Intracellular Targeting Specificity of Novel Phthalocyanines Assessed in a Host-Parasite Model for Developing Potential Photodynamic Medicine

Photodynamic therapy, unlikely to elicit drug-resistance, deserves attention as a strategy to counter this outstanding problem common to the chemotherapy of all diseases. Previously, we have broadened the applicability of this modality to photodynamic vaccination by exploiting the unusual properties of the trypanosomatid protozoa, Leishmania, i.e., their innate ability of homing to the phagolysosomes of the antigen-presenting cells and their selective photolysis therein, using transgenic mutants endogenously inducible for porphyrin accumulation. Here, we extended the utility of this host-parasite model for in vitro photodynamic therapy and vaccination by exploring exogenously supplied photosensitizers. Seventeen novel phthalocyanines (Pcs) were screened in vitro for their photolytic activity against cultured Leishmania. Pcs rendered cationic and soluble (csPcs) for cellular uptake were phototoxic to both parasite and host cells, i.e., macrophages and dendritic cells. The csPcs that targeted to mitochondria were more photolytic than those restricted to the endocytic compartments. Treatment of infected cells with endocytic csPcs resulted in their accumulation in Leishmania-containing phagolysosomes, indicative of reaching their target for photodynamic therapy, although their parasite versus host specificity is limited to a narrow range of csPc concentrations. In contrast, Leishmania pre-loaded with csPc were selectively photolyzed intracellularly, leaving host cells viable. Pre-illumination of such csPc-loaded Leishmania did not hinder their infectivity, but ensured their intracellular lysis. Ovalbumin (OVA) so delivered by photo-inactivated OVA transfectants to mouse macrophages and dendritic cells were co-presented with MHC Class I molecules by these antigen presenting cells to activate OVA epitope-specific CD8+T cells. The in vitro evidence presented here demonstrates for the first time not only the potential of endocytic csPcs for effective photodynamic therapy against Leishmania but also their utility in photo-inactivation of Leishmania to produce a safe carrier to express and deliver a defined antigen with enhanced cell-mediated immunity

Reducing the environmental impact of surgery on a global scale: systematic review and co-prioritization with healthcare workers in 132 countries

Abstract Background Healthcare cannot achieve net-zero carbon without addressing operating theatres. The aim of this study was to prioritize feasible interventions to reduce the environmental impact of operating theatres. Methods This study adopted a four-phase Delphi consensus co-prioritization methodology. In phase 1, a systematic review of published interventions and global consultation of perioperative healthcare professionals were used to longlist interventions. In phase 2, iterative thematic analysis consolidated comparable interventions into a shortlist. In phase 3, the shortlist was co-prioritized based on patient and clinician views on acceptability, feasibility, and safety. In phase 4, ranked lists of interventions were presented by their relevance to high-income countries and low–middle-income countries. Results In phase 1, 43 interventions were identified, which had low uptake in practice according to 3042 professionals globally. In phase 2, a shortlist of 15 intervention domains was generated. In phase 3, interventions were deemed acceptable for more than 90 per cent of patients except for reducing general anaesthesia (84 per cent) and re-sterilization of ‘single-use’ consumables (86 per cent). In phase 4, the top three shortlisted interventions for high-income countries were: introducing recycling; reducing use of anaesthetic gases; and appropriate clinical waste processing. In phase 4, the top three shortlisted interventions for low–middle-income countries were: introducing reusable surgical devices; reducing use of consumables; and reducing the use of general anaesthesia. Conclusion This is a step toward environmentally sustainable operating environments with actionable interventions applicable to both high– and low–middle–income countries

Photodynamic Sensitization of Leishmania amazonensis in both Extracellular and Intracellular Stages with Aluminum Phthalocyanine Chloride for Photolysis In Vitro

Leishmania amazonensis, a causative agent of cutaneous leishmaniasis, is susceptible in vitro to light-mediated cytolysis in the presence of or after pretreatment with the photosensitizer aluminum phthalocyanine chloride. Cytolysis of both promastigotes and axenic amastigotes required less photosensitizer (e.g., one μg · ml(−1)) and a lower light dose (e.g., 1.5 J · cm(−2)) than did the mammalian cells examined for comparison. Exposure of Leishmania cells to the photosensitizer alone had little effect on their viability, as judged from their motility, growth, and/or retention of green fluorescent proteins genetically engineered for episomal expression. Fluorimetric assays for cell-associated and released green fluorescence proteins proved to be even more sensitive for the evaluation of cell viability than microscopy for the evaluation of motility and/or integrity. Axenic amastigotes pretreated with the photosensitizer infected macrophages of the J774 line but were lysed intracellularly when the infected cells were exposed to light. Addition of the photosensitizer to the already infected cells produced no effect on their intracellular parasites. However, light irradiation lysed these macrophages and also those infected with parasites preincubated with the photosensitizer at a concentration of 5 μg · ml(−1) or higher. Photosensitized Leishmania cells are highly susceptible to cytolysis, apparently due to the generation of reactive oxidative species on light illumination, suggestive of inefficiency of their antioxidant mechanisms. Efficient delivery of photosensitizers to intracellular Leishmania is expected to increase their therapeutic potentials against leishmaniasis

New "light" for one-world approach toward safe and effective control of animal diseases and insect vectors from leishmaniac perspectives

Camila Indiani de Oliveira. FIOCRUZ. Centro de Pesquisas Gonçalo Moniz. Salvador, BA, BrazilRamesh B. Batchu, PhD, Director, Division of Surgical Oncology & Developmental, Therapeutics, Associate Professor, The Michael and Marian Ilitch Department of Surgery, Wayne State University, 4646 John R Road, Detroit, MI 48201, USA. [email protected] Hui-Wen (Winni) Chen, DVM, PhD (陳慧文), Assistant Professor, Department of Veterinary Medicine, National Taiwan University, 1 Sec. 4 Roosevelt Rd., Taipei, 10617 Taiwan. [email protected] Larry Ming C. Chow, ScD (周銘祥), Associate Head and Professor, Department of Applied Biology and Chemical Technology, Hong Kong Polytechnic University, Hong Kong. [email protected] Robert Elliott, MD/Jonathan F. Head, PhD, Elliott Mastology Center, 541 Shadows Lane, Baton Rouge, LA 70806, USA. [email protected] Chia-Kwung Fan PhD (范家堃), Professor and Chairman, Department of Molecular Parasitology and Tropical Diseases, School of Medicine and Center for International Tropical Medicine, College of Medicine, 250 Wu-Xing Street, Taipei Medical University, Taipei, Taiwan. [email protected] Chen-Hsiung Hung, PhD (洪政雄), Investigator and Vice Director, Institute of Chemistry, Academia Sinica, 128, Sec. 2, Academia Road,, Nankang, Taipei, Taiwan 11529. [email protected] Dar-Der Ji, PhD (嵇達德), Associate Professor, Department of Tropical Medicine/International Health Programs, National Yang-Ming University, Beitou, Taipei 112, Taiwan. [email protected] Zhao-Rong Lun, PhD (伦照荣), Professor & Director, Center for Parasitic Organisms, School of Life Sciences, Sun Yat-Sen University, Guangzhou 510275, P.R. China. [email protected] Laura Manna, PhD, DVM, School of Veterinary Medicine and Animal Productions, University of Naples, Federico II, Naples, Italy. [email protected] Yoshitsugu Matsumoto, DVM, PhD, Professor, Laboratory of Molecular Immunology, School of Agricultural and Life Sciences, The University of Tokyo, Yayoi 1-1-1, Tokyo, Japan 113-8657 [email protected] Dennis KP Ng, DPhil (吳基培), Professor, Department of Chemistry, Chinese University of Hong Kong, Sha Tin, N. T., Hong Kong. [email protected] Camila I. de Oliveira PhD/Sayonara Melo, Centro de Pesquisas Gonçalo Moniz. FIOCRUZ, R. Waldemar, Falcão, 121 Candeal, Salvador- BA -40296-710, Brazil. [email protected] Yusuf Ozbel, MD, Professor, Ege University Faculty of Medicine, Department of Parasitology, 35100 Bornova, Izmir,TURKEY. [email protected] Ahmet Özbilgin. PhD, Professor, Celal Bayar Üniversitesi, Tip Fakültesi Parazitoloji A.D, Dekanlik Binasi Uncubozköy, 45030, Manisa, Türkiye. [email protected] Joseph Reynolds, PhD, Department of Microbiology/Immunology, Chicago Medical School/RFUMS, N Chicago, IL 60064, USA. [email protected] Chizu Sanjoba, PhD, Laboratory of Molecular Immunology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1Yayoi, Bunkyo-ku, Tokyo 113-8657 Japan. [email protected], Shin-Hong Shiao, PhD (蕭信宏), Assistant Professor, Department of Parasitology, Institute of Microbiology, School of Medicine, National Taiwan University, Taipei, Taiwan. [email protected] Nang-Yao Shih, PhD (施能耀), Associate Investigator, National Institute of Cancer Research, National Health Research Institutes, Tainan, Taiwan. [email protected] Chi-Wei Tsai, PhD (蔡志偉), Associate Professor, Department of Entomology, College of Natural Resources and Agriculture, National Taiwan University, 1 Sec. 4 Roosevelt Rd., Taipei, 10617 Taiwan. [email protected] Maria da Graça H. Vicente, PhD, Distinguished Professor, Department of Chemistry, 249 Chemistry and Materials Building, Highland Rd, Louisiana State University, Baton Rouge, LA 70803, USA. [email protected] Petr Volf, PhD, Professor, Department of Parasitology, Faculty of Science, Charles University Vinicna 7, 128 44 Praha 2, Czech Republic. [email protected] Yueh-Lung Wu, PhD (吳岳隆), Assistant Professor, Department of Entomology, College of Natural Resources and Agriculture, National Taiwan University, 1 Sec. 4 Roosevelt Rd., Taipei, 10617 Taiwan. [email protected] Chao-Lan Yu, PhD (郁兆蘭), Professor, Department of Biomedical Sciences, Chang Gung University, No. 259, Wenhua 1st Rd., Guishan Dist., Taoyuan City 33302, Taiwan. [email protected] Xiao-Nong Zhou, PhD (周晓农),Professor and Director, National Institute of Parasitic Diseases, Chinese Center for Disease Control and Prevention, 207 Rui Jin Er Road, Shanghai 200025, P.R. China. [email protected]; [email protected] by Ana Maria Fiscina Sampaio ([email protected]) on 2017-03-17T18:29:15Z No. of bitstreams: 1 Chang Kwang Poo New light for....pdf: 2642368 bytes, checksum: bba693f0a7e83e78a21b1ce134e231a2 (MD5)Approved for entry into archive by Ana Maria Fiscina Sampaio ([email protected]) on 2017-03-17T18:54:53Z (GMT) No. of bitstreams: 1 Chang Kwang Poo New light for....pdf: 2642368 bytes, checksum: bba693f0a7e83e78a21b1ce134e231a2 (MD5)Made available in DSpace on 2017-03-17T18:54:53Z (GMT). No. of bitstreams: 1 Chang Kwang Poo New light for....pdf: 2642368 bytes, checksum: bba693f0a7e83e78a21b1ce134e231a2 (MD5) Previous issue date: 2016NIH/NIAID Grant # AI097830, AI-7712375, AI-68835Chicago Medical School. Department of Microbiology, Immunology / Rosalind Franklin University of Medicine and Science. North Chicago, IL, EUAChicago Medical School. Department of Microbiology, Immunology / Rosalind Franklin University of Medicine and Science. North Chicago, IL, EUALight Group - Múltipla - ver em NotasLight is known to excite photosensitizers (PS) to produce cytotoxic reactive oxygen species (ROS) in the presence of oxygen. This modality is attractive for designing control measures against animal diseases and pests. Many PS have a proven safety record. Also, the ROS cytotoxicity selects no resistant mutants, unlike other drugs and pesticides. Photodynamic therapy (PDT) refers to the use of PS as light activable tumoricides, microbicides and pesticides in medicine and agriculture.Here we describe "photodynamic vaccination" (PDV) that uses PDT-inactivation of parasites, i.e. Leishmania as whole-cell vaccines against leishmaniasis, and as a universal carrier to deliver transgenic add-on vaccines against other infectious and malignant diseases. The efficacy of Leishmania for vaccine delivery makes use of their inherent attributes to parasitize antigen (vaccine)-presenting cells. Inactivation of Leishmania by PDT provides safety for their use. This is accomplished in two different ways: (i) chemical engineering of PS to enhance their uptake, e.g. Si-phthalocyanines; and (ii) transgenic approach to render Leishmania inducible for porphyrinogenesis. Three different schemes of Leishmania-based PDV are presented diagrammatically to depict the cellular events resulting in cell-mediated immunity, as seen experimentally against leishmaniasis and Leishmania-delivered antigen in vitro and in vivo. Safety versus efficacy evaluations are under way for PDT-inactivated Leishmania, including those further processed to facilitate their storage and transport. Leishmania transfected to express cancer and viral vaccine candidates are being prepared accordingly for experimental trials.We have begun to examine PS-mediated photodynamic insecticides (PDI). Mosquito cells take up rose bengal/cyanosine, rendering them light-sensitive to undergo disintegration in vitro, thereby providing a cellular basis for the larvicidal activity seen by the same treatments. Ineffectiveness of phthalocyanines and porphyrins for PDI underscores its requirement for different PS. Differential uptake of PS by insect versus other cells to account for this difference is under study.The ongoing work is patterned after the one-world approach by enlisting the participation of experts in medicinal chemistry, cell/molecular biology, immunology, parasitology, entomology, cancer research, tropical medicine and veterinary medicine. The availability of multidisciplinary expertise is indispensable for implementation of the necessary studies to move the project toward product development

Low dose UV-B induced modification of chromophore conformation and it's interaction with microenvironment in cyanobacterial phycobilisomes

486-490Phycobilisomes (Pbsomes) are the supra macromolecular pigment protein complexes of cyanobacteria. Synechococcus  Pbsomes are comprised of phycocyanins (PC) and allophycocyanins (APC).Pbsomes are major light harvesting antennae and also absorb ultraviolet-B (UV-B) radiation (280-320 nm). Synechococcus Pbsomes, upon exposure to low dose of UV-B (0.28 mW cm-2) for different time intervals showed profound alteration in their steady state absorption, fluorescence excitation and emission characteristics (Sah et. al. Biochem. Mol. Biol.Int., Vol. 44, No.2, 245-247). In the present study, we  investigated the effect of low dose of UV -B on isolated Pbsome of Synechococus. <span style="mso-bidi-font-style: italic">Our re<span style="font-size:14.0pt;font-family:HiddenHorzOCR; mso-bidi-font-family:HiddenHorzOCR">sults demonstrate the following alterations. Absorbance at 623 nm initially showed a sharp decrease with increasing exposure time to UV-B radiation. The changes in the visible to near ultraviolet absorption and excitation ratio indicated a change in chromophore conformation, upon prolonged exposure of Pbsomes to UV-B radiation. This modification of chromophore conformation appeared to be associated with the loss of energy transfer from PC to APC. Circular dichroism spectra in the amide region showed a significant loss of the α helical content of Pbsomes when exposed for longer duration to UV-B. CD spectra in the visible region revealed a marked decrease in the rotational strength at 620 nm. Close monitoring of CD signals emanating in the 500 to 700 nm range further revealed that the decrease in the rotational strength was closely associated with an initial red shift in the positive CD band of Pbsomes when exposed to UV -B for short duoration. However, the peak became constant over prolonged exposure to UV-B radiation and accompanied a prominent blue shoulder in the positive CD band which suggests the modification and uncoupling of the various phycocyanobilin (PCB) chromophores of the Synechococcus  Pbsomes. </span

Photodynamic Vaccination of BALB/c Mice for Prophylaxis of Cutaneous Leishmaniasis Caused by Leishmania amazonensis

Submitted by Ana Maria Fiscina Sampaio ([email protected]) on 2018-04-16T19:07:59Z No. of bitstreams: 1 Viana S Photodynamic vaccination of BALBc mice ....pdf: 1972289 bytes, checksum: d8f1288efe88a2a888d4aea1d0c76945 (MD5)Approved for entry into archive by Ana Maria Fiscina Sampaio ([email protected]) on 2018-04-16T19:19:02Z (GMT) No. of bitstreams: 1 Viana S Photodynamic vaccination of BALBc mice ....pdf: 1972289 bytes, checksum: d8f1288efe88a2a888d4aea1d0c76945 (MD5)Made available in DSpace on 2018-04-16T19:19:02Z (GMT). No. of bitstreams: 1 Viana S Photodynamic vaccination of BALBc mice ....pdf: 1972289 bytes, checksum: d8f1288efe88a2a888d4aea1d0c76945 (MD5) Previous issue date: 2018NIH-NIAID grant # AI-68835, AI-7712375, and AI-097830 to KC. Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) – (Programa Visitante Especial) Processo 402312/2012-0 to CdO. SV, FC, and LR were supported by CNPq fellowships and CdO is a sênior investigator for CNPq.Fundação Oswaldo Cruz. Instituto Gonçalo Moniz. Salvador, BA, Brasil.Fundação Oswaldo Cruz. Instituto Gonçalo Moniz. Salvador, BA, Brasil.Fundação Oswaldo Cruz. Instituto Gonçalo Moniz. Salvador, BA, Brasil.Rosalind Franklin University of Medicine and Science. Chicago Medical School. Department of Microbiology/Immunology. North Chicago, IL, USA.The Chinese University of Hong Kong. Department of Chemistry. Hong Kong, Hong Kong.Rosalind Franklin University of Medicine and Science. Chicago Medical School. Department of Microbiology/Immunology. North Chicago, IL, USA.Fundação Oswaldo Cruz. Instituto Gonçalo Moniz. Salvador, BA, Brasil / Instituto Nacional de Ciência e Tecnologia. Instituto de Investigação em Imunologia. São Paulo, SP, Brasil.Background: Photosensitizers (PS), like porphyrins and phthalocyanines (PC) are excitable by light to generate cytotoxic singlet oxygen and other reactive oxygen species in the presence of atmospheric O2. Photodynamic inactivation of Leishmania by this means renders them non-viable, but preserves their effective use as vaccines. Leishmania can be photo-inactivated after PS-sensitization by loading via their endocytic uptake of PC or endogenous induction of transgenic mutants with delta-aminolevulinate (ALA) to accumulate cytosolic uroporphyrin I (URO). Here, PS-sensitization and photo-inactivation of Leishmaniaamazonensis was further examined in vitro and in vivo for vaccination against cutaneous leishmaniasis (CL). Methods and Results:Leishmania amazonensis promastigotes were photodynamically inactivated in vitro by PC-loading followed by exposure to red light (1-2 J/cm2) or ALA-induction of uroporphyrinogenic transfectants to accumulate cytosolic URO followed by longwave UV exposure. When applied individually, both strategies of photodynamic inactivation were found to significantly, albeit incompletely abolish the MTT reduction activities of the promastigotes, their uptake by mouse bone marrow-derived macrophages in vitro and their infectivity to mouse ear dermis in vivo. Inactivation of Leishmania to completion by using a combination of both strategies was thus used for the sake of safety as whole-cell vaccines for immunization of BALB/c mice. Different cutaneous sites were assessed for the efficacy of such photodynamic vaccination in vivo. Each site was inoculated first with in vitro doubly PS-sensitized promastigotes and then spot-illuminated with white light (50 J/cm2) for their photo-inactivation in situ. Only in ear dermis parasites were photo-inactivated beyond detection. Mice were thus immunized once in the ear and challenged 3 weeks later at the tail base with virulent L. amazonensis. Prophylaxis was noted in mice photodynamically vaccinated with doubly photo-inactivated parasites, as indicated by a significant delay in the onset of lesion development and a substantial decrease in the parasite loads. Conclusion: Leishmania doubly PS-sensitized and in situ photo-inactivated as described proved to be safe and effective when used for one-time immunization of ear dermis, as indicated by its significant protection of the inherently very susceptible BALB/c mice against CL

New light for one-world approach toward safe and effective control of animal diseases and insect vectors from leishmaniac perspectives

© 2016 The Author(s). Light is known to excite photosensitizers (PS) to produce cytotoxic reactive oxygen species (ROS) in the presence of oxygen. This modality is attractive for designing control measures against animal diseases and pests. Many PS have a proven safety record. Also, the ROS cytotoxicity selects no resistant mutants, unlike other drugs and pesticides. Photodynamic therapy (PDT) refers to the use of PS as light activable tumoricides, microbicides and pesticides in medicine and agriculture. Here we describe photodynamic vaccination (PDV) that uses PDT-inactivation of parasites, i.e. Leishmania as whole-cell vaccines against leishmaniasis, and as a universal carrier to deliver transgenic add-on vaccines against other infectious and malignant diseases. The efficacy of Leishmania for vaccine delivery makes use of their inherent attributes to parasitize antigen (vaccine)-presenting cells. Inactivation of Leishmania by PDT provides safety for their use. This is accomplished in two different ways: (i) chemical engineering of PS to enhance their uptake, e.g. Si-phthalocyanines; and (ii) transgenic approach to render Leishmania inducible for porphyrinogenesis. Three different schemes of Leishmania-based PDV are presented diagrammatically to depict the cellular events resulting in cell-mediated immunity, as seen experimentally against leishmaniasis and Leishmania-delivered antigen in vitro and in vivo. Safety versus efficacy evaluations are under way for PDT-inactivated Leishmania, including those further processed to facilitate their storage and transport. Leishmania transfected to express cancer and viral vaccine candidates are being prepared accordingly for experimental trials. We have begun to examine PS-mediated photodynamic insecticides (PDI). Mosquito cells take up rose bengal/cyanosine, rendering them light-sensitive to undergo disintegration in vitro, thereby providing a cellular basis for the larvicidal activity seen by the same treatments. Ineffectiveness of phthalocyanines and porphyrins for PDI underscores its requirement for different PS. Differential uptake of PS by insect versus other cells to account for this difference is under study. The ongoing work is patterned after the one-world approach by enlisting the participation of experts in medicinal chemistry, cell/molecular biology, immunology, parasitology, entomology, cancer research, tropical medicine and veterinary medicine. The availability of multidisciplinary expertise is indispensable for implementation of the necessary studies to move the project toward product development