Langerhans cells are negative regulators of the anti-Leishmania response

Langerhans cells suppress the immune response to low-dose Leishmania major infection in part by inducing regulatory T cells

Treatment of melanoma cells with the synthetic retinoid CD437 induces apoptosis via activation of AP-1 in vitro, and causes growth inhibition in xenografts in vivo

Human malignant melanoma is notoriously resistant to pharmacological modulation. We describe here for the first time that the synthetic retinoid CD437 has a strong dose-dependent antiproliferative effect on human melanoma cells (IC50: 5 x 10(-6) M) via the induction of programmed cell death, as judged by analysis of cell morphology, electron microscopical features, and DNA fragmentation. Programmed cell death was preceded by a strong activation of the AP-1 complex in CD437-treated cells as demonstrated by gel retardation and chloramphenicol transferase (CAT) assays. Northern blot analysis showed a time-dependent increase in the expression of c-fos and c-jun encoding components of AP-1, whereas bcl-2 and p53 mRNA levels remained constant. CD437 also exhibited a strong growth inhibitory effect on MeWo melanoma cells in a xenograft model. In tissue sections of CD437-treated MeWo tumors from these animals, apoptotic melanoma cells and c-fos overexpressing cells were colocalized by TdT-mediated deoxyuridine triphosphate-digoxigenin nick end labeling (TUNEL) staining and in situ hybridization. Taken together, this report identifies CD437 as a retinoid that activates and upregulates the transcription factor AP-1, leading eventually to programmed cell death of exposed human melanoma cells in vitro and in vivo. Further studies are needed to evaluate whether synthetic retinoids such as CD437 represent a new class of retinoids, which may open up new ways to a more effective therapy of malignant melanoma

Suppression of LPS-induced inflammatory responses in macrophages infected with Leishmania

<p>Abstract</p> <p>Background</p> <p>Chronic inflammation activated by macrophage innate pathogen recognition receptors such as TLR4 can lead to a range of inflammatory diseases, including atherosclerosis, Crohn's disease, arthritis and cancer. Unlike many microbes, the kinetoplastid protozoan pathogen <it>Leishmania </it>has been shown to avoid and even actively suppress host inflammatory cytokine responses, such as LPS-induced IL-12 production. The nature and scope of <it>Leishmania</it>-mediated inflammatory cytokine suppression, however, is not well characterized. Advancing our knowledge of such microbe-mediated cytokine suppression may provide new avenues for therapeutic intervention in inflammatory disease.</p> <p>Methods</p> <p>We explored the kinetics of a range of cytokine and chemokine responses in primary murine macrophages stimulated with LPS in the presence versus absence of two clinically distinct species of <it>Leishmania </it>using sensitive multiplex cytokine analyses. To confirm that these effects were parasite-specific, we compared the effects of <it>Leishmania </it>uptake on LPS-induced cytokine expression with uptake of inert latex beads.</p> <p>Results</p> <p>Whilst <it>Leishmania </it>uptake alone did not induce significant levels of any cytokine analysed in this study, <it>Leishmania </it>uptake in the presence of LPS caused parasite-specific suppression of certain LPS-induced pro-inflammatory cytokines, including IL-12, IL-17 and IL-6. Interestingly, <it>L. amazonensis </it>was generally more suppressive than <it>L. major</it>. We also found that other LPS-induced proinflammatory cytokines, such as IL-1α, TNF-α and the chemokines MIP-1α and MCP-1 and also the anti-inflammatory cytokine IL-10, were augmented during <it>Leishmania </it>uptake, in a parasite-specific manner.</p> <p>Conclusions</p> <p>During uptake by macrophages, <it>Leishmania </it>evades the activation of a broad range of cytokines and chemokines. Further, in the presence of a strong inflammatory stimulus, <it>Leishmania </it>suppresses certain proinflammatory cytokine responses in a parasite-specific manner, however it augments the production of other proinflammatory cytokines. Our findings highlight the complexity of inflammatory cytokine signalling regulation in the context of the macrophage and <it>Leishmania </it>interaction and confirm the utility of the <it>Leishmania</it>/macrophage infection model as an experimental system for further studies of inflammatory regulation. Such studies may advance the development of therapies against inflammatory disease.</p

Keratinocytes Determine Th1 Immunity during Early Experimental Leishmaniasis

Experimental leishmaniasis is an excellent model system for analyzing Th1/Th2 differentiation. Resistance to Leishmania (L.) major depends on the development of a L. major specific Th1 response, while Th2 differentiation results in susceptibility. There is growing evidence that the microenvironment of the early affected tissue delivers the initial triggers for Th-cell differentiation. To analyze this we studied differential gene expression in infected skin of resistant and susceptible mice 16h after parasite inoculation. Employing microarray technology, bioinformatics, laser-microdissection and in-situ-hybridization we found that the epidermis was the major source of immunomodulatory mediators. This epidermal gene induction was significantly stronger in resistant mice especially for several genes known to promote Th1 differentiation (IL-12, IL-1β, osteopontin, IL-4) and for IL-6. Expression of these cytokines was temporally restricted to the crucial time of Th1/2 differentiation. Moreover, we revealed a stronger epidermal up-regulation of IL-6 in the epidermis of resistant mice. Accordingly, early local neutralization of IL-4 in resistant mice resulted in a Th2 switch and mice with a selective IL-6 deficiency in non-hematopoietic cells showed a Th2 switch and dramatic deterioration of disease. Thus, our data indicate for the first time that epidermal cytokine expression is a decisive factor in the generation of protective Th1 immunity and contributes to the outcome of infection with this important human pathogen

Pathogen- and Host-Directed Antileishmanial Effects Mediated by Polyhexanide (PHMB)

BACKGROUND:Cutaneous leishmaniasis (CL) is a neglected tropical disease caused by protozoan parasites of the genus Leishmania. CL causes enormous suffering in many countries worldwide. There is no licensed vaccine against CL, and the chemotherapy options show limited efficacy and high toxicity. Localization of the parasites inside host cells is a barrier to most standard chemo- and immune-based interventions. Hence, novel drugs, which are safe, effective and readily accessible to third-world countries and/or drug delivery technologies for effective CL treatments are desperately needed. METHODOLOGY/PRINCIPAL FINDINGS:Here we evaluated the antileishmanial properties and delivery potential of polyhexamethylene biguanide (PHMB; polyhexanide), a widely used antimicrobial and wound antiseptic, in the Leishmania model. PHMB showed an inherent antileishmanial activity at submicromolar concentrations. Our data revealed that PHMB kills Leishmania major (L. major) via a dual mechanism involving disruption of membrane integrity and selective chromosome condensation and damage. PHMB's DNA binding and host cell entry properties were further exploited to improve the delivery and immunomodulatory activities of unmethylated cytosine-phosphate-guanine oligodeoxynucleotides (CpG ODN). PHMB spontaneously bound CpG ODN, forming stable nanopolyplexes that enhanced uptake of CpG ODN, potentiated antimicrobial killing and reduced host cell toxicity of PHMB. CONCLUSIONS:Given its low cost and long history of safe topical use, PHMB holds promise as a drug for CL therapy and delivery vehicle for nucleic acid immunomodulators

Photodynamic therapy for multi-resistant cutaneous Langerhans cell histiocytosis

Langerhans cell histiocytosis is a rare group of proliferative disorders. Beside cutaneous involvement, other internal organs can be affected. The treatment of cutaneous lesions is difficult and relies on topical corticosteroids, carmustine, nitrogen mustard, and photochemotherapy. Systemic steroids and vinblastine are used for recalcitrant skin lesions. However, some cases fail to respond. An 18-month old boy presented a CD1a+, S100a+ Langerhans cell histocytosis with cutaneous and severe scalp involvement. Topical corticosteroids and nitrogen mustard failed to improve the skin lesions. Systemic corticosteroids and vinblastine improved the truncal involvement but had no effect on the scalp lesions. Methylaminolevulinate (MAL) based photodynamic therapy (PDT) resulted in a significant regression of the scalp lesions. Control histology revealed an almost complete clearance of the tumor infiltrate. Clinical follow-up after six months showed no recurrence

The IFN-γ-Inducible GTPase, Irga6, Protects Mice against Toxoplasma gondii but Not against Plasmodium berghei and Some Other Intracellular Pathogens

Clearance of infection with intracellular pathogens in mice involves interferon-regulated GTPases of the IRG protein family. Experiments with mice genetically deficient in members of this family such as Irgm1(LRG-47), Irgm3(IGTP), and Irgd(IRG-47) has revealed a critical role in microbial clearance, especially for Toxoplasma gondii. The in vivo role of another member of this family, Irga6 (IIGP, IIGP1) has been studied in less detail. We investigated the susceptibility of two independently generated mouse strains deficient in Irga6 to in vivo infection with T. gondii, Mycobacterium tuberculosis, Leishmania mexicana, L. major, Listeria monocytogenes, Anaplasma phagocytophilum and Plasmodium berghei. Compared with wild-type mice, mice deficient in Irga6 showed increased susceptibility to oral and intraperitoneal infection with T. gondii but not to infection with the other organisms. Surprisingly, infection of Irga6-deficient mice with the related apicomplexan parasite, P. berghei, did not result in increased replication in the liver stage and no Irga6 (or any other IRG protein) was detected at the parasitophorous vacuole membrane in IFN-γ-induced wild-type cells infected with P. berghei in vitro. Susceptibility to infection with T. gondii was associated with increased mortality and reduced time to death, increased numbers of inflammatory foci in the brains and elevated parasite loads in brains of infected Irga6-deficient mice. In vitro, Irga6-deficient macrophages and fibroblasts stimulated with IFN-γ were defective in controlling parasite replication. Taken together, our results implicate Irga6 in the control of infection with T. gondii and further highlight the importance of the IRG system for resistance to this pathogen

Peranan Modal Dari Keluarga Pada 3 UKM di Bandung

Usaha kecil dan menengah menjadi bagian terbesar dari dunia usaha di Indonesia ,  yang telah  terbukti mampu bertahan selama masa krisis, tahun 1997-1998 dan tahun 2008. Usaha skala kecil dan menengah ini (UKM) memiliki peran penting terhadap perkembangan ekonomi, menyerap jumlah tenaga kerja yang banyak memberikan kontribusi terhadap GDP. Ada suatu fakta yang belum cukup  dipahami , bahwa berdirinya usaha skala kecil dan menengah umumnya dimulai dengan melibatkan anggota keluarga atau tidak langsung  mendapat dukungan dan bantuan keluarga . Penelitian ini difokuskan untuk menemukan   peran  modal  yang diberikan oleh keluarga pada saat pendirian usaha dan dalam perkembangannya,  baik modal yang  berupa uang dan finansial (harta tetap atau harta lancar), maupun yang bersifat nilai-nilai (family capital), dan emosi-waktu-perhatian (psychological capital) , dsb . Penelitian dilaksanakan dengan metode kualitatif yakni Multi Case Studies. Tiga UKM  di Bandung  dipilih sebagai obyek penelitian.  Teknik wawancara dan observasi digunakan saling melengkapi   . Hasil transkrip wawancara dikategorisasi (coding) sesuai tema penelitian. Teori modal Bourdieu (1996) digunakan untuk menganalisis dan untuk mendapatkan pemaknaan dari temuan. Temuan menunjukkan bahwa modal ekonomi bukanlah satu-satunya faktor yang berperan dalam memulai dan mengembangkan usaha dalam 3 kasus ini.  Ada  modal yang lain yaitu modal budaya dan modal sosial yang punya peran penting  terhadap awal dan  kelangsungan usaha. Modal simbolik dapat  dibangun dari   modal budaya  sejalan dengan berjalannya usaha dalam  jangka waktu yang cukup   lama. Kata-kata Kunci : modal ekonomi, modal budaya, modal sosial, modal simboli

Differential impact of LPG-and PG-deficient Leishmania major mutants on the immune response of human dendritic cells

<div><p>Background</p><p><i>Leishmania major</i> infection induces robust interleukin-12 (IL12) production in human dendritic cells (hDC), ultimately resulting in Th1-mediated immunity and clinical resolution. The surface of <i>Leishmania</i> parasites is covered in a dense glycocalyx consisting of primarily lipophosphoglycan (LPG) and other phosphoglycan-containing molecules (PGs), making these glycoconjugates the likely pathogen-associated molecular patterns (PAMPS) responsible for IL12 induction.</p><p>Methodology/Principal Findings</p><p>Here we explored the role of parasite glycoconjugates on the hDC IL12 response by generating <i>L</i>. <i>major</i> Friedlin V1 mutants defective in LPG alone, (FV1 <i>lpg1-</i>), or generally deficient for all PGs, (FV1 <i>lpg2-</i>). Infection with metacyclic, infective stage, <i>L</i>. <i>major</i> or purified LPG induced high levels of <i>IL12B</i> subunit gene transcripts in hDCs, which was abrogated with FV1 <i>lpg1-</i> infections. In contrast, hDC infections with FV1 <i>lpg2-</i> displayed increased <i>IL12B</i> expression, suggesting other PG-related/<i>LPG2</i> dependent molecules may act to dampen the immune response. Global transcriptional profiling comparing WT, FV1 <i>lpg1-</i>, FV1 <i>lpg2-</i> infections revealed that FV1 <i>lpg1-</i> mutants entered hDCs in a silent fashion as indicated by repression of gene expression. Transcription factor binding site analysis suggests that LPG recognition by hDCs induces IL-12 in a signaling cascade resulting in Nuclear Factor κ B (NFκB) and Interferon Regulatory Factor (IRF) mediated transcription.</p><p>Conclusions/Significance</p><p>These data suggest that <i>L</i>. <i>major</i> LPG is a major PAMP recognized by hDC to induce IL12-mediated protective immunity and that there is a complex interplay between PG-baring <i>Leishmania</i> surface glycoconjugates that result in modulation of host cellular IL12.</p></div

HisAK70: Progress towards a vaccine against different forms of leishmaniosis

Background: Leishmania major and Leishmania infantum are among the main species that are responsible for cutaneous leishmaniosis (CL) and visceral leishmaniosis (VL), respectively. The leishmanioses represent the second-largest parasitic killer in the world after malaria. Recently, we succeeded in generating a plasmid DNA (pCMV-HISA70m2A) and demonstrated that immunized mice were protected against L. major challenge. The efficacy of the DNA-vaccine was further enhanced by the inclusion of KMP-11 antigen into the antibiotic-free plasmid pVAX1-asd. Methods: Here, we describe the use of a HisAK70 DNA-vaccine encoding seven Leishmania genes (H2A, H2B, H3, H4, A2, KMP11 and HSP70) for vaccination of mice to assess the induction of a resistant phenotype against VL and CL. Results: HisAK70 was successful in vaccinated mice, resulting in a high amount of efficient sterile hepatic granulomas associated with a hepatic parasite burden fully resolved in the VL model; and resulting in 100 % inhibition of parasite visceralization in the CL model. Conclusions: The results suggest that immunization with the HisAK70 DNA-vaccine may provide a rapid, suitable, and efficient vaccination strategy to confer cross-protective immunity against VL and CL.This work was partially supported by grants from the Spanish Ministry of Economy and Competitiveness (AGL2010-17394 and AGL2013-44100R) and PLATESA (P2013/ABI-2906) from the Comunidad de Madrid (Spain).Peer Reviewe