Delta-Aminolevulinate-Induced Host-Parasite Porphyric Disparity for Selective Photolysis of Transgenic Leishmania in the Phagolysosomes of Mononuclear Phagocytes: a Potential Novel Platform for Vaccine Delivery

Dutta, Sandeep; Chang, Chau‐Lyan; Kolli, Bala K.; Sassa, Shigeru; Yousef, Malik; Showe, Michael K.; Showe, Louise C.; Chang, Kwang-Poo

doi:10.1128/ec.05202-11

Cited by 9 publications

(18 citation statements)

References 39 publications

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“…S2C and D in the supplemental material). Macrophages of the J774 cell line were cultured at 35°C without 5% (vol/vol) CO 2 atmosphere in RPMI 1640 medium HEPES buffered to pH 7.4 plus 10% (vol/vol) HIFBS (22)(23)(24).…”

Section: Methodsmentioning

confidence: 99%

“…The cell suspensions in 24-well culture plates and monolayers of J774 macrophages in 25-cm Ϫ2 tissue culture (TC) flasks were exposed from the bottom to red light (wavelengths, Ͼ650 nm) via a red filter (product no. 650021; Smith-Victor Co., Bartlett, IL, USA) over a light box for 20 to 30 min at a fluence of 1 to 2 J cm Ϫ2 (22,23). Cell viability assay.…”

Section: Methodsmentioning

confidence: 99%

“…Cell viability assay. An MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] reduction assay was done, according to the manufacturer's protocol (Molecular Probe kits; Sigma) and as previously described (22,23). A freshly prepared MTT stock solution (5 mg ml Ϫ1 ) was added to promastigote and axenic amastigote cell suspensions at 0.5 ϫ 10 8 to 1 ϫ 10 8 ml Ϫ1 .…”

Section: Methodsmentioning

confidence: 99%

“…Nikon Eclipse 80i and TE2000-S microscopes with CCD cameras were used with the Metamorphosis (version 6.1) or NIS Elements version AR 4.20.01 software for image capture and analysis (18,(22)(23)(24). The cells were examined under living conditions by placing concentrated cell suspensions in 3-l aliquots each on a glass slide covered with an 18-mm 2 glass coverslip.…”

Section: Methodsmentioning

confidence: 99%

“…Photodynamic vaccination of hamsters based on this principle produced cell-mediated immunity that is adaptively transferable to naive animals against visceral leishmaniasis (21). The porphyrinogenic mutants have the potential to serve as a universal platform for vaccine delivery, as shown by the findings that ALA-induced porphyrinogenesis, followed by photoinactivation of Leishmania in situ, reversed the immunosuppression of macrophages caused by the infection (22). For endosomal accumulation of PS in Leishmania, silicon(IV)-phthalocyanines (PC) attached axially with bulky ligands (PC14 and PC15) were identified as effective by screening a panel of ϳ17 novel PCs (23).…”

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confidence: 99%

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Aminophthalocyanine-Mediated Photodynamic Inactivation of Leishmania tropica

Al‐Qahtani

Alkahtani

Kolli

et al. 2016

Antimicrob Agents Chemother

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dPhotodynamic inactivation of Leishmania spp. requires the cellular uptake of photosensitizers, e.g., endocytosis of silicon(IV)-phthalocyanines (PC) axially substituted with bulky ligands. We report here that when substituted with amino-containing ligands, the PCs (PC1 and PC2) were endocytosed and displayed improved potency against Leishmania tropica promastigotes and axenic amastigotes in vitro. The uptake of these PCs by both Leishmania stages followed saturation kinetics, as expected. Sensitive assays were developed for assessing the photodynamic inactivation of Leishmania spp. by rendering them fluorescent in two ways: transfecting promastigotes to express green fluorescent protein (GFP) and loading them with carboxyfluorescein succinimidyl ester (CFSE). PC-sensitized Leishmania tropica strains were seen microscopically to lose their motility, structural integrity, and GFP/CFSE fluorescence after exposure to red light (wavelength, ϳ650 nm) at a fluence of 1 to 2 J cm ؊2 . Quantitative fluorescence assays based on the loss of GFP/CFSE from live Leishmania tropica showed that PC1 and PC2 dose dependently sensitized both stages for photoinactivation, consistent with the results of a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) cell viability assay. Leishmania tropica strains are >100 times more sensitive than their host cells or macrophages to PC1-and PC2-mediated photoinactivation, judging from the estimated 50% effective concentrations (EC 50 s) of these cells. Axial substitution of the PC with amino groups instead of other ligands appears to increase its leishmanial photolytic activity by up to 40-fold. PC1 and PC2 are thus potentially useful for photodynamic therapy of leishmaniasis and for oxidative photoinactivation of Leishmania spp. for use as vaccines or vaccine carriers.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

See 3 more Smart Citations

Aminophthalocyanine-Mediated Photodynamic Inactivation of Leishmania tropica

Al‐Qahtani

Alkahtani

Kolli

et al. 2016

Antimicrob Agents Chemother

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Photodynamic Antimicrobial Chemotherapy

Phoenix

Dennison

Harris

2014

Novel Antimicrobial Agents and Strategies

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Photodynamic inactivation of Leishmania braziliensis doubly sensitized with uroporphyrin and diamino-phthalocyanine activates effector functions of macrophages in vitro

Sharma

Viana

et al. 2020

Sci Rep

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Photodynamic inactivation of Leishmania has been shown to render them non-viable, but retain their immunological activities. Installation of dual photodynamic mechanisms ensures complete inactivation of species in the Leishmania subgenus, raising the prospect of their safe and effective application as whole-cell vaccines against leishmaniasis. Here, we report the successful extension of this approach to L. braziliensis in the Viannia subgenus, viz. genetic engineering of promastigotes for cytosolic accumulation of UV-sensitive uroporphyrin (URO) and their loading with red light excitable phthalocyanines (PC) that was cationized by chemical engineering. The transgenic strategy used previously produced L. braziliensis transfectants, which gave the same phenotype of aminolevulinate (ALA)-inducible uroporphyria as found in Leishmania subgenus, indicative of pre-subgenus evolutionary origin for similar genetic deficiencies in porphyrin/heme biosynthesis. In the present study, 12 independent clones were obtained and were invariably ALA-responsive, albeit to different extent for uroporphyrinogenesis and UV-inactivation. In a separate study, L. braziliensis was also found, like other Leishmania spp., to take up diamino-PC (PC2) for red light inactivation. In vitro interactions of a highly uroporphyrinogenic clone with primary macrophages were examined with the intervention of URO/PC2-medated double-photodynamic inactivation to ascertain its complete loss of viability. Doubly sensitized L. braziliensis transfectants were photo-inactivated before (Strategy #1) or after (Strategy #2) loading of macrophages. In both cases, macrophages were found to take up L. braziliensis and degrade them rapidly in contrast to live Leishmania infection. The effector functions of macrophages became upregulated following their loading with L. braziliensis photodynamically inactivated by both strategies, including CD86 expression, and IL6 and NO production. This was in contrast to the immunosuppressive infection of macrophages with live parasites, marked by IL10 production. The results provide evidence that photodynamically inactivated L. braziliensis are susceptible to the degradative pathway of macrophages with upregulation of immunity relevant cytokine and co-stimulatory markers. The relative merits of the two loading strategies with reference to previous experimental vaccination were discussed in light of the present findings with L. braziliensis.

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Delta-Aminolevulinate-Induced Host-Parasite Porphyric Disparity for Selective Photolysis of Transgenic Leishmania in the Phagolysosomes of Mononuclear Phagocytes: a Potential Novel Platform for Vaccine Delivery

Cited by 9 publications

References 39 publications

Aminophthalocyanine-Mediated Photodynamic Inactivation of Leishmania tropica

Aminophthalocyanine-Mediated Photodynamic Inactivation of Leishmania tropica

Photodynamic Antimicrobial Chemotherapy

Photodynamic inactivation of Leishmania braziliensis doubly sensitized with uroporphyrin and diamino-phthalocyanine activates effector functions of macrophages in vitro

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