal cord injury via altering microglial activity: Introduction of Roflumilast as an alternative therapy

Despite the great search for an effective approach to minimize secondary injury in spinal cord injury (SCI) setting, there have been limited advances. Roflumilast is a selective inhibitor of phosphodiesterase 4 with potent anti-inflammatory properties. Here, we sought to explore Roflumilast efficacy in the improvement of locomotor and sensory deficits of SCI. In an animal setting, 50 male rats were randomly assigned to five groups: an SCI group receiving Placebo, three SCI groups receiving Roflumilast at the doses of 0.25, 0.5, and 1 mg/kg prior to T9 vertebra laminectomy, and a sham-operated group. Locomotor, mechanical, and thermal activities were evaluated for 28 days. At the end of the study, spinal cord samples were taken to assess the relative ratio of microglial subtypes, including M1 and M2, histopathological changes, levels of pro-inflammatory (TNF-α and IL-1β) and anti-inflammatory (IL-10) biomarkers, and cAMP level. Repeated measure analysis revealed significant effect for time-treatment interaction on locomotion [F (24, 270) = 280.7, p < 0.001], thermal sensitivity [F (16, 180) = 4.35, p < 0.001], and mechanical sensitivity [F (16, 180) = 7.96, p < 0.001]. As expected, Roflumilast significantly increased the expression of spinal cAMP. H&E staining exhibited lesser histopathological disruptions in Roflumilast-treated rodents. We also observed a significant reduction in the M1/M2 ratio (p values < 0.001) as well as in pro-inflammatory biomarkers following the administration of Roflumilast to the injured rats. Furthermore, IL-10 level was increased in rodents receiving 1 mg/kg of the reagent. In conclusion, the increased spinal cAMP following Roflumilast therapy might attenuate neuroinflammation via altering microglial activity; therefore, it could be considered as an alternative therapeutic agent for SCI complications.

Are Islamic indices a viable investment avenue? An empirical study of Islamic and conventional indices in India

PurposeThis study aims to empirically compare the performance of Islamic indices vis-à-vis to their conventional counterparts in India.Design/methodology/approachThe performance of the Islamic and selected conventional indices is evaluated using various risk-adjusted performance measures such as Sharpe ratio, Treynor ratio, M-square (M2) ratio, information ratio, capital asset pricing model (CAPM), Fama-French three-factor model and Carhart four-factor model in India context. The period of study is from December 2006 to 2018.FindingsThe risk-adjusted performance measures based on the Sharpe ratio, Treynor ratio, information ratio, the M2 ratio show that the return of Islamic indices provides slightly superior performance. However, performance investigated using CAPM, Fama-French and Carhart benchmarks produce a statistically insignificant differences in return of the Islamic and conventional benchmarks.Research limitations/implicationsThe Sharīʿah-compliant indices can provide a viable, ethical and alternative investment avenue for faith-based investors as it will not make them worse off in comparison to the conventional benchmarks. This also offers opportunity to conventional investors for portfolio diversification. The promotion of faith-based investment can serve as a tool for financial inclusion to attract a huge segment of Indian population in the formal financial system. The findings of the study suffer from the limitation of small sample size and empirical methods used.Originality/valueThis study contributes to the literature on the comparative performance of Islamic and conventional indices in general and emerging markets, in particular, using most recent data and covering a relatively long span of time. To the best of the knowledge, this is the first comprehensive study examining the performance of Islamic indices, using multiple Islamic indices and various risk-adjusted measures in the Indian context.

1956-P: Inhibition of Tissue Macrophage Proliferation Improves Insulin Resistance in Liver and Adipose Tissue

Objective: Insulin resistance (IR) is based on chronic inflammation in part through the activation of adipose tissue macrophages (Mϕ). Nonalcoholic fatty liver disease (NAFLD) is accepted to be strongly associated with IR. Mϕ in adipose tissue and liver have been reported to proliferate, but the pathophysiological significance of tissue Mϕ proliferation remains unclear. We examined the effects of Mϕ proliferation on IR and tissue inflammation using high-fat fed mice whose Mϕ proliferation was specifically inhibited. Methods and Results: Mϕ specific inhibition of proliferation in mice was achieved by transgenic expression of cyclin-dependent kinase inhibitor 1B (p27kip) under the regulation of scavenger receptor promoter/enhancer. After 10-week high-fat diet load in control and Tg mice, glucose tolerance and IR were assessed, and liver and epididymal white adipose tissue (eWAT) were collected. Significant improvement of IR was showed in Tg mice in ipGTT, ipITT, ipPTT, and tissue Akt phosphorylation after insulin injection. Increased peripheral insulin sensitivity and reduced hepatic glucose production in Tg mice were confirmed by glucose clump. In Immunohistochemistry, proliferation of Mϕ was suppressed in liver and eWAT from Tg mice. Liver triglyceride content decreased in Tg mice. The mRNA expression of M1 Mϕ markers were decreased, and that of M2 Mϕ markers were increased in liver and eWAT of Tg mice. Crown-like structure formation was less observed in Tg eWAT. The mRNA expression of inflammatory cytokines/chemokines was suppressed, and that of M2 Mϕ marker was increased in Tg eWAT. Flow cytometry analysis confirmed decreased M1/​​M2 ratio in liver and eWAT Mϕ from Tg mice. Conclusions: Proliferation of Mϕ is involved in the development of IR in liver and adipose tissue, and the progression of hepatic steatosis. Inhibition of Mϕ proliferation ameliorates tissue inflammation, therefore could potentially be the therapeutic target for IR and type 2 diabetes. Disclosure T. Wada: None. T. Senokuchi: None. S. Yamada: None. Y. Morita: None. H. Motoshima: None. T. Matsumura: None. E. Araki: Advisory Panel; Self; Abbott. Speaker’s Bureau; Self; ARKRAY, Astellas Pharma Inc., AstraZeneca, Eli Lilly Japan K.K., Merck &amp; Co., Inc., Novo Nordisk Inc., WebMD LLC. Other Relationship; Self; Kowa Company, Ltd., Mitsubishi Tanabe Pharma Corporation, Novartis Pharma K.K., Sumitomo Dainippon Pharma Co., Ltd., Takeda Pharmaceutical Company Limited.

199-LB: Diabetes, Macrophage Dysfunction, and Mesh Complications

Objective: Diabetic women who receive polypropylene meshes in the surgical repair of stress urinary incontinence and/or pelvic organ prolapse have a ~5-fold increased risk in developing mesh complications, mainly mesh exposure and pain. Macrophage plays important roles in inflammation and mesh-tissue integration. This study aims to establish an association between diabetes and macrophage dysfunction at the mesh implantation site. Methods: Diabetic (n=20) vs. nondiabetic (n=28) mesh-tissue complexes excised from women with mesh complications were assessed for macrophage phenotype transition, efferocytosis and immune mediator production with TUNEL apoptotic cell labeling, immunofluorescence, NanoString and Luminex multiplex assays. Significance of all statistical tests was set at 0.05 level. Results: Demographic parameters were not different except that BMI was higher in diabetic women (p=0.04). In diabetic vs. nondiabetic specimens, the ratio of pro-healing M2 to pro-inflammatory M1 was 50% lower; the ratio of apoptotic cells/CD68+ macrophages was 50% higher; normal correlations of molecules involved in cell apoptosis (BCL10 and TRAF3) and CD68 were disrupted (Figure); and levels of chemokines CCL11, CCL13, CXCL10 were significantly increased (all p&amp;lt;0.05). Conclusion: Diabetes was associated with macrophage dysfunction at the mesh implantation site, which may contribute to short-term and long-term mesh complications. Disclosure R. Liang: None. S. Lopa: None. Funding Pennsylvania Department of Health

SNOW: Sitravatinib and nivolumab in oral cavity cancer (OCC) window of opportunity study.

6569 Background: Sitravatinib (receptor TKI against TYRO3, AXL, MERTK and VEGF family of receptors) is predicted to increase M1-type tumor-associated macrophages (TAMs) and decrease MDSCs in the tumor microenvironment. SNOW is a window-of-opportunity study evaluating the immunogenic and antitumor effects of preoperative sitravatinib and nivolumab in patients (pts) with OCC. Early results demonstrated the combination was safe and active (Oliva et al, SITC 2019). Biomarker analyses and updated results are presented. Methods: Pts with untreated T2-4a, N0-2 or T1&gt;1cm-N2 OCC are eligible. All pts receive oral sitravatinib 120mg daily from day (D) 1 up to 48h pre-surgery and 1 dose of Nivolumab 240mg on D15. Surgery planned between D23-D30. Standard of care adjuvant radiotherapy given based on clinical stage. Tumor pictures, fresh tumor biopsies, blood samples taken at baseline, D15 and pre-surgery. Tumor flow cytometry and multiplex immunofluorescence staining performed on all biopsies to study changes in immune-cell populations. Tumor whole-exome sequencing (WES) performed on baseline biopsies. Results: As of Jan 31st 2020, 10 out of 12 planned pts were enrolled. Study treatment was well-tolerated: only 1 pt had grade (G) &gt;3 toxicity (hypertension) and 1 pt required surgery delay due to G2 thrombocytopenia. None had intraoperative complications. 1 pt had wound infection and tracheostomy bleeding 11 days post-surgery, possibly-related to study drugs. All pts had tumor reduction, 9/10 had pathological downstaging, including 1 complete response (Table). All had clear margins with no extranodal extension; none required adjuvant chemotherapy. All pts are alive with no recurrence (median follow-up= 69 weeks). Lower % of MDSCs and increased % of M1-TAMs and M1:M2 ratio trend was seen at D15 and pre-surgery, with stronger effect in major responders. Best responders (Pts S1-S2) had higher % of PD-L1+ TAMs at baseline. Tumor WES revealed an HRAS G12D mutation in pt S2 and a BLM mutation (DNA repair) in pt S6 (no downstaging). Conclusions: Pharmacodynamic analyses support the antitumor and immune effects of sitravatinib and nivolumab in OCC. Immune pathological response assessment and transcriptomics are on-going. Clinical trial information: NCT03575598 . [Table: see text]

Ratio of CD68/CD163 in Breast Carcinoma with and without Axillary Lymph Node Metastatic

Tumor and its microenvironment can interact each other. Macrophage is part of tumor microenvironment. New drugs targeting specific superficial receptor of macrophage or cytokine of macrophage polarization have been found. Therefore, macrophage phenotype and its ratio of M1/M2 macrophage need to be identified. This identification could lead us to prognose breast cancer and monitor its therapy. Analytical observational study with cross sectional approach, conducted on paraffin block sample of breast carcinoma from Anatomical Pathology Laboratory of Dr. Soetomo General Hospital. The samples divided into four groups based on nodal metastasis staging (N0, N1, N2, and N3) and stained with antibody against CD68 and CD163. The ratio of CD68 and CD163 were analyzed with Anova test. There were difference expression of CD68 as M1 macrophage marker in various axillary node metastasis groups (p=0.015). There were difference ratio of CD68 as M1 macrophage marker and CD163 as M2 macrophage marker in various axillary node metastasis groups (p=0.005). There were difference ratio of macrophage M1 and macrophage M2 between N0 and N3 group, N2 and N3 group also.

Nonlinear response to cancer nanotherapy due to macrophage interactions revealed by mathematical modeling and evaluated in a murine model via CRISPR-modulated macrophage polarization.

Tumor-associated macrophages (TAMs) have been shown to both aid and hinder tumor growth, with patient outcomes potentially hinging on the proportion of M1, pro-inflammatory/growth-inhibiting, to M2, growth-supporting, phenotypes. Strategies to stimulate tumor regression by promoting polarization to M1 are a novel approach that harnesses the immune system to enhance therapeutic outcomes, including chemotherapy. We recently found that nanotherapy with mesoporous particles loaded with albumin-bound paclitaxel (MSV-nab-PTX) promotes macrophage polarization towards M1 in breast cancer liver metastases (BCLM). However, it remains unclear to what extent tumor regression can be maximized based on modulation of the macrophage phenotype, especially for poorly perfused tumors such as BCLM. Here, for the first time, a CRISPR system is employed to permanently modulate macrophage polarization in a controlled in vitro setting. This enables the design of 3D co-culture experiments mimicking the BCLM hypovascularized environment with various ratios of polarized macrophages. We implement a mathematical framework to evaluate nanoparticle-mediated chemotherapy in conjunction with TAM polarization. The response is predicted to be not linearly dependent on the M1:M2 ratio. To investigate this phenomenon, the response is simulated via the model for a variety of M1:M2 ratios. The modeling indicates that polarization to an all-M1 population may be less effective than a combination of both M1 and M2. Experimental results with the CRISPR system confirm this model-driven hypothesis. Altogether, this study indicates that response to nanoparticle-mediated chemotherapy targeting poorly perfused tumors may benefit from a fine-tuned M1:M2 ratio that maintains both phenotypes in the tumor microenvironment during treatment.

Critical Analysis of Sharpe, Treynor and Jensen Methods in Analyzing Stock Portfolio Performance LQ-45 Stock Studies

The purpose of this study is to determine the stocks of the LQ-45 Index which form a portfolio based on a single index model, analyzed by Sharpe, Treynor, and Jensen methods, whether there are differences in the results of calculations and which is the most appropriate in measuring portfolio performance. The method used in this research is descriptive research with a quantitative approach and the data source used is secondary data from the Indonesia Stock Exchange. The population used is the LQ-45 Index shares for the period 2013-2018 and based on established criteria, a sample of 22 shares was obtained. Based on the methods of Sharpe, Treynor, and Jensen, the portfolio performance that is formed experiences a similar and fluctuating movement. Because the three methods produce almost the same value movement, the determination of slope (linear curve) requires accuracy, therefore re-testing is done by using Robustness Test. With the regression analysis it shows that the Sharpe method is most appropriate to be used in measuring portfolio performance in this study. Researchers suggest for further research, in the formation of portfolios can use other portfolio formation models. This research in the formation of LQ-45 stock portfolios using the single index model method, the portfolio formed is only one portfolio each year which causes less number of portfolios compared. In addition to the Sharpe, Treynor, and Jensen methods, researchers can then use other performance measurement methods for comparisons, such as M2 (Modigliani-Modigliani) and Information Ratio (RI).

Delayed Administration of Angiotensin II Type 2 Receptor (AT2R) Agonist Compound 21 Prevents the Development of Post-stroke Cognitive Impairment in Diabetes Through the Modulation of Microglia Polarization.

A disabling consequence of stroke is cognitive impairment, occurring in 12%-48% of patients, for which there is no therapy. A critical barrier is the lack of understanding of how post-stroke cognitive impairment (PSCI) develops. While 70% of stroke victims present with comorbid diseases such as diabetes and hypertension, the limited use of comorbid disease models in preclinical research further contributes to this lack of progress. To this end, we used a translational model of diabetes to study the development of PSCI. In addition, we evaluated the application of compound 21 (C21), an angiotensin II Type 2 receptor agonist, for the treatment of PSCI by blinding the treatment assignment, setting strict inclusion criteria, and implementing a delayed administration time point. Diabetes was induced by a high-fat diet (HFD) and low-dose streptozotocin (STZ) combination. Control and diabetic rats were subjected to 1 h middle cerebral artery occlusion (MCAO) or sham surgery. Adhesive removal task (ART) and two-trial Y-maze were utilized to test sensorimotor and cognitive function. Three days post-stroke, rats that met the inclusion criteria were administered C21 or vehicle in drinking water at a dose of 0.12 mg/kg/day for 8 weeks. Samples from freshly harvested brains were analyzed by flow cytometry and immunohistochemistry (IHC). Diabetes exacerbated the development of PSCI and increased inflammation and demyelination. Delayed administration of C21 3 days post-stroke reduced mortality and improved sensorimotor and cognitive deficits. It also reduced inflammation and demyelination through modulation of the M1:M2 ratio in the diabetic animals.

PATH-59. HISTOPATHOLOGICAL INVESTIGATION OF THE 1p/19q-CODELETED GLIOMAS RESECTED FOLLOWING ALKYLATING AGENTS CHEMOTHERAPY

Abstract BACKGROUND Little is known about histopathological changes after chemotherapy in lower-grade gliomas (LrGGs). METHODS We investigated 15 1p/19q-codeleted gliomas resected following tumor volume decrease after alkylating agents chemotherapy in comparison with their pre-chemotherapy specimens. Histopathological changes by chemotherapy were evaluated by hematoxilyn-eosin staining and immunohistochemistry for Ki-67/MIB-1, CD68 as pan macrophage/monocyte marker, CD163 as presumed marker of M2 polarity, and nestin and CD133 as markers of glioma stem cells (GSCs). RESULTS Histologically, there were several presumed chemotherapy-related changes in the post-chemotherapy specimens, with the most frequent findings being sparse glial background and abundant foamy cell infiltration. The Ki-67/MIB-1 indices significantly decreased, and CD68+ cells significantly increased after chemotherapy. The increasing rate of CD68+ cells in the post-/pre-chemotherapy specimens was prone to be associated with patients’ progression-free survival (PFS) and overall survival (OS), but not tumor response. The number of CD163+ cells and the ratio of nestin+ cells and CD133+ cells significantly increased after chemotherapy. The number of CD163+ cells, the ratio of nestin+ cells and CD133+ cells, and M2 (CD163+)/M1+M2 (CD68+) ratio in the post-chemotherapy specimens were negatively correlated with patients’ PFS and OS. There was no difference between chemotherapy regimens (temozolomide versus nitrosourea-based) in the number of CD163+ cells and the ratio of nestin+ and CD133+ cells. CONCLUSIONS GSCs in conjunction with M2 macrophages constitute the mechanism of resistance to and recurrence after alkylating agents chemotherapy in LrGGs.

Abstract P2021: Cutaneous Wound Healing in Diabetic Mice is Improved by Topical Mineralocorticoid Receptor Blockade

Introduction: Chronic skin ulcers resulting from impaired wound healing are a serious complication of diabetes. Unresolved inflammation, associated with the dysregulation of both the phenotype and function of macrophages, is involved in the poor healing of diabetic wounds. Objective: To investigate the role of the mineralocorticoid receptor in the healing delay and specifically in the impaired angiogenesis and prolonged inflammation of diabetic wounds. Methods: 6 mm wounds were created on the back skin of female streptozotocin (STZ)-induced or db/db mice. Wounds were treated topically with canrenoate (Canre: 0.5mM) or PBS twice a day, for 5 (STZ mice) or 7 (db/db mice) days. Wound area was measured at indicated time post-wounding. Wounded skin was collected for histology and molecular analysis. Results: topical pharmacological inhibition of the MR by Canre can resolve prolonged inflammation to improve delayed skin wound healing in diabetic mouse models. The improvement is demonstrated by increasing of 39% of neo-epidermisation and 62% of epidermal keratinocyte proliferation in db/db wounds treated by Canre. Moreover, Canre could improve the reduced blood vessel density in the wound beds of STZ mice (STZ + PBS: 5.33% ± 0.32 vs STZ + Canre: 8.26% ± 0.64) and of db/db mice (db/db + PBS: 1.32% ± 0.29 vs db/db + Canre: 3.31% ± 0.33). The beneficial effect of Canre is associated with an increased ratio of anti-inflammatory M2 macrophages to pro-inflammatory M1 macrophages in diabetic wounds (ratio of M2 to M1 in db/db + PBS vs db/db + Canre: 0.72 ± 0.15 vs 1.72 ± 0.29, respectively). Furthermore, we show that MR blockade leads to downregulated expression of a MR target, lipocalin 2 (Lcn2), which may facilitate macrophage polarization towards the M2 phenotype and promote impaired angiogenesis in diabetic wounds. Indeed, diabetic Lcn2-deficient mice showed improved wound healing, associated with macrophage M2 polarization and angiogenesis. In addition, recombinant Lcn2 protein prevented IL4-induced macrophages switch from M1 to M2 phenotype. Conclusion: topical MR blockade accelerates skin wound healing in diabetic mice via Lcn2 reduction, M2 macrophage polarization, prevention of prolonged inflammation, and induction of angiogenesis.

Avian Influenza A Virus Polymerase Recruits Cellular RNA Helicase eIF4A3 to Promote Viral mRNA Splicing and Spliced mRNA Nuclear Export.

The influenza A virus replicates in a broad range of avian and mammalian species by hijacking cellular factors and processes. Avian influenza A viruses (AIVs) generally propagated poorly in mammalian cells, but some mutants of virus-encoded RNA polymerase components, especially PB2 subunit, can overcome host restriction. Host factors associated with PB2 may be essential for efficient AIV replication in mammalian cells. Here, we infected human cells with the PB2 Flag-tagged replication-competent recombinant AIV and identified cellular proteins that coprecipitate with PB2 protein by mass spectrometry. We confirmed one of the coprecipitating host factors, DEAD-box protein eIF4A3, that interacts with viral PB2, PB1, and NP proteins. Depletion of endogenous eIF4A3 significantly reduced virus replication. Later studies showed that eIF4A3 is essential for viral RNA polymerase activity and viral RNAs synthesis. Upon systematic dissection of the influenza virus progeny mRNA generation, from pre-mRNA processing to nuclear export, we found that the depletion of eIF4A3 resulted in significant defects in the ratio of M2 to M1 and NS2 to NS1, and the proportion of viral spliced mRNA in the nucleus increased, indicating that eIF4A3 plays a significant function in viral nascent intron mRNA splicing and spliced mRNA (M2 and NS2) nuclear export. Additionally, we confirmed that in specific deletion of eIF4A3, the synthesis of reduced NS2 can significantly impair neo-synthetized viral ribonucleoprotein (vRNP) nuclear export. Taken together, our findings revealed that eIF4A3 is a key mediator of AIV polymerase activity, mRNA splicing, and spliced mRNA nuclear export.

Tumor-associated macrophage infiltration in meningioma.

BackgroundMeningioma, a most common brain tumor, has a high rate of recurrence. Tumor-associated macrophages (TAMs) are the most abundant immune cell type in meningioma. TAMs display functional phenotypic diversity and may establish either an inflammatory and anti-tumoral or an immunosuppressive and pro-tumoral microenvironment. TAM subtypes present in meningioma and potential contribution to growth and recurrence is unknown.MethodsImmunofluorescence staining was used to quantify M1 and M2 TAM populations in tissues obtained from 30 meningioma patients. Associations between M1 and M2 cells, M1:M2 cell ratio to tumor characteristics, WHO grade, recurrence, size, location, peri-tumoral edema, and patient demographics such as age and sex were examined.ResultsTAM cells accounted for ~18% of all cells in meningioma tissues. More than 80% of infiltrating TAMs were found to be of pro-tumoral M2 phenotype and correlated to tumor size (P = .0409). M1:M2 cell ratio was significantly decreased in WHO grade II, compared to grade I tumors (P = .009). Furthermore, a 2.3-fold difference in M1:M2 ratio between primary (0.14) and recurrent (0.06) tumors was observed (n = 18 and 12 respectively, P = .044).ConclusionThis study is the first to confirm existence of pro-tumoral M2 TAMs in the meningioma microenvironment, emphasizing its potential role in tumor growth and recurrence.

PAM3 supports the generation of M2-like macrophages from lupus patient monocytes and improves disease outcome in murine lupus

Abstract Systematic Lupus Erythematosus (SLE) is an autoimmune syndrome of unclear etiology. While T and B cell abnormalities contribute to disease pathogenesis, recent work suggests that inflammatory M1-like macrophages also play a role. Previous work showed that the TLR2/1 agonist PAM3CSK4 (PAM3) could stimulate normal human monocytes to preferentially differentiate into immunosuppressive M2-like rather than inflammatory M1-like macrophages. This raised the possibility of PAM3 being used to normalize the M1:M2 ratio in SLE. Consistent with that possibility, monocytes from lupus patients differentiated into M2-like macrophages when treated with PAM3 in vitro. Furthermore, lupus-prone NZB x NZW F1 mice responded similarly to weekly PAM3 treatment. Normalization of the M2 macrophage frequency was associated with delayed disease progression, decreased autoantibody and inflammatory cytokine synthesis, reduced proteinuria and prolonged survival in NZB x NZW F1 mice. The ability of PAM3 to bias monocyte differentiation in favor of immunosuppressive macrophages may represent a novel approach to the therapy of SLE.