Photochemical Strategies toward Precision Targeting against Multidrug-Resistant Bacterial Infections.

Infectious diseases pose a serious threat and a substantial economic burden on global human and public health security, especially with the frequent emergence of multidrug-resistant (MDR) bacteria in clinical settings. In response to this urgent need, various photobased anti-infectious therapies have been reported lately. This Review explores and discusses several photochemical targeted antibacterial therapeutic strategies for addressing bacterial infections regardless of their antibiotic susceptibility. In contrast to conventional photobased therapies, these approaches facilitate precise targeting of pathogenic bacteria and/or infectious microenvironments, effectively minimizing toxicity to mammalian cells and surrounding healthy tissues. The highlighted therapies include photodynamic therapy, photocatalytic therapy, photothermal therapy, endogenous pigments-based photobleaching therapy, and polyphenols-based photo-oxidation therapy. This comprehensive exploration aims to offer updated information to facilitate the development of effective, convenient, safe, and alternative strategies to counter the growing threat of MDR bacteria in the future.

Oregano Oil and Harmless Blue Light to Synergistically Inactivate Multidrug-Resistant Pseudomonas aeruginosa.

Blue light (BL) at 405 nm and oregano essential oil (OEO) have shown bactericidal activity by its own. Here, we demonstrated that the two synergistically killed multidrug-resistant (MDR) Pseudomonas aeruginosa (Pa). Pa ATCC19660 and HS0065 planktonic cells and mature biofilms were reduced by more than 7 log10 after treatment by BL combined with OEO, in sharp contrast to no significant bacterial reduction with the monotreatment. The duo also sufficiently eliminated acute or biofilm-associated infection of open burn wounds in murine without incurring any harmful events in the skin. The synergic bactericide was attributed mainly to the ability of OEO to magnify cytotoxic reactive oxygen species (ROS) production initiated by BL that excited endogenous tetrapyrrole macrocycles in bacteria while completely sparing the surrounding tissues from the phototoxic action. OEO ingredient analysis in combination with microbial assays identified carvacrol and its isomer thymol to be the major phytochemicals that cooperated with BL executing synergic killing. The finding argues persuasively for valuable references of carvacrol and thymol in assessing and standardizing the bactericidal potential of various OEO products.

Blue light potentiates safety and bactericidal activity of p-Toluquinone.

Fewer antibiotics are available for effective management of bacterial infections to date owing to increasing multiple-drug resistance (MDR). Here, we expand our early success in combination of 405 nm blue light irradiation with phenolic compounds to sufficiently kill blue light-refractory MDR Escherichia coli (E. coli). p-Toluquinone (p-TQ) alongside blue light inactivated 7.3 log10E. coli within 6 min, whereas either alone was totally ineffective. A similar killing efficacy was attained with four other pathogens commonly seen in hospital-acquired infections and Enterococcus faecalis (Ef) that don't produce porphyrins-like molecules. The combinatory therapy prevented recurrence of E. coli infection in skin scratch wounds of murine. The bactericidal activity was ascribed to reactive oxygen species (ROS) generation triggered by blue light-mediated excitation of p-TQ, which is less likely to induce resistance because of multi-targeted and non-specific nature of ROS. Remarkably, toxic p-TQ became harmless to mammalian cells after brief exposure to blue light while retaining its bactericidal activity. The opposite effect of blue light on p-TQ activity unravels a novel, simple strategy to detoxify p-TQ and its combination with blue light as a safe and efficacious bactericidal modality for managing MDR bacterial infections.

Corrigendum: Bactericidal Property of Oregano Oil Against Multidrug-Resistant Clinical Isolates.

[This corrects the article DOI: 10.3389/fmicb.2018.02329.].

Thermosensitive and Conductive Hybrid Polymer for Real-Time Monitoring of Spheroid Growth and Drug Responses.

Three-dimensional (3D) cell culture based on polymer scaffold provides a promising tool to mimic a physiological microenvironment for drug testing; however, the next-generation cell activity monitoring technology for 3D cell culture is still challenging. Conventionally, drug efficacy evaluation and cell growth heavily rely on cell staining assays, using optical devices or flow cytometry. Here, we report a dual-function polymer scaffold (DFPS) composed of thermosensitive, silver flake- and gold nanoparticle-decorated polymers, enabling conductance change upon cell proliferation or death for in situ cell activity monitoring and drug screening. The cell activity can be quantitatively monitored via measuring the conductance change induced by polymeric network swelling or shrinkage. This novel dual-function system (1) provides a 3D microenvironment to enable the formation and growth of tumor spheroid in vitro and streamlines the harvesting of tumor spheroids through the thermosensitive scaffold and (2) offers a simple and direct quantitative method to monitor 3D cell culture in situ for drug responses. As a proof of concept, we demonstrated that a breast cancer stem cell line MDA-MB-436 was able to form cell spheroids in the scaffold, and the conductance change of the sensor exhibited a linear relationship with cell concentration. To examine its potential in drug screening, cancer spheroids in the cell sensor were treated with paclitaxel (PTX) and docetaxel (DTX), and predicted quantitative evaluation of the cytotoxic effect of drugs was established. Our results indicated that this cell sensing system may hold promising potential in expanding into an array device for high-throughput drug screening.

830 nm photobiomodulation therapy promotes engraftment of human umbilical cord blood-derived hematopoietic stem cells.

Human umbilical cord blood (hUCB)-derived hematopoietic stem cells (HSCs) are an important source for HSCs in allogeneic HSC transplantation, but a limited number and a low efficacy of engraftment greatly restrict their clinical use. Here, we report the ability of photobiomodulation therapy (PBMT) to significantly enhance the engraftment efficacy of hUCB HSCs and progenitor cells (HSPCs). hUCB CD34+ cells were illuminated at a fluence of 2 J/cm2 with a near-infrared light (830 nm) transmitted by an array of light-emitting diodes (LED) prior to infusion of NOD/SCID-IL2Rγ-/- mice. The pre-treatment resulted in a threefold higher of the mean percentage of human CD45+ cells in the periphery of the mice compared to sham-treated CD34+ cells. The enhanced engraftment may result from a PBMT-mediated increase of intracellular reactive oxygen species (ROS) levels and Src protein phosphorylation in CD34+ cells. The two events were causally related as suggested by the finding that elevation of ROS by hydrogen peroxide increased Src phosphorylation, while ROS reduction by N-acetyl cysteine partially reversed the phosphorylation. The investigation demonstrates that PBMT can promote engraftment of hUCB HPSCs, at least in part, via ROS-mediated Src signaling pathway. PBMT can be potentially a safe, convenient, and cost-effective modality to improve hematological reconstitution in patients.

Engineering the Protein Corona Structure on Gold Nanoclusters Enables Red-Shifted Emissions in the Second Near-infrared Window for Gastrointestinal Imaging.

The application of NIR-II emitters for gastrointestinal (GI) tract imaging remains challenging due to fluorescence quenching in the digestive microenvironment. Herein, we report that red-shifting of the fluorescence emission of Au nanoclusters (AuNCs) into NIR-II region with improved quantum yields (QY) could be achieved by engineering a protein corona structure consisting of a ribonuclease-A (RNase-A) on the particle surfaces. RNase-A-encapsulated AuNCs (RNase-A@AuNCs) displayed emissions at 1050 nm with a 1.9 % QY. Compared to rare earth and silver-based NIR-II emitters, RNase-A@AuNCs had excellent biocompatibility, showing >50-fold higher sensitivity in GI tract, and migrated homogenously during gastrointestinal peristalsis to allow visualization of the detailed structures of the GI tract. RNase-A@AuNCs could successfully examine intestinal tumor mice from healthy mice, indicating a potential utility for early diagnosis of intestinal tumors.

Pulmonary surfactant-biomimetic nanoparticles potentiate heterosubtypic influenza immunity.

Current influenza vaccines only confer protection against homologous viruses. We synthesized pulmonary surfactant (PS)-biomimetic liposomes encapsulating 2',3'-cyclic guanosine monophosphate-adenosine monophosphate (cGAMP), an agonist of the interferon gene inducer STING (stimulator of interferon genes). The adjuvant (PS-GAMP) vigorously augmented influenza vaccine-induced humoral and CD8+ T cell immune responses in mice by simulating the early phase of viral infection without concomitant excess inflammation. Two days after intranasal immunization with PS-GAMP-adjuvanted H1N1 vaccine, strong cross-protection was elicited against distant H1N1 and heterosubtypic H3N2, H5N1, and H7N9 viruses for at least 6 months while maintaining lung-resident memory CD8+ T cells. Adjuvanticity was then validated in ferrets. When alveolar epithelial cells (AECs) lacked Sting or gap junctions were blocked, PS-GAMP-mediated adjuvanticity was substantially abrogated in vivo. Thus, AECs play a pivotal role in configuring heterosubtypic immunity.

Uncoupling of CD71 shedding with mitochondrial clearance in reticulocytes in a subset of myelodysplastic syndromes.

Reticulocytes shed CD71 from the cell membrane and eliminate mitochondria during terminal maturation, but it is unknown whether these two events are coordinated. We demonstrate that timely removal of CD71 is coupled with mitochondrial clearance, which can be disrupted by null mutation of immediate early response gene X-1 (IEX-1), leading to generation of aberrant CD71-positive and mitochondria-negative (CD71+Mito-) reticulocytes. CD71+Mito- reticulocytes were also present in a subset of patients with myelodysplastic syndromes (MDS) in direct proportion to reduced mitochondrial membrane potential (∆ψm). Mitochondrial abnormality caused by either IEX-1 deficiency or agents that dissipate ∆ψm could trigger premature clearance of mitochondria in reticulocytes. Premature clearance of mitochondria or addition of anti-oxidants lowered intracellular reactive oxygen species (ROS) that in turn hindered CD71 shedding and reticulocyte maturation. In contrast, introduction of ROS accelerated CD71 shedding via release of exosomes that contained a high proportion of Fe3+ over Fe2+, suggesting dual functions of CD71 shedding both in removal of toxic Fe3+ from reticulocytes and in limiting importation of Fe3+ into the cells. These observations emphasize the coordination of mitochondrial and CD71 clearance in erythroid terminal maturation and offer new insights into a role for mitochondrial degeneration in the pathogenesis of some MDS-associated anemia.

Potentials for prolonging shelf-life of platelets by near infrared low-level light.

Platelets are uniquely stored at room temperature, during which they gradually loss their quality owing to deteriorating functions of mitochondria over time. Given the well-documented beneficial effect of near infrared low-level light (LLL) on mitochondrial functions, we explored a potential for LLL to protect mitochondrial function and extend the shelf-life of platelets beyond the current 5 days. We found that exposure of a platelet-containing storage bag to 830 nm light-emitting diode (LED) light at 0.5 J/cm2 prior to storage could significantly retain a pH value and viability of the platelets stored for 8 days with improved quality compared to those stored similarly for 5 days in controls. The LLL inhibited reactive oxygen species (ROS) and lactate production, while sustaining ATP synthesis and mitochondrial membrane potential and morphology in the stored platelets. It also sustained aggregation capacity and in vivo survival of stored platelets, concomitant with no significant activation, as suggested by similar CD62p expression and enhanced agonist-induced aggregation and recovery following infusion in the presence compared to absence of LLL treatment. This simple, additive-free, cost-effective, noninvasive approach can be readily incorporated into the current platelet storage system to potentially improve quality of stored platelets.

Bactericidal Property of Oregano Oil Against Multidrug-Resistant Clinical Isolates.

Development of non-antibiotic alternatives to treat infections caused by multidrug-resistant (MDR) microbes represents one of the top priorities in healthcare and community settings, especially in the care of combat trauma-associated wound infections. Here, we investigate efficacy of oregano oil against pathogenic bacteria including MDR isolates from the combat casualties in vitro and in a mouse burn model. Oregano oil showed a significant anti-bacterial activity against 11 MDR clinical isolates including four Acinetobacter baumannii, three Pseudomonas aeruginosa, and four methicillin-resistant Staphylococcus aureus (MRSA) obtained from combat casualties and two luminescent strains of PA01 and MRSA USA300, with a MIC ranging from 0.08 mg/ml to 0.64 mg/ml. Oregano oil also effectively eradicated biofilms formed by each of the 13 pathogens above at similar MICs. Transmission electron microscopy (TEM) and scanning electron microscopy (SEM) revealed that oregano oil damaged bacterial cells and altered the morphology of their biofilms. While efficiently inactivating bacteria, there was no evidence of resistance development after up to 20 consecutive passages of representative bacterial strains in the presence of sublethal doses of oregano oil. In vivo study using the third-degree burn wounds infected with PA01 or USA300 demonstrated that oregano oil, topically applied 24 h after bacterial inoculation, sufficiently reduced the bacterial load in the wounds by 3 log10 in 1 h, as measured by drastic reduction of bacterial bioluminescence. This bactericidal activity of oregano oil concurred with no significant side effect on the skin histologically or genotoxicity after three topical applications of oregano oil at 10 mg/ml for three consecutive days. The investigation suggests potentials of oregano oil as an alternative to antibiotics for the treatment of wound-associated infections regardless of antibiotic susceptibility.

Physical activation of innate immunity by spiky particles.

Microbial biochemicals have been indicated as the primary stimulators of innate immunity, the first line of the body's defence against infections. However, the influence of topological features on a microbe's surface on immune responses remains largely unknown. Here we demonstrate the ability of TiO2 microparticles decorated with nanospikes (spiky particles) to activate and amplify the immune response in vitro and in vivo. The nanospikes exert mechanical stress on the cells, which results in potassium efflux and inflammasome activation in macrophages and dendritic cells during phagocytosis. The spiky particles augment antigen-specific humoral and cellular immune responses in the presence of monophosphoryl lipid A and elicit protective immunity against tumour growth and influenza viral infection. The study offers insights into how surface physical cues can tune the activation of innate immunity and provides a basis for engineering particles with increased immunogenicity and adjuvanticity.

A clinical review of phototherapy for psoriasis.

Psoriasis is an autoimmune inflammatory skin disease. In the past several decades, phototherapy has been widely used to treat stable psoriatic lesions, including trunk, scalp, arms and legs, and partial nail psoriasis. A variety of light/lasers with different mechanisms of action have been developed for psoriasis including ultraviolet B (UVB), psoralen ultraviolet A (PUVA), pulsed dye laser (PDL), photodynamic therapy (PDT), intense pulsed light (IPL), light-emitting diodes (LED), and so on. Because light/laser each has specific therapeutic and adverse effects, it is important to adequately choose the sources and parameters in management of psoriasis with different pathogenic sites, severities, and duration of the disorder. This review aims at providing most updated clinic information to physicians about how to select light/laser sources and individual therapeutic regimens. To date, UV light is primarily for stable plaque psoriasis and PDL for topical psoriatic lesions with small area, both of which are safe and effective. On the other hand, PUVA has better curative effects than UVB for managing refractory psoriasis plaques, if its side effects can be better controlled. PDL provides optimal outcomes on nail psoriasis compared with other lasers. Although the trails of low-level light/laser therapy (LLLT) are still small, the near infrared (NIR) and visible red light with low energy show promise for treating psoriasis due to its strong penetration and encouraging photobiomodulation. IPL is rarely reported for psoriasis treatment, but PDT-IPL has been found to offer a moderate effect on nail psoriasis. In brief, various phototherapies have been used either in different combinations or as monotherapy. The modality has become a mainstay in the treatment of mild-to-moderate psoriasis without systemic adverse events in today's clinical practice.

Sustained epidermal powder drug delivery via skin microchannels.

Transdermal delivery of hydrophilic drugs is challenging. This study presents a novel sustained epidermal powder delivery technology (sEPD) for safe, efficient, and sustained delivery of hydrophilic drugs across the skin. sEPD is based on coating powder drugs into high-aspect-ratio, micro-coating channels (MCCs) followed by topical application of powder drug-coated array patches onto ablative fractional laser-generated skin MCs to deliver drugs into the skin. We found sEPD could efficiently deliver chemical drugs without excipients and biologics drugs in the presence of sugar excipients into the skin with a duration of ~12h. Interestingly the sEPD significantly improved zidovudine bioavailability by ~100% as compared to oral gavage delivery. sEPD of insulin was found to maintain blood glucose levels in normal range for at least 6h in chemical-induced diabetes mice, while subcutaneous injection failed to maintain blood glucose levels in normal range. sEPD of anti-programmed death-1 antibody showed more potent anti-tumor efficacy than intraperitoneal injection in B16F10 melanoma models. Tiny skin MCs and 'bulk' drug powder inside relatively deep MCCs are crucial to induce the sustained drug release. The improved bioavailability and functionality warrants further development of the novel sEPD for clinical use.

Analgesic Microneedle Patch for Neuropathic Pain Therapy.

Neuropathic pain caused by nerve injury is debilitating and difficult to treat. Current systemic pharmacological therapeutics for neuropathic pain produce limited pain relief and have undesirable side effects, while current local anesthetics tend to nonspecifically block both sensory and motor functions. Calcitonin gene related peptide (CGRP), a neuropeptide released from sensory nerve endings, appears to play a significant role in chronic neuropathic pain. In this study, an analgesic microneedle (AMN) patch was developed using dissolvable microneedles to transdermally deliver selective CGRP antagonist peptide in a painless manner for the treatment of localized neuropathic pain. Local analgesic effects were evaluated in rats by testing behavioral pain sensitivity in response to thermal and mechanical stimuli using neuropathic pain models such as spared-nerve injury and diabetic neuropathy pain, as well as neurogenic inflammatory pain model induced by ultraviolet B (UVB) radiation. Unlike several conventional therapies, the AMN patches produced effective analgesia on neuropathic pain without disturbing the normal nociception and motor function of the rat, resulting from the high specificity of the delivered peptide against CGRP receptors. The AMN patches did not cause skin irritation or systemic side effects. These results demonstrate that dissolvable microneedle patches delivering CGRP antagonist peptide provide an effective, safe, and simple approach to mitigate neuropathic pain with significant advantages over current treatments.

Low-level light treatment ameliorates immune thrombocytopenia.

Immune thrombocytopenia (ITP) is an immune-mediated acquired bleeding disorder characterized by abnormally low platelet counts. We reported here the ability of low-level light treatment (LLLT) to alleviate ITP in mice. The treatment is based on noninvasive whole body illumination 30 min a day for a few consecutive days by near infrared light (830 nm) transmitted by an array of light-emitting diodes (LEDs). LLLT significantly lifted the nadir of platelet counts and restored tail bleeding time when applied to two passive ITP models induced by anti-CD41 antibody. The anti-platelet antibody hindered megakaryocyte differentiation from the progenitors, impaired proplatelet and platelet formation, and induced apoptosis of platelets. These adverse effects of anti-CD41 antibody were all mitigated by LLLT to varying degrees, owing to its ability to enhance mitochondrial biogenesis and activity in megakaryocytes and preserve mitochondrial functions in platelets in the presence of the antibody. The observations argue not only for contribution of mitochondrial stress to the pathology of ITP, but also clinical potentials of LLLT as a safe, simple, and cost-effective modality of ITP.

Noninvasive low-level laser therapy for thrombocytopenia.

Thrombocytopenia is a common hematologic disorder that is managed primarily by platelet transfusions. We report here that noninvasive whole-body illumination with a special near-infrared laser cures acute thrombocytopenia triggered by γ-irradiation within 2 weeks in mice, as opposed to a 5-week recovery time required in controls. The low-level laser (LLL) also greatly accelerated platelet regeneration in the presence of anti-CD41 antibody that binds and depletes platelets, and prevented a severe drop in platelet count caused by a common chemotherapeutic drug. Mechanistically, LLL stimulated mitochondrial biogenesis specifically in megakaryocytes owing to polyploidy of the cells. LLL also protected megakaryocytes from mitochondrial injury and apoptosis under stress. The multifaceted effects of LLL on mitochondria bolstered megakaryocyte maturation; facilitated elongation, branching, and formation of proplatelets; and doubled the number of platelets generated from individual megakaryocytes in mice. LLL-mediated platelet biogenesis depended on megakaryopoiesis and was inversely correlated with platelet counts, which kept platelet biogenesis in check and effectively averted thrombosis even after repeated uses, in sharp contrast to all current agents that stimulate the differentiation of megakaryocyte progenitors from hematopoietic stem cells independently of platelet counts. This safe, drug-free, donor-independent modality represents a paradigm shift in the prophylaxis and treatment of thrombocytopenia.

Natural STING Agonist as an "Ideal" Adjuvant for Cutaneous Vaccination.

A potent adjuvant that induces strong protective immunity without incurring any significant skin reactogenicity is urgently needed for cutaneous vaccination. Here, we report that a natural agonist of stimulator of interferon genes (STING), 2'3'- cyclic guanosine monophosphate-adenosine monophosphate (cGAMP), robustly augmented and prolonged the cellular and humoral immune responses provoked by H5N1 and 2009 H1N1 pandemic influenza vaccines after a single dose of intradermal, but not intramuscular, immunization. The potency of cGAMP for cutaneous vaccination was ascribed to a large number of antigen-presenting cells resident in the skin and ready for immediate activation when cGAMP was injected. However, its potency was severely compromised in the muscle, because antigen-presenting cells could not be promptly recruited to the injection site before the injected cGAMP was diffused out. The superior adjuvant effect and safety of cGAMP were also confirmed in a more clinically relevant swine model of skin. The vigorous immune responses elicited by cGAMP with no overt skin irritation was attributable to its stay in the skin, which was brief but sufficient to activate dermal dendritic cells. This small and well-characterized self-molecule holds great promise as an ideal adjuvant for cutaneous vaccination.

IEX-1 deficiency induces browning of white adipose tissue and resists diet-induced obesity.

Chronic inflammation plays a crucial role in the pathogenesis of obesity and insulin resistance. However, the primary mediators that affect energy homeostasis remain ill defined. Here, we report an unexpected role for immediate early response gene X-1 (IEX-1), a downstream target of NF-κB, in energy metabolism. We found that IEX-1 expression was highly induced in white adipose tissue (WAT) in both epidydmal and subcutaneous depots but not in interscapular brown adipose tissue (BAT) in mice fed a high fat diet (HFD). Null mutation of IEX-1 protected mice against HFD-induced adipose and hepatic inflammation, hepatic steatosis, and insulin resistance. Unexpectedly, IEX-1 knockout (IEX-1(-/-)) mice gained markedly less weight on HFD for 20 weeks as compared to wild-type (WT) littermates (37 ± 3 versus 48 ± 2 gm) due to increased energy expenditure. Mechanistically, we showed that IEX-1 deficiency induced browning and activated thermogenic genes program in WAT but not in BAT by promoting alternative activation of adipose macrophages. Consequently, IEX-1(-/-) mice exhibited enhanced thermogenesis (24 ± 0.1 versus 22 ± 0.1 kcal/hour/kg in WT mice) explaining increased energy expenditure and lean phenotype in these mice. In conclusion, the present study suggests that IEX-1 is a novel physiological regulator of energy homeostasis via its action in WAT.

Low-level light in combination with metabolic modulators for effective therapy of injured brain.

Vascular damage occurs frequently at the injured brain causing hypoxia and is associated with poor outcomes in the clinics. We found high levels of glycolysis, reduced adenosine triphosphate generation, and increased formation of reactive oxygen species and apoptosis in neurons under hypoxia. Strikingly, these adverse events were reversed significantly by noninvasive exposure of injured brain to low-level light (LLL). Low-level light illumination sustained the mitochondrial membrane potential, constrained cytochrome c leakage in hypoxic cells, and protected them from apoptosis, underscoring a unique property of LLL. The effect of LLL was further bolstered by combination with metabolic substrates such as pyruvate or lactate both in vivo and in vitro. The combinational treatment retained memory and learning activities of injured mice to a normal level, whereas other treatment displayed partial or severe deficiency in these cognitive functions. In accordance with well-protected learning and memory function, the hippocampal region primarily responsible for learning and memory was completely protected by combination treatment, in marked contrast to the severe loss of hippocampal tissue because of secondary damage in control mice. These data clearly suggest that energy metabolic modulators can additively or synergistically enhance the therapeutic effect of LLL in energy-producing insufficient tissue-like injured brain.