Additive amelioration of ALS by co-targeting independent pathogenic mechanisms.

OBJECTIVE: Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease in which glia are central mediators of motor neuron (MN) death. Since multiple cell types are involved in disease pathogenesis, the objective of this study was to determine the benefit of co-targeting independent pathogenic mechanisms in a familial ALS mouse model. METHODS: Recently, our laboratory identified that ALS microglia induce MN death in an NF-κB-dependent mechanism. We also demonstrated that a single, post-natal, intravenous injection of adeno-associated viral vector serotype 9 encoding a shRNA against mutant SOD1 is able to traverse the blood-brain barrier of ALS mice and reduce SOD1-expression in astrocytes and MNs. Reducing mutant SOD1 in MNs and astrocytes led to a robust increase in survival. To evaluate the benefit of co-targeting multiple cell types in ALS, we combined microglial NF-κB suppression with SOD1 reduction in astrocytes and MNs. RESULTS: Targeting both astrocytes and microglia resulted in an additive increase in survival and motor function by delaying both onset and progression. Strikingly, targeting all three cell types (astrocytes, motor neurons [MNs], and microglia) resulted in an additive increase in lifespan and motor function, with maximum survival reaching 204 days, 67 days longer than the mean survival of untreated control animals. INTERPRETATION: Our data suggest that a combinatorial approach co-targeting different pathogenic mechanisms in independent cell types is a beneficial therapeutic strategy for ALS

Vascular endothelial growth factor restores delayed tumor progression in tumors depleted of macrophages

Genetic depletion of macrophages in Polyoma Middle T oncoprotein (PyMT)‐induced mammary tumors in mice delayed the angiogenic switch and the progression to malignancy. To determine whether vascular endothelial growth factor A (VEGF‐A) produced by tumor‐associated macrophages regulated the onset of the angiogenic switch, a genetic approach was used to restore expression of VEGF‐A into tumors at the benign stages. This stimulated formation of a high‐density vessel network and in macrophage‐depleted mice, was followed by accelerated tumor progression. The expression of VEGF‐A led to a massive infiltration into the tumor of leukocytes that were mostly macrophages. This study suggests that macrophage‐produced VEGF regulates malignant progression through stimulating tumor angiogenesis, leukocytic infiltration and tumor cell invasion

α-Mangostin Effectively Inhibits Chikungunya Virus Replication in HepG2 Cells

Chikungunya virus (CHIKV) is an arthropod-transmitted Alphavirus endemic to countries in Africa and Asia, including Indonesia, which causes debilitating arthralgia which can last several years. The rapid spread of CHIKV to new areas makes the discovery of antiviral agents a high priority. α-mangostin is a xanthone from mangosteen (Garcinia mangostana) pericarp and has antiviral activity against Hepatitis C and Dengue viruses. We investigated the antiviral activity of α-mangostin against CHIKV in HepG2 cells in pre-, post- and combination treatments compared to the common antiviral medicine ribavirin, as well their cytotoxicity. Our results show dose-responsive reductions in viral titer in all treatment regimes, with post- and combination treatments being more effective than pre-treatment only (IC50 = 7.79, 5.99 and 6.39 µM, respectively), but with poor specificity (SI = 1.39, 1.81 and 1.70, respectively) compared to ribavirin. Neither compound showed a direct virucidal effect. These results suggest α-mangostin effectively inhibits CHIKV replication in this cell line

Mena deficiency delays tumor progression and decreases metastasis in polyoma middle-T transgenic mouse mammary tumors

Introduction The actin binding protein Mammalian enabled (Mena), has been implicated in the metastatic progression of solid tumors in humans. Mena expression level in primary tumors is correlated with metastasis in breast, cervical, colorectal and pancreatic cancers. Cells expressing high Mena levels are part of the tumor microenvironment for metastasis (TMEM), an anatomical structure that is predictive for risk of breast cancer metastasis. Previously we have shown that forced expression of Mena adenocarcinoma cells enhances invasion and metastasis in xenograft mice. Whether Mena is required for tumor progression is still unknown. Here we report the effects of Mena deficiency on tumor progression, metastasis and on normal mammary gland development. Methods To investigate the role of Mena in tumor progression and metastasis, Mena deficient mice were intercrossed with mice carrying a transgene expressing the polyoma middle T oncoprotein, driven by the mouse mammary tumor virus. The progeny were investigated for the effects of Mena deficiency on tumor progression via staging of primary mammary tumors and by evaluation of morbidity. Stages of metastatic progression were investigated using an in vivo invasion assay, intravital multiphoton microscopy, circulating tumor cell burden, and lung metastases. Mammary gland development was studied in whole mount mammary glands of wild type and Mena deficient mice. Results Mena deficiency decreased morbidity and metastatic dissemination. Loss of Mena increased mammary tumor latency but had no affect on mammary tumor burden or histologic progression to carcinoma. Elimination of Mena also significantly decreased epidermal growth factor (EGF) induced in vivo invasion, in vivo motility, intravasation and metastasis. Non-tumor bearing mice deficient for Mena also showed defects in mammary gland terminal end bud formation and branching. Conclusions Deficiency of Mena decreases metastasis by slowing tumor progression and reducing tumor cell invasion and intravasation. Mena deficiency during development causes defects in invasive processes involved in mammary gland development. These findings suggest that functional intervention targeting Mena in breast cancer patients may provide a valuable treatment option to delay tumor progression and decrease invasion and metastatic spread leading to an improved prognostic outcome.National Cancer Institute (U.S.). Integrative Cancer Biology Program (grant U54 CA112967)Virginia and D.K. Ludwig Fund for Cancer Researc

Infeksi Virus Dengue pada Nyamuk Aedes aegypti Menggunakan Artificial Blood Feeding dan Deteksi Virus Dengue Menggunakan Teknik Molekular

Abstract. Artificial blood-feeding using the parafilm-M membrane can be used as an alternative solution andsubstitute live animals as a source of blood. This method is not only be used for blood-feeding but also to infectthe dengue virus (DENV) to mosquitoes. This study was aimed to determine the effectiveness artificial bloodfeeding using parafilm-M membrane in Aedes mosquitoes originated in Indonesia and determine the positivityof mosquitoes infected by Indonesia DENV-1. DENV-1 was isolated from patient and propagated in Vero cellculture. The feeding was done in cardboard cups after mosquitos have been starved for 4-17 hours before beingfed with human blood. A conical 50ml tube was prepared, and a hole was created in the tube lid. The tubeopening was covered with parafilm. Glycerol was added into conical tube and heated in water bath for an hourat 55oC. A mixture of blood and DENV-1 was made with concentration of 10%. Detection of DENV in bloodfedmosquitos was carried out by using Simplexa Dengue Real-Time RT-PCR assay. The results showed thatthe prevalence of blood-fed mosquitoes reached 66.67% with fasting period for 17 hours. Blood feedingmosquitoes are affected by duration of fasting period, blood-feeding time, and attractants from human skinrubbed into parafilm-M membrane. The prevalence of blood-fed Ae. aegypti infected by DENV was 20.83%.This study provides information on the effectiveness of artificial parafilm membrane blood-feeding in alaboratory setting that will be useful for vector control study in Indonesia

Siglec-H is a microglia-specific marker that discriminates microglia from CNS-associated macrophages and CNS-infiltrating monocytes

International audienceSeveral types of myeloid cell are resident in the CNS. In the steady state, microglia are present in the CNS parenchyma, whereas macrophages reside in boundary regions of the CNS, such as perivascular spaces, the meninges and choroid plexus. In addition, monocytes infiltrate into the CNS parenchyma from circulation upon blood-brain barrier breakdown after CNS injury and inflammation. Although several markers, such as CD11b and ionized calcium-binding adapter molecule 1 (Iba1), are frequently used as microglial markers, they are also expressed by other types of myeloid cell and microglia-specific markers were not defined until recently. Previous transcriptome analyses of isolated microglia identified a transmembrane lectin, sialic acid-binding immunoglobulin-like lectin H (Siglec-H), as a molecular signature for microglia; however, this was not confirmed by histological studies in the nervous system and the reliability of Siglec-H as a microglial marker remained unclear. Here, we demonstrate that Siglec-H is an authentic marker for microglia in mice by immunohistochemistry using a Siglec-H-specific antibody. Siglec-H was expressed by parenchymal microglia from developmental stages to adulthood, and the expression was maintained in activated microglia under injury or inflammatory condition. However, Siglec-H expression was absent from CNS-associated macrophages and CNS-infiltrating monocytes, except for a minor subset of cells. We also show that the Siglech gene locus is a feasible site for specific targeting of microglia in the nervous system. In conclusion, Siglec-H is a reliable marker for microglia that will allow histological identification of microglia and microglia-specific gene manipulation in the nervous system

Dengue score: a proposed diagnostic predictor for pleural effusion and/or ascites in adults with dengue infection

BACKGROUND: There are several limitations in diagnosing plasma leakage using the World Health Organization (WHO) guidelines of dengue hemorrhagic fever. We conducted a study to develop a dengue scoring system to predict pleural effusion and/or ascites using routine laboratory parameters. METHODS: A prospective observational study was carried out at Cipto Mangunkusumo Hospital and Persahabatan Hospital, Jakarta, Indonesia. Dengue-infected adults admitted on the third febrile day from March, 2010 through August, 2015 were included in the study. A multivariate analysis was conducted to determine the independent diagnostic predictors of pleural effusion and/or ascites and to convert the prediction model into a scoring system. RESULTS: A total of 172 dengue-infected adults were enrolled in the study. Of the 172 patients, 101 (58.7 %) developed pleural effusion and/or ascites. A multivariate analysis was conducted to determine the independent diagnostic predictors of pleural effusion and/or ascites in dengue-infected adults. The predictors were scored based on the following calculations: hemoconcentration ≥15.1 % had a score of 1 (OR, 3.11; 95 % CI, 1.41–6.88), lowest albumin concentration at critical phase ≤3.49 mg/dL had a score of 1 (OR, 4.48; 95 % CI, 1.87–10.77), lowest platelet count ≤49,500/μL had a score of 1 (OR, 3.62; 95 % CI, 1.55–8.49), and elevated ratio of AST ≥2.51 had a score of 1 (OR 2.67; 95 % CI, 1.19–5.97). At a cut off of ≥ 2, the Dengue Score predicted pleural effusion and/or ascites diagnosis with positive predictive value of 79.21 % and negative predictive value of 74.63 %. This prediction model is suitable for calibration and good discrimination. CONCLUSIONS: We have developed a Dengue Score that could be used to identify pleural effusion and/or ascites and might be useful to stratify dengue-infected patients at risk for developing severe dengue

Real-Time Imaging Reveals the Dynamics of Leukocyte Behaviour during Experimental Cerebral Malaria Pathogenesis

During experimental cerebral malaria (ECM) mice develop a lethal neuropathological syndrome associated with microcirculatory dysfunction and intravascular leukocyte sequestration. The precise spatio-temporal context in which the intravascular immune response unfolds is incompletely understood. We developed a 2-photon intravital microscopy (2P-IVM)-based brain-imaging model to monitor the real-time behaviour of leukocytes directly within the brain vasculature during ECM. Ly6Chi monocytes, but not neutrophils, started to accumulate in the blood vessels of Plasmodium berghei ANKA (PbA)-infected MacGreen mice, in which myeloid cells express GFP, one to two days prior to the onset of the neurological signs (NS). A decrease in the rolling speed of monocytes, a measure of endothelial cell activation, was associated with progressive worsening of clinical symptoms. Adoptive transfer experiments with defined immune cell subsets in recombinase activating gene (RAG)-1-deficient mice showed that these changes were mediated by Plasmodium-specific CD8+ T lymphocytes. A critical number of CD8+ T effectors was required to induce disease and monocyte adherence to the vasculature. Depletion of monocytes at the onset of disease symptoms resulted in decreased lymphocyte accumulation, suggesting reciprocal effects of monocytes and T cells on their recruitment within the brain. Together, our studies define the real-time kinetics of leukocyte behaviour in the central nervous system during ECM, and reveal a significant role for Plasmodium-specific CD8+ T lymphocytes in regulating vascular pathology in this disease. © 2014 Pai et al

Antiviral activities of curcumin and 6‐gingerol against infection of four dengue virus serotypes in A549 human cell line in vitro

Dengue virus (DENV) is the most geographically widespread arbovirus causing dengue disease epidemics in tropical and subtropical regions. Nature provides abundant plants as a source for lead molecules against various diseases including DENV infection. We investigated the antiviral effect of curcumin and 6‐gingerol, the major active constituent of turmeric (Curcuma longa Linn.) and ginger (Zingiber officinale Roscoe), respectively, against all four serotypes of DENV infecting human lung epithelial carcinoma (A549) cell line in vitro. Both compounds generated cell cytotoxicity to A549 cells at CC50 values of 108 µM for curcumin and 210 µM for 6‐gingerol. The compound curcumin showed antiviral properties as described by IC50 of 20.60, 13.95, 25.54, and 12.35 µM, while 6‐gingerol of 14.70, 14.17, 78.76, and 112.84 µM for DENV‐1, ‐2, ‐3, and ‐4, respectively. Different levels of antiviral properties were observed between DENV serotypes. Our findings suggest that the antiviral assay of compounds against DENV should be performed to all four serotypes and not limited to a particular serotype. In conclusion, curcumin and 6‐gingerol exhibit antiviral properties against DENV infection and could provide a new therapeutic approach for dengue disease treatment strategies

Chikungunya virus infection in Indonesia: a systematic review and evolutionary analysis

Abstract Background Despite the high number of chikungunya cases in Indonesia in recent years, comprehensive epidemiological data are lacking. The systematic review was undertaken to provide data on incidence, the seroprevalence of anti-Chikungunya virus (CHIKV) IgM and IgG antibodies, mortality, the genotypes of circulating CHIKV and travel-related cases of chikungunya in the country. In addition, a phylogenetic and evolutionary analysis of Indonesian CHIKV was conducted. Methods A systematic review was conducted to identify eligible studies from EMBASE, MEDLINE, PubMed and Web of Science as of October 16th 2017. Studies describing the incidence, seroprevalence of IgM and IgG, mortality, genotypes and travel-associated chikungunya were systematically reviewed. The maximum likelihood phylogenetic and evolutionary rate was estimated using Randomized Axelerated Maximum Likelihood (RAxML), and the Bayesian Markov chain Monte Carlo (MCMC) method identified the Time to Most Recent Common Ancestors (TMRCA) of Indonesian CHIKV. The systematic review was registered in the PROSPERO database (CRD42017078205). Results Chikungunya incidence ranged between 0.16-36.2 cases per 100,000 person-year. Overall, the median seroprevalence of anti-CHIKV IgM antibodies in both outbreak and non-outbreak scenarios was 13.3% (17.7 and 7.3% for outbreak and non-outbreak events, respectively). The median seroprevalence of IgG antibodies in both outbreak and non-outbreak settings was 18.5% (range 0.0–73.1%). There were 130 Indonesian CHIKV sequences available, of which 120 (92.3%) were of the Asian genotype and 10 (7.7%) belonged to the East/Central/South African (ECSA) genotype. The ECSA genotype was first isolated in Indonesia in 2008 and was continually sampled until 2011. All ECSA viruses sampled in Indonesia appear to be closely related to viruses that caused massive outbreaks in Southeast Asia countries during the same period. Massive nationwide chikungunya outbreaks in Indonesia were reported during 2009–2010 with a total of 137,655 cases. Our spatio-temporal, phylogenetic and evolutionary data suggest that these outbreaks were likely associated with the introduction of the ECSA genotype of CHIKV to Indonesia. Conclusions Although no deaths have been recorded, the seroprevalence of anti-CHIKV IgM and IgG in the Indonesian population have been relatively high in recent years following re-emergence in early 2001. There is sufficient evidence to suggest that the introduction of ECSA into Indonesia was likely associated with massive chikungunya outbreaks during 2009–2010.https://deepblue.lib.umich.edu/bitstream/2027.42/148282/1/12879_2019_Article_3857.pd