Hematoporphyrin derivative-mediated photodynamic techniques for the diagnosis and treatment of chordoma.

BACKGROUND: Chordoma is a rare congenital low-grade malignant tumor characterized by infiltrative growth. It often tends to compress important intracranial nerves and blood vessels, making its surgical treatment extremely difficult. Besides, the efficacy of radiotherapy and chemotherapy is limited. The photosensitizer hematoporphyrin derivative (HPD) can emit red fluorescence under 405 nm excitation and produce reactive oxygen species for tumor therapy under 630 nm excitation. Herein, we investigated the effects of the photosensitizer hematoporphyrin derivative (HPD) on different cell lines of chordoma and xenograft tumors under 405 nm and 630 nm excitation. METHODS: The photosensitizer hematoporphyrin derivative (HPD) and Two different chordoma cell lines (U-CH1, JHC7) were used for the test. The in vitro experiments were as follows: (1) the fluorescence intensity emitted by chordoma cells excited by different 405 nm light intensities was observed under a confocal microscope; (2) the Cell Counting Kit-8 (CCK-8) assay was performed to detect the effects of different photosensitizer concentrations and 630 nm light energy densities on the activity of chordoma cells. In the in vivo experiments, (3) Fluorescence visualization of chordoma xenograft tumors injected with photosensitizer via tail vein under 405 nm excitation; (4) Impact of 630 nm excitation of photosensitizer on the growth of chordoma xenograft tumors. RESULTS: (1) The photosensitizers in chordoma cells and chordoma xenografts of nude mice were excited by 405 nm to emit red fluorescence; (2) 630 nm excitation photosensitizer reduces chordoma cell activity and inhibits chordoma xenograft tumor growth in chordoma nude mice. CONCLUSION: Photodynamic techniques mediated by the photosensitizer hematoporphyrin derivatives can be used for the diagnosis and treatment of chordoma.

Preparation of Hematoporphyrin-Poly(Lactic Acid) Nanoparticles Encapsulated Perfluoropentane/Salicylic Acid for Enhanced US/CEST MR Bimodal Imaging.

Background: Medical imaging modalities, such as magnetic resonance imaging (MRI), ultrasound, and fluorescence imaging, have gained widespread acceptance in clinical practice for tumor diagnosis. Each imaging modality has its own unique principles, advantages, and limitations, thus necessitating a multimodal approach for a comprehensive disease understanding of the disease process. To enhance diagnostic precision, physicians frequently integrate data from multiple imaging modalities, driving research advancements in multimodal imaging technology research. Methods: In this study, hematoporphyrin-poly (lactic acid) (HP-PLLA) polymer was prepared via ring-opening polymerization and thoroughly characterized using FT-IR, 1H-NMR, XRD, and TGA. HP-PLLA based nanoparticles encapsulating perfluoropentane (PFP) and salicylic acid were prepared via emulsion-solvent evaporation. Zeta potential and mean diameter were assessed using DLS and TEM. Biocompatibility was evaluated via cell migration, hemolysis, and cytotoxicity assays. Ultrasonic imaging was performed with a dedicated apparatus, while CEST MRI was conducted using a 7.0 T animal scanner. Results: We designed and prepared a novel dual-mode nanoimaging probe SA/PFP@HP-PLLA NPs. PFP enhanced US imaging, while salicylic acid bolstered CEST imaging. With an average size of 74.43 ± 1.12 nm, a polydispersity index of 0.175 ± 0.015, and a surface zeta potential of -64.1 ± 2.11 mV. These NPs exhibit excellent biocompatibility and stability. Both in vitro and in vivo experiments confirmed the SA/PFP@HP-PLLA NP's ability to improve tumor characterization and diagnostic precision. Conclusion: The SA/PFP@HP-PLLA NPs demonstrate promising dual-modality imaging capabilities, indicating their potential for preclinical and clinical use as a contrast agent.

Sequence-dependent effects of hematoporphyrin derivatives (HPD) photodynamic therapy and cisplatin on lung adenocarcinoma cells.

BACKGROUND: Hematoporphyrin derivatives (HPD)-Photodynamic therapy (PDT) in combination with cisplatin (DDP) is an effective anticancer strategy. However, whether the order of combination affects efficacy has not been studied. METHODS: The human lung adenocarcinoma (LUAD) A549 cells were used as the study subjects. After A549 cells were treated with a single medication (PDT/DDP) or a sequential combination (PDT + DDP / DDP + PDT), the cell viability was assayed using the cell counting kit-8 method. Hoechst staining, Annexin-V/propidium iodide (PI) double staining, western blotting, and a real-time quantitative polymerase chain reaction (RT-qPCR) were performed to examine the mechanisms behind the combined effects. RESULTS: A synergistic impact between HPD-PDT and DDP was found. The cell viability in the PDT+DDP group was significantly lower than in the DDP+PDT group. A significant apoptotic profile and a high apoptotic rate were seen in the PDT + DDP group. The western blot showed that the expression levels of Bcl2-associated x(Bax) and cleaved-poly ADP-ribose polymerase (PARP) increased, and those of B-cell lymphoma-2 (Bcl-2) and Caspase-9 decreased in the PDT + DDP group. At the same time, the RT-qPCR revealed the upregulation of Bax and PARP mRNA and the downregulation of Bcl-2 and Caspase-9 mRNA. CONCLUSION: The order of the combination therapy (PDT + DDP / DDP + PDT) was important. The HPD-PDT followed by DDP significantly inhibited LUAD cell viability, which may be related to the mitochondrial apoptotic pathway.

Simulating PDT of port-wine stains in the in vivo chicken wattle model using Hemoporfin and radiation at 532 nm: Comparison of a LED and a laser source.

Port-wine stain (PWS) birthmarks are congenital capillary malformations occurring in 0.3 %∼0.5 % of newborns. Hemoporfin-mediated vascular-acting photodynamic therapy (Hemoporfin PDT) is an emerging option for treating PWS. This in vivo study aimed to compare laser and light-emitting diodes (LED) as light source for Hemoporfin PDT. Chicken wattles were used as the animal model. Color and histopathological changes were evaluated after combining Hemoporfin with KTP laser or LED light source of 532 nm at the same doses. Both PDT approaches could induce significant vascular injury and color bleaching. Although the use of the laser resulted in a greater vascular clearance, the LED showed more uniform distribution both in the beam profiles and tissue reaction and exhibited better safety. This in vivo study suggests that the LED is a favorable choice for larger PWS lesion.

Enhanced cavitation dose and reactive oxygen species production in microbubble-mediated sonodynamic therapy for inhibition of Escherichia coli and biofilm.

Sonodynamic therapy (SDT) is an emerging antibacterial therapy. This work selected hematoporphyrin monomethyl ether (HMME) as the sonosensitizer, and studied the enhanced inhibition effect of Escherichia coli and biofilm by microbubble-mediated cavitation in SDT. Firstly, the influence of microbubble-mediated cavitation effect on different concentrations of HMME (10 µg/ml, 30 µg/ml, 50 µg/ml) was studied. Using 1,3-diphenylisobenzofuran (DPBF) as an indicator, the effect of microbubble-mediated cavitation on the production of reactive oxygen species (ROS) was studied by absorption spectroscopy. Secondly, using agar medium, laser confocal microscopy and scanning electron microscopy, the effect of microbubble-mediated cavitation on the activity and morphology of bacteria was studied. Finally, the inhibitory effect of cavitation combined with SDT on biofilm was evaluated by laser confocal microscopy. The research results indicate that: (1) Microbubble-mediated ultrasound cavitation can significantly increase cavitation intensity and production of ROS. (2) Microbubble-mediated acoustic cavitation can alter the morphological structure of bacteria. (3) It can significantly enhance the inhibition of SDT on the activity of Escherichia coli and its biofilm. Compared with the control group, the addition of microbubbles resulted in an increase in the number of dead bacteria by 61.7 %, 71.6 %, and 76.2 %, respectively. The fluorescence intensity of the biofilm decreased by 27.1 %, 80.3 %, and 98.2 %, respectively. On the basis of adding microbubbles to ensure antibacterial and biofilm inhibition effects, this work studied the influence of cavitation effect in SDT on bacterial structure, providing a foundation for further revealing the intrinsic mechanism of SDT.

Influential Factors in the Efficacy of Hemoporfin-Mediated Photodynamic Therapy for Port-wine Stains.

Hemoporfin-mediated photodynamic therapy (HMME-PDT) is commonly used in the treatment of port-wine stains (PWS). However, the influential factors for the efficacy of the treatment are not well defined. This study intends to observe the influential factors for the efficacy of HMME-PDT in the treatment of port-wine stains (PWS). A total of 551 patients with PWS of head and neck was enrolled in this retrospective study. Further screening the patients of facial PWS, 484 patients were chosen. Patients were treated with HMME-PDT. All patients received 1~3 sessions of treatment with 2~3-month intervals. We photographed the lesions before each session and 2~3 months after the last session. Ages, sessions, lesion subtypes, and previous treatment history were related to the response of HMME-PDT (P =0.032, P<0.001, P=0.012, P=0.003 respectively). Treatment sessions were the independent factor correlated with efficacy after 3 sessions of treatment. Patients with no treatment history targeting PWS showed higher efficacy than those were treated with laser or other photodynamic treatment (P<0.05). The efficacy was higher by increasing the sessions of treatment. The efficacy was higher for lesion on maxillary prominence area and mandibular prominence area that on frontonasal prominence area and optic vesicle area (P<0.05). HMME-PDT is an effective in the treatment of PWS. Patients received no previous treatment for PWS, total treatment sessions and lesion on maxillary prominence area and mandibular prominence area are positive factors.

Study of hematoporphyrin monomethyl ether-mediated blue-light photodynamic therapy in the treatment of oral ulcers infected with Staphylococcus aureus: In vivo evaluation.

Clinical suppurative infection is mainly caused by Staphylococcus aureus. Although many antibiotics can be used to kill S. aureus, the resulting resistance problem is difficult to solve. Therefore, it is necessary to seek a new sterilizing method to solve the problem of drug resistance of S. aureus and improve the therapeutic effect of infectious diseases. Photodynamic therapy (PDT) has become an alternative for the treatment of a variety of drug-resistant infectious diseases due to its advantages of non-invasive, specific targeting, and no drug resistance. We have confirmed the advantages and experimental parameters of blue-light PDT sterilization in vitro experiments. This study aimed to treat buccal mucosa ulcer of hamster infected with S. aureus according to the parameters obtained in vitro experiment, and observe the bactericidal effect of hematoporphyrin monomethyl ether (HMME) mediated blue-light PDT in vivo and its therapeutic effect on tissue infection. The results indicated that HMME mediated blue-light PDT can effectively kill S. aureus in vivo and promote the healing of the oral infectious wound.The study findings lay a foundation for carrying out more HMME mediated blue-light PDT sterilizing therapy.

Nanotechnology-augmented sonodynamic therapy and associated immune-mediated effects for the treatment of pancreatic ductal adenocarcinoma.

PURPOSE: Sonodynamic therapy (SDT) is emerging as a cancer treatment alternative with significant advantages over conventional therapies, including its minimally invasive and site-specific nature, its radical antitumour efficacy with minimal side effects, and its capacity to raise an antitumour immune response. The study explores the efficacy of SDT in combination with nanotechnology against pancreatic ductal adenocarcinoma. METHODS: A nanoparticulate formulation (HPNP) based on a cathepsin B-degradable glutamate-tyrosine co-polymer that carries hematoporphyrin was used in this study for the SDT-based treatment of PDAC. Cathepsin B levels in BxPC-3 and PANC-1 cells were correlated to cellular uptake of HPNP. The HPNP efficiency to induce a sonodynamic effect at varying ultrasound parameters, and at different oxygenation and pH conditions, was investigated. The biodistribution, tumour accumulation profile, and antitumour efficacy of HPNP in SDT were examined in immunocompetent mice carrying bilateral ectopic murine pancreatic tumours. The immune response profile of excised tumour tissues was also examined. RESULTS: The HPNP formulation significantly improved cellular uptake of hematoporphyrin for both BxPC-3 and PANC-1 cells, while increase of cellular uptake was positively correlated in PANC-1 cells. There was a clear SDT-induced cytotoxicity at the ultrasound conditions tested, and the treatment impaired the capacity of both BxPC-3 and PANC-1 cells to form colonies. The overall acoustic energy and pulse length, rather than the power density, were key in eliciting the effects observed in vitro. The SDT treatment in combination with HPNP resulted in 21% and 27% reduction of the target and off-target tumour volumes, respectively, within 24 h. A single SDT treatment elicited an antitumour effect that was characterized by an SDT-induced decrease in immunosuppressive T cell phenotypes. CONCLUSION: SDT has significant potential to serve as a monotherapy or adjunctive treatment for inoperable or borderline resectable PDAC.

Docetaxel prodrug and hematoporphyrin co-assembled nanoparticles for anti-tumor combination of chemotherapy and photodynamic therapy.

To realize the synergistic anti-tumor effect of chemotherapy and photodynamic therapy, the mono sulfide-modified docetaxel (DTX) prodrugs (DSD) provided by our laboratory and hematoporphyrin (HP) were used to physically prepare co-assembled nanoparticles (DSD/HP NPs) by nano-precipitation. For the first time, this study showed its characteristics, in vitro anti-tumor activity, pharmacokinetic behavior in rats, in vivo distribution, and pharmacodynamic effects on 4T1 tumor-bearing Bal b/c mice. DSD/HP NPs optimized by single-factor and response surface optimization had several distinct characteristics. First, it had dark purple appearance with particle size of 105.16 ± 1.24 nm, PDI of 0.168 ± 0.15, entrapment efficiency and drug loading of DSD and HP in DSD/HP NPs of 96.27 ± 1.03% and 97.70 ± 0.20%, 69.22 ± 1.03% and 20.03 ± 3.12%, respectively. Second, it had good stability and could release DTX and HP slowly in the media of pH 7.4 PBS with 10 mM DTT (H2O2). Moreover, DSD/HP NPs along with NiR treatment significantly inhibited 4T1 cells proliferation, and induced more reactive oxygen species and cells apoptosis. In vivo pharmacokinetic and pharmacodynamic studies showed that DSD/HP NPs could prolong the drug circulation time in rats, increase drug distribution in tumor site, obviously inhibit tumor growth, and decrease the exposure of drug to normal tissues. Therefore, DSD/HP NPs as a promising co-assembled nano-drug delivery system could potentially improve the therapeutic efficiency of chemotherapeutic drug and achieve better anti-tumor effects due to the combination of chemotherapy and photodynamic therapy.

Hematoporphyrin monomethyl ether mediated photodynamic therapy inhibits oral squamous cell carcinoma by regulating the P53-miR-21-PDCD4 axis via singlet oxygen.

The objective of this study was to determine the mechanism and effect of hematoporphyrin monomethyl ether mediated photodynamic therapy (HMME-PDT) on oral squamous cell carcinoma (OSCC). Human OSCC CAL-27 cells were randomly divided into four groups: control group, HMME group, laser group, and HMME-PDT group. Cell viability was detected by the CCK-8 method. Cell cycle distribution was evaluated by flow cytometry. GEO database was used to screen differentially expressed microRNAs (DEMs), and TCGA database was performed to verify DEM expression in OSCC and normal tissues. The effects of HMME-PDT on DEM expression were assayed by real-time PCR, and the expressions of miRNAs target genes were measured by western blot. Fluorescence probes were used to determine the production of singlet oxygen (1O2). Compared with the other three groups, HMME-PDT dramatically inhibited CAL-27 cell proliferation and induced G0/G1 cycle arrest. The expressions of miR-21 and miR-155 were significantly upregulated in OSCC. HMME-PDT downregulated the expression of miR-21 but had no obvious effect on miR-155. HMME-PDT remarkably upregulated the levels of P53 and miR-21 target proteins, such as PDCD4, RECK, and SPRY2. 1O2 was generated during HMME-PDT, and inhibition of 1O2 production could reverse the regulation of HMME-PDT on P53, miR-21, and its target proteins, thus restoring cell viability. HMME-PDT can significantly inhibit the growth of OSCC cells, and the mechanism of this effect is related to the regulation of the P53-miR-21-PDCD4 axis via 1O2 induced by HMME-PDT.

Disinfection efficacy and fracture strength of PMMA denture-based polymer with chlorhexidine, PDT utilizing Rose Bengal and hematoporphyrin, and Er, Cr: YSGG laser.

OBJECTIVE: The study aimed to assess and compare disinfection efficacy and fracture strength of PMMA based DBPs using chlorhexidine (CHX), PDT utilizing Rose Bengal (RB) and hematoporphyrin HPD, and Er,Cr:YSGG laser (ECL) induced antimicrobial action against in-vitro biofilms colonized with C.albicans, S.aureus, S. mutans, and E. coli. MATERIAL AND METHODS: American Type Culture Collection (ATCC) of C.albicans, S.aureus, S. mutans and E. coli were cultured. Forty-eight PMMA-based denture base plates (DBPs) were prepared by heat-cure acrylic resin and contaminated by in-vitro biofilm under-stimulated in-vitro conditions. DBPs were treated with group1; ECL group 2; RB 5µm, group 3; HPD 500 mg/L and group 4; 0.12% CHX (controls) respectively, for the disinfection of biofilms. All photosensitizers (PS) were activated by LED at a different wavelength. Each contaminated DBP was sprayed on all its surfaces with the aforementioned photosensitizers and CHX. One-way analysis of variance (ANOVA) was used to test the efficacy of disinfection and fracture load testing. Tukey multiple comparison tests were performed to compare means of CFU/mL (log10) for exposed E. coli, C. albicans, S aureus, and S. mutans. RESULTS: Specimens in group 1 disinfected with erbium laser, group 3 disinfected with HPD, and group 4 sterilized with chemical disinfection were effective in decreasing bacterial load CFU/mL (log10) against C.albicans, S. aureus, S.mutans, and E. Coli (p>0.05). Intergroup comparison demonstrated ECL, CHX and HPD demonstrated a comparable reduction against C.albicans and S.aureus (p>0.05). Similarly, S.mutans and E.coli were sensitive against all experimental groups (p>0.05). no significant difference in fractural load analysis among the different investigated groups was noted (p>0.05). CONCLUSION: ECL and HPD photosensitizer revealed a significant reduction in CFU/ml of exposed viable colonies of C.albicans, S.aureus, S. mutans, and E. coli comparable to 0.12% CHX mediated disinfection of PMMA-based DBPs. Irrespective of the type of disinfection no influence of disinfection was noted on the fracture load of PMMA DBPs.

Co-delivery of gefitinib and hematoporphyrin by aptamer-modified fluorinated dendrimer for hypoxia alleviation and enhanced synergistic chemo-photodynamic therapy of NSCLC.

Although epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs)-based molecular targeted therapy are proved to be effective in the treatment of non-small cell lung cancer (NSCLC) with EGFR mutation, its efficacy is limited by the acquired drug resistance. The combination of EGFR-TKIs with photodynamic therapy (PDT) has been explored to combat NSCLC with promising synergistic results. However, hypoxic tumor microenvironment is associated with the development of EGFR-TKIs resistance and severely limits the efficacy of PDT. Here, we synthesized an aptamer modified fluorinated dendrimer (APF) as a drug carrier and prepared nanocomplexes APFHG by encapsulation of gefitinib (Gef) and hematoporphyrin (Hp). APF has good oxygen-carrying capacity, high drug entrapment efficiency, and could release Gef and Hp in response to intracellular pH. APF can specifically recognize EGFR-positive NSCLC cells and effectively improve the tumor hypoxic microenvironment due to the targeting effect of aptamer and the good oxygen-carrying capacity of the fluorinated dendrimer. Under the laser irradiation, APFHG can significantly increase the production of the intracellular reactive oxygen species and produce a synergistic therapeutic effect in inhibition of cellular growth and induction of cell cycle arrest and apoptosis on both Gef-sensitive and Gef-resistant EGFR-mutant NSCLC cells through PDT/molecular targeted therapy. This work indicates that fluorinated dendrimer could be a potent drug delivery platform to overcome hypoxia-related resistance and the co-delivery of EGFR-TKI and photosensitizer by the fluorinated dendrimer could be a promising therapeutic approach for reversal of EGFR-TKIs resistance in EGFR mutation-positive NSCLC.

A new antibacterial nano-system based on hematoporphyrin-carboxymethyl chitosan conjugate for enhanced photostability and photodynamic activity.

To promote bactericidal activity, improve photostability and safety, novel antibacterial nanoparticle system based on photodynamic action (PDA) was prepared here through conjugation of photosensitizer hematoporphyrin (HP) onto carboxymethyl chitosan (CMCS) via amide linkage and followed by ultrasonic treatment. The system was stable in PBS (pH 7.4) and could effectively inhibit the photodegradation of conjugated HP because of aggregation-caused quenching effect. ROS produced by the conjugated HP under light exposure could change the structure of nanoparticles by oxidizing the CMCS skeleton and thereby significantly promote the photodynamic activity of HP and its photodynamic activity after 6 h was higher than that of HP·2HCl under the same conditions. Antibacterial experiments showed that CMCS-HP nanoparticles had excellent photodynamic antibacterial activity, and the bacterial inhibition rates after 60 min of light exposure were greater than 97%. Safety evaluation exhibited that the nanoparticles were safe to mammalian cells, showing great potential for antibacterial therapy.

Photodynamic activity of Photogem® in Leishmania promastigotes and infected macrophages.

Objectives: This study aimed to evaluate the effect of photodynamic therapy (PDT) with Photogem® in promastigotes of Leishmania braziliensis and Leishmania major, and in infected macrophages. Materials & methods: The following parameters were analyzed: Photogem® internalization, mitochondrial activity, viability, tubulin marking and morphological alterations in promastigotes and viability in infected macrophages. Results: Photogem® accumulated in the cytosol and adhered to the flagellum. Changes were observed in the mitochondrial activity in groups maintained in the dark, with no viability alteration. After PDT, viability decreased up to 80%, and morphology was affected. Conclusion: The results point out that PDT with Photogem® can reduce parasite and macrophage viability.

Crosstalk between oxidative stress, autophagy and apoptosis in hemoporfin photodynamic therapy treated human umbilical vein endothelial cells.

BACKGROUND: Photodynamic therapy (PDT) provides a treatment for port-wine stain (PWS) using hemoporfin (hematoporphyrin monomethyl ether, HMME), a novel photosensitizer, reporting better efficacy and lower recurrence rate. This study investigated the effects of HMME-PDT on human umbilical vein endothelial cells (HUVECs) as well as underlying mechanisms. METHODS: Cell proliferation ability was measured by CCK8 assay and cell apoptosis was determined by TUNEL assay and Western blot analysis. Confocal fluorescence microscopy monitoring RFP-GFP-LC3 transfected HUVECs and Western blot analysis were used to evaluate autophagy. 3-Methyladenine (3-MA), Z-VAD-FMK, N-acetylcysteine (NAC) were used for inhibitor studies. RESULTS: HMME-PDT decreased cell proliferation ability in an HMME concentration and light dose-dependent manner. Oxidative stress played an important role in HMME-PDT induced cell apoptosis and autophagy in HUVECs. Pretreatment with Z-VAD-FMK, the inhibitor of apoptosis, enhanced HMME-PDT induced autophagy. 3-MA, the suppressor of autophagy, significantly increased HMME-PDT induced apoptosis rates. CONCLUSIONS: Our study demonstrated that HMME-PDT induced both apoptosis and autophagy in HUVECs via oxidative stress. Our data suggested that HMME-PDT- induced autophagy was able to prevent apoptotic cell death of HUVECs and rendered them more resistant to HMME-PDT induced toxicity.

The penetration effect of HMME-mediated low-frequency and low-intensity ultrasound against the Staphylococcus aureus bacterial biofilm.

BACKGROUND: The purpose of this study was to observe the effect of hematoporphyrin monomethyl ether (HMME)-mediated low-frequency and low-intensity ultrasound on mature and stable Staphylococcus aureus (S. aureus) biofilms under different ultrasound parameters. METHODS: The biofilm was formed after 48-h culture with stable concentration of bacterial solution. Different types of ultrasound and time were applied to the biofilm, and the ultrasonic type and time of our experiments were determined when the biofilm was not damaged. The penetration effects of low-frequency and low-intensity ultrasound were decided by the amount of HMME that penetrated into the biofilm which was determined by fluorescence spectrometry. RESULTS: The destruction of biofilms by pulse waveform was the strongest. Sinusoidal low-frequency and low-intensity ultrasound can enhance the biofilm permeability. For a period of time after the ultrasound was applied, the biofilm permeability increased, however, changes faded away over time. CONCLUSIONS: Low-frequency and low-intensity sinusoidal ultrasound significantly increased the permeability of the biofilms, which was positively correlated with the time and the intensity of ultrasound. Simultaneous action of ultrasound and HMME was the most effective way to increase the permeability of the biofilms.

Immunoconjugates to increase photoinactivation of bovine alphaherpesvirus 1 in semen.

Artificial insemination and in vitro embryo production are increasingly used to improve the reproductive efficiency of herds, however success of these techniques depends on the sanitary quality of the semen. Insemination centers commonly use antibiotics in their routine procedure, but they are not able against viruses. In this paper, we demonstrate a new approach for disinfecting virus in bovine semen using photoimmunoinactivation, an adaptation of the photodynamic inactivation (PDI) methodology. The photosensitizers (PSs), hematoporphyrin (HP) and zinc tetracarboxy-phthalocyanine (ZnPc) were conjugated to Immunoglobulin Y (IgY) anti-bovine alphaherpesvirus 1 (BoHV-1) and used for PDI against the BoHV-1 viruses in cell culture and compared to the unconjugated PSs. Both treatments proved to be efficient, but a significant decrease in the irradiation time required to completely eliminate the virus was observed in the samples treated with the immunoconjugates. Photophysical measurements help us to understand the coupling between PSs and IgY and the evaluated production of singlet oxygen. Following the cell culture test, the same approach was applied in semen artificially infected with BoHV-1. The immunoconjugates were also efficient for complete virus inactivation up to 5 min of irradiation and proved to be safe using several parameters of sperm viability, demonstrating the feasibility of our strategy for disinfection viruses in semen.

Effect of 532nm photodynamic therapy with hemoporfin on the expression of vascular endothelial growth factor in cultured human vascular endothelial cells.

BACKGROUND: Photodynamic therapy (PDT) with hemoporfin (hematoporphyrinmonomethyl ether, HMME) and 532 nm continuous-wave lasers is effective for port-wine stains (PWS) and is considered as a promising treatment modality. The vascular endothelial growth factor (VEGF) is involved in the development of PWS. This study aimed to investigate the effect of 532 nm-HMME-PDT on the in vitro expression of VEGF in human umbilical vein endothelial cells (HUVECs) exposed to different power levels of 532 nm laser and different concentrations of HMME. METHOD: The CCK-8 assay was performed to assess cell viability. Enzyme-linked immunosorbent assay (ELISA) was performed to measure VEGF secretion. VEGF and VEGF receptor (VEGFR) mRNA expression levels were detected by quantitative reverse transcription-polymerase chain reaction (qRT-PCR). RESULTS: Within 72 h after HMME-PDT, cell viability was inhibited, secretion and expression of VEGF was decreased, and expression of VEGFR mRNA was significantly decreased with the increase of HMME concentration. CONCLUSION: 532 nm-HMME-PDT can downregulate the expression of VEGF and VEGFR mRNA. Future studies are necessary to examine the HMME doses to achieve better efficacy with fewer adverse effects.

Hematoporphyrin monomethyl ether mediated sonodynamic antimicrobial chemotherapy on porphyromonas gingivalis in vitro.

This study aimed to evaluate the efficacy of hematoporphyrin monomethyl ether (HMME)-mediated sonodynamic antimicrobial chemotherapy (SACT) on Porphyromonas gingivalis (P. gingivalis). P. gingivalis (ATCC 33277) was used in the present study. The bacterial suspension was randomly divided into five groups: Group 1 was incubated for 2 h in the dark with HMME in various concentrations (10, 20, 30 and 40 µg/mL). Then exposed to 1 MHz ultrasound frequency with 3 W/cm2 ultrasound intensity for 10 min. Group 2 was incubated with 40 µg/mL HMME and then irradiated with 2, 4, 6, 8 and 10 min ultrasonic time. Group 3 received different HMME concentration (10, 20, 30 and 40 µg/mL) treatment alone with no ultrasound as the HMME control group. Group 4 received ultrasound treatment alone in different ultrasonic time (2, 4, 6, 8 and 10 min) with no HMME as the ultrasound control group. Group 5 received no treatment as the no treatment control group. After the SACT, the bactericidal effect was determined by the colony forming unit assay. The intracellular content of reactive oxygen species (ROS) was detected using the laser scanning confocal microscope based on DCFH-DA. 4.7 lg reduction in CFU, When P. gingivalis was treated with ultrasound (3 W/cm2 for 10 min) at 40 µg/mL HMME concentration (P < 0.01). The intracellular ROS in SDT group had a significant difference in comparison with the no treatment control group (P < 0.01). HMME mediated SACT can be a potential antibacterial therapy to significantly inhibit P. gingivalis growth.

Multi-path tumor inhibition via the interactive effects between tumor microenvironment and an oxygen self-supplying delivery system for a photosensitizer.

The tumor microenvironment (TME) affects not only tumor growth and metastasis, but also therapy efficacy. In the present study, an oxygen self-supplying delivery system for a photosensitizer (O2-PDS), with an oxygen source of calcium peroxide (CPO), has been designed to induce multi-path tumor apoptosis through interactive effects with the TME. In anti-tumor experiments, the CPO decomposition and O2 released from the O2-PDS are site-activated and accelerated by high interstitial pressure and low pH level of the TME. The CPO decomposition products of O2 and Ca2+ lead to direct tumor apoptosis by irradiation generated singlet oxygen and mitochondrial calcium overload. The decomposition products of OH- and O2 relieves the acid and hypoxic state of TME, inducing a decrease in tumor proliferation and metastasis. This multi-path tumor apoptosis leads to a positive therapeutic effect on an animal tumor model and nontoxicity in normal tissue.