Preventive and therapeutic vascular photobiomodulation decreases the inflammatory markers and enhances the muscle repair process in an animal model.

Photobiomodulation therapy (PBM) has shown positive effects when applied locally to modulate the inflammatory process and facilitate muscle repair. However, the available literature on the mechanisms of action of vascular photobiomodulation (VPBM), a non-invasive method of vascular irradiation, specifically in the context of local muscle repair, is limited. Thus, this study aimed to assess the impact of vascular photobiomodulation (VPBM) using a low-level laser (LLL) on the inflammatory response and the process of skeletal muscle repair whether administered prior to or following cryoinjury-induced acute muscle damage in the tibialis anterior (TA) muscles. Wistar rats (n = 85) were organized into the following experimental groups: (1) Control (n = 5); (2) Non-Injury + VPBM (n = 20); (3) Injured (n = 20); (4) Pre-VPBM + Injury (n = 20); (5) Injury + Post-VPBM (n = 20). VPBM was administered over the vein/artery at the base of the animals' tails (wavelength: 780 nm; power: 40 mW; application area: 0.04 cm2; energy density: 80 J/cm2). Euthanasia of the animals was carried out at 1, 2, 5, and 7 days after inducing the injuries. Tibialis anterior (TA) muscles were collected for both qualitative and quantitative histological analysis using H&E staining and for assessing protein expression of TNF-α, MCP-1, IL-1ß, and IL-6 via ELISA. Blood samples were collected and analyzed using an automatic hematological analyzer and a leukocyte differential counter. Data were subjected to statistical analysis (ANOVA/Tukey). The results revealed that applying VPBM prior to injury led to an increase in circulating neutrophils (granulocytes) after 1 day and a subsequent increase in monocytes after 2 and 5 days, compared to the Non-Injury + VPBM and Injured groups. Notably, an increase in erythrocytes and hemoglobin concentration was observed in the Non-Injury + VPBM group on days 1 and 2 in comparison to the Injured group. In terms of histological aspects, only the Prior VPBM + Injured group exhibited a reduction in the number of inflammatory cells after 1, 5, and 7 days, along with an increase in blood vessels at 5 days. Both the Prior VPBM + Injured and Injured + VPBM after groups displayed a decrease in myonecrosis at 1, 2, and 7 days, an increase in newly-formed and immature fibers after 5 and 7 days, and neovascularization after 1, 2, and 7 days. Regarding protein expression, there was an increase in MCP-1 after 1 and 5 days, TNF-α, IL-6, and IL-1ß after 1, 2, and 5 days in the Injured + VPBM after group when compared to the other experimental groups. The Prior VPBM + Injured group exhibited increased MCP-1 production after 2 days, in comparison to the Non-Injury + VPBM and Control groups. Notably, on day 7, the Injured group continued to show elevated MCP-1 protein expression when compared to the VPBM groups. In conclusion, VPBM effectively modulated hematological parameters, circulating leukocytes, the protein expression of the chemokine MCP-1, and the proinflammatory cytokines TNF-α and IL-1ß, ultimately influencing the inflammatory process. This modulation resulted in a reduction of myonecrosis, restoration of tissue architecture, increased formation of newly and immature muscle fibers, and enhanced neovascularization, with more pronounced effects when VPBM was applied prior to the muscle injury.

Cholesterol depletion induces mesenchymal properties in oral squamous cell carcinoma cell line.

BACKGROUND: Cholesterol in cell membranes is crucial for cell signaling, adhesion, and migration. Membranes feature cholesterol-rich caveolae with caveolin proteins, playing roles in epithelial-mesenchymal transition and cancer progression. Despite elevated cholesterol levels in tumors, its precise function and the effects of its depletion in oral squamous cell carcinoma remain unclear. The aim of this study was to evaluate the influence of cholesterol depletion in oral squamous cell carcinoma cell line and epithelial-mesenchymal transition process. METHODS: Cholesterol depletion was induced on SCC-9 cells by methyl-ß-cyclodextrin and cell viability, proliferation, apoptosis, and colony formation capacities were evaluated. Gene and protein expressions were evaluated by reverse transcription polymerase chain reaction (RT-qPCR) and Western Blot, respectively, and cell sublocalization was assessed by immunofluorescence. RESULTS: Cholesterol depletion resulted in alteration of oral squamous cell carcinoma cell morphology at different concentrations of methyl-ß-cyclodextrin, as well as decreased cell proliferation and viability rates. Analysis of CAV1 transcript expression revealed increased gene expression in the treated SCC-9 during the 24 h period, at different concentrations of methyl-ß-cyclodextrin: 5 , 7.5, 10, and 15 mM, in relation to parental SCC-9. CAV1 protein expression was increased, with subsequent dose-dependent decrease. A statistically significant difference was observed in samples treated with 5 mM of methyl-ß-cyclodextrin (p = 0.02, Kruskal-Wallis test). The immunofluorescence assay showed lower cytoplasmic and membrane labeling intensity in the treated samples for CAV1. CONCLUSION: These findings indicate the modulation of cholesterol as a possible mechanism underlying the regulation of these molecules and activation of epithelial-mesenchymal transition in oral squamous cell carcinoma.

The supportive use of photobiomodulation on salivary glands: a narrative review and meta-analysis.

PURPOSE: Radiotherapy is one of the main strategies used in the treatment of cancer patients and it can cause early or late xerostomia and/or hyposalivation. Therapeutic management of xerostomia includes oral hygiene, sialogenic agents among others. METHODS: This study reviews the use of extra-oral salivary glands photobiomodulation in treating xerostomia and/or hyposalivation after radiotherapy and performs a meta-analysis of this data. RESULTS: After a broad search of the literature, eight clinical studies were selected. DISCUSSION: In a safe way, the studies found that extra-oral stimulation of the salivary glands has benefits in the hyposalivation and changes in salivary flow resulting from lesions by radiotherapy. A meta-analysis found significant values in pain comparing the pre- and post-treatment moments (MD - 3.02, I2 95%, IC - 5.56; - 0.48) and for stimulated salivary flow at 30 days after the end of radiotherapy (MD 2.90, I2 95%, IC 1.96; 3.84). CONCLUSION: The most promising parameters comprise wavelengths between 630 and 830 nm, radiant exposure from 2 to 10 J/cm2, two-to-three times a week, before the radiotherapy damage, and homogeneously in the glands. Therefore, Light-Emitting Diode (LED) stimulation of larger areas than the punctual stimulation of small millimeters of the Low-Level Laser Therapy (LLLT) appears to be promising.

Effects of amber LED on inflammatory and regulatory monocytes and lymphocytes.

The primary objective of the present study was to assess the impact of amber LED photobiomodulation (PBM) on human monocytes and lymphocytes that were polarized into proinflammatory and regulatory/reparative phenotypes. Human leukocytes were polarized with LPS or LPS + IL-4 for 2 h and irradiated after 2 and 6 h with amber LED (590 nm). Cell absorbance spectrum and gene and protein expression of IL-1ß, IL-6, IL-10, IL-17, TNF-α and IFNγ determined after 24 h. The results showed that irradiation did not significantly alter absorbance of non-polarized monocytes, whereas irradiated polarized monocytes presented reduction in absorbance in 625-850 nm region. Irradiated monocytes polarized with LPS + IL-4 presented reduction in absorbance in 600-725 nm region compared to non-irradiated group. Irradiated non-polarized lymphocytes presented absorbance peaks between 650 and 820 nm not seen in non-irradiated group. No difference was found in absorbance pattern of polarized lymphocytes after irradiation. Irradiation led to reduction in protein synthesis of IL-6 and TNFα in monocytes polarized to proinflammatory phenotype and increase in production of IL-17 in lymphocytes. Irradiation reduced production of IL-10 in monocytes and lymphocytes polarized to immunoregulatory phenotype. In conclusion, amber LED modulates light absorbance and expression of important cytokines in inflammatory/repair processes in monocytes and lymphocytes.

Effects of photobiomodulation in the experimental acetic acid-induced colitis: comparison between male and female.

Ulcerative colitis (UC) is an important chronic and multifactorial disease, which alters the colon mucosal with a significant impact on life quality affecting both men and women. The difference between genders causes changes in the inflammatory processes, modulating the development of several diseases. The available drugs to treat UC exhibit limited outcomes and side effects; thus, new therapies are needed. Photobiomodulation (PBM) emerges as potential treatment by modulating the inflammatory process without side effects and low costs. The aim of this study was to evaluate the effects of PBM in acetic acid-induced UC comparing the responses between male and females. For this purpose, male and female Wistar rats (36) were submitted to induction of UC by rectal administration of 10% acetic acid (colitis group) and treated or not with PBM (colitis-PBM group) (LED, 660 nm, 100 mW, 150 s) in three points: right side and left of the ventral surface and in the external anal region. Non-manipulated rats were used as control (basal group). We investigated the disease activity index (DAI score), myeloperoxidase enzyme activity (MPO) and release of cytokines in the intestine homogenates, and histological analysis. PBM reduces DAI score, MPO activity, and mast cell degranulation while increased mucous production in both females and males. Moreover, PBM reduced histopathological score as well as the levels of IL-6 and IL-4 in the bowel only in males. We also showed reduced levels of IL-1beta and TNF-alpha after PBM in both males and females, while the levels of IL-10 and IFN-gamma were increased. In conclusion, despite our study has shown some differences between males and females, PBM attenuated the biomarkers of UC in both genders constituting a potential combined treatment that is non-invasive and low cost.

Effects of the invasive and non-invasive systemic photobiomodulation using low-level laser in experimental models: A systematic review.

Systemic photobiomodulation (PBM) of the blood or over blood vessels has been associated with bio-stimulating, vasodilating, and anti-inflammatory properties. This treatment modality has been used for modulating inflammatory processes, tissue repair, atherosclerosis, and systemic arterial hypertension, and is described more often in clinical studies than experimental models. Therefore, the aim of the present study was to conduct a literature review regarding the effect of systemic PBM involving the intravascular laser irradiation of blood (ILIB) or non-invasive vascular photobiomodulation (VPBM) using low-level laser (LLL) in experimental (animal) models. The PubMed/MEDLINE®, Scopus, SPIE Digital Library, and Web of Science databases were searched for articles on the use of VPBM with LLL in animal models. Nine original articles met the inclusion criteria and were critically evaluated. The variables of interest were the dosimetric laser parameters, different methods for delivering energy, and the main results. The use laser in the red spectrum was more prevalent and VPBM (non-invasive) predominated over ILIB (invasive). No standardization was found in the dosimetric parameters. However, the studies showed the positive effects of VPBM on arterial pressure and blood circulation, the positive effects of ILIB on blood composition and hematological markers, as well as positive effects of both forms of systemic PBM (ILIB and VPBM) on the tissue repair process. In conclusion, the studies evaluated in the present review showed that the use of systemic PBM with ILIB or non-invasive VPBM induced positive effects, modulating metabolic conditions and tissue repair. However, there is a need for standardization in the dosimetric parameters for the different conditions and processes evaluated using experimental models.

Loss of Caveolin-1 Expression in Tumor Cells is Associated with Increased Aggressiveness and Cell Invasion in Oral Squamous Cell Carcinoma.

BACKGROUND: Changes in Caveolin-1 (CAV-1) expression are related to tumorigenesis. The aim of this study was to evaluate the role of CAV-1 in tumor progression in oral squamous cell carcinoma (SCC) tissue samples and the effect of CAV-1 silencing on two oral tongue SCC (OTSCC) cell lines (SCC-25, from a primary tumor, and HSC-3 from lymph node metastases). METHODS: Mycroarray hybridization, mRNA expression, and immunohistochemistry were performed on OSCC tissue samples and corresponding non-tumoral margin tissues. The effects of CAV-1 silencing (siCAV-1) on cell viability, membrane fluidity, on the expression of epithelial to mesenchymal transition (EMT) markers and on cell migration and invasion capacity of OTSCC cell lines were evaluated. RESULTS: Microarray showed a greater CAV-1 expression (1.77-fold) in OSCC tumors than in non-tumoral tissues and 2.0-fold more in less aggressive OSCCs. However, significant differences in CAV-1 gene expression were not seen between tumors and non-tumoral margins nor CAV-1 with any clinicopathological parameters. CAV-1 protein was localized both in carcinoma and in spindle cells of the tumor microenvironment (TME), and CAV-1 positive TME cells were associated with smaller/more aggressive tumors, independent of the carcinoma cells' expression. Silencing of CAV-1 increased cell viability only in SCC-25 cells. It also stimulated the invasion of HSC-3 cells and increased ECAD and BCAT mRNA in these cells; however, the protein levels of the EMT markers were not affected. CONCLUSION: Decreased expression of CAV-1 by tumor cells in OSCC and an increase in the TME were associated with increased cell invasiveness and tumor aggressiveness.

Secretory carcinoma of the salivary gland is rich in lactoferrin: a possible lactational-like differentiation?

PURPOSE: It has been hypothesised that secretory carcinoma of the salivary gland (SCsg) might have a lactational-like differentiation. Therefore, we aimed to assess the immunoexpression of breast hormonal receptors and milk-related proteins in cases of SCsg and other salivary gland tumours with prominent secretory activity. METHODS: Immunohistochemistry against prolactin and growth hormone receptors, lactoferrin, human milk fat globule 1, MUC 1 and MUC4 was performed in twelve cases of SCsg and 47 other salivary gland tumours. RESULTS: Most cases of SCsg were negative for prolactin and growth hormone receptors. All cases of SCsg showed enhanced membranous-cytoplasmic staining for human milk fat globule 1, a pattern seen in other tumour groups. Only SCsg showed widespread strong staining for lactoferrin, concomitantly in the cell compartment and secretion. The other positive tumour types exhibited restricted staining. MUC1 and MUC4 showed no distinct pattern of expression. CONCLUSION: Although SCsg failed to demonstrate a complete lactational-like differentiation, lactoferrin showed a distinctive expression pattern in SCsg compared to other tumour types, which makes it a good marker to help in its differential diagnosis.

Mechanisms involved in cancer stem cell resistance in head and neck squamous cell carcinoma.

Despite scientific advances in the Oncology field, cancer remains a leading cause of death worldwide. Molecular and cellular heterogeneity of head and neck squamous cell carcinoma (HNSCC) is a significant contributor to the unpredictability of the clinical response and failure in cancer treatment. Cancer stem cells (CSCs) are recognized as a subpopulation of tumor cells that can drive and maintain tumorigenesis and metastasis, leading to poor prognosis in different types of cancer. CSCs exhibit a high level of plasticity, quickly adapting to the tumor microenvironment changes, and are intrinsically resistant to current chemo and radiotherapies. The mechanisms of CSC-mediated therapy resistance are not fully understood. However, they include different strategies used by CSCs to overcome challenges imposed by treatment, such as activation of DNA repair system, anti-apoptotic mechanisms, acquisition of quiescent state and Epithelial-mesenchymal transition, increased drug efflux capacity, hypoxic environment, protection by the CSC niche, overexpression of stemness related genes, and immune surveillance. Complete elimination of CSCs seems to be the main target for achieving tumor control and improving overall survival for cancer patients. This review will focus on the multi-factorial mechanisms by which CSCs are resistant to radiotherapy and chemotherapy in HNSCC, supporting the use of possible strategies to overcome therapy failure.

Intravascular laser irradiation of blood (ILIB) used to treat lung diseases: a short critical review.

Intravascular laser irradiation of blood (ILIB) was developed to treat cardiovascular diseases due to its rheological effects. In its original form, ILIB was applied by an intravenous optical fiber, restricting its application. However, this technique was modified to non-invasive irradiation through the radial artery, now called vascular photobiomodulation (VPBM). Many studies have used both, ILIB and VPBM, to treat lung diseases. It is well established that lung diseases affect more than 300 million people worldwide with high morbidity and mortality rates. In this short critical review, we discuss the potential benefits of photobiomodulation to treat lung diseases using these two approaches. The search was performed in the electronic database of MEDLINE (Medical Literature Analysis and Retrieval System Online) via PubMed. The data search was carried out from 1991 to 2017. We selected a total of 10 clinical studies using either ILIB or VPBM, in addition to 2 experimental studies in animals. The respiratory diseases treated in these studies included bronchitis, asthma, pneumonia, and tuberculosis. The results showed overall beneficial effects on lung diseases, characterized by a reduction in the inflammatory cascade and antioxidant effects, improvement of hemodynamic parameters, the efficiency of gas exchange, and reduction of hospitalization periods. In conclusion, all studies showed promising effects of ILIB in both animal and human studies. The studies did not discuss any disadvantages or contraindications. However, further studies are needed in order to understand the dosimetry, and the literature is lacking in randomized, controlled clinical trials. Thus, this review highlights the need for additional studies using this approach.

Cholesterol depletion affects caveolin-1 expression, migration and invasion of oral tongue squamous cell carcinoma cell lines.

INTRODUCTION: Cholesterol is a key lipid molecule within cell membranes. This is especially true in cavelolas, invaginated membrane nanodomains, which present the protein caveolin-1 (CAV-1). It is important to note that this structure is involved in many cell signalling pathways. Additionally, high cholesterol is seen in different tumor types but little is known in regards to oral tongue squamous cell carcinoma (OTSCC). The aim of this study was to evaluate the influence of cholesterol depletion on primary (SCC-25) and metastatic (HSC-3) OTSCC cell lines. MATERIALS AND METHODS: Cell membrane fluidity, cell viability, gene and protein expression of CAV-1 and of epithelial-mesenchymal transition (EMT) markers, cell migration in Myogel and invasion-myoma assay were evaluated after cholesterol depletion with methyl-ß-cyclodextrin (MßCD - 7.5, 10 or 15 mM) RESULTS: Decreased cell viability and increased membrane fluidity of SCC-25 cells was seen with cholesterol depletion but cell viability was less affected and there was no effect on membrane fluidity in HSC-3. Cholesterol depletion also decreased CAV-1 at 6 h but increased it after 24 h.; both epithelial and mesenchymal EMT genes were upregulated after 6 h, followed by downregulation at 24 h in SCC-25. In HSC-3, CAV-1 was downregulated, and E-cadherin gene (ECAD) was upregulated at 6 h. Only the protein ß-catenin in SCC-25 was affected, and cell migration of both cell lines was decreased, affecting SCC-25 more intensely. The invasive capacity within human myoma organotypic model was increased in SCC-25 and decreased in HSC-3. CONCLUSION: Cholesterol depletion affects CAV-1 and ECAD inversely. This affect also depends on cell type since the invasive capacity was augmented in primary cells while decreased in metastatic cells.

Periodontal ligament cells mobilized by transforming growth factor-beta 1 and migrated without stimuli showed enhanced osteogenic differentiation.

OBJECTIVE: This study aimed to analyze the ability of G-CSF and TGF-ß1 to mobilize periodontal ligament stem cells to obtain populations with better potential for proliferation and osteogenic differentiation. DESIGN: Primary cultures were established from the periodontal ligament of Wistar rats. After a cell migration assay, four experimental groups were obtained: PDLSC, composed of the primary culture, non-mobilized cells; MPDLSC, the spontaneously migrated cells; MPDLSC-GCSF, the cells mobilized with G-CSF; and MPDLSC-TGF-ß1, the cells mobilized with TGF-ß1. The expression of mesenchymal stem cell markers was assessed by flow cytometry. Clonogenicity, viability, proliferative potential, and osteogenic differentiation capacity were also analyzed. RESULTS: All the study groups expressed well-known mesenchymal stem cell markers and exhibited clonogenic capacity. The higher proliferation potential was seen in the PDLSC and MPDLSC groups, while the MPDLSC and MPDLSC-TGFß1 groups showed a higher number of mineralized deposits in vitro and higher ALP activity after osteogenic differentiation induction. Cells of all the groups also expressed mRNA of genes associated with osteogenic differentiation without previous induction. CONCLUSIONS: Both agents were able to mobilize stem cells from the periodontal ligament, but G-CSF did not show an advantage, whereas TGF-ß1 appears to direct the cells towards a state of increased osteogenic differentiation. Furthermore, spontaneous cell migration through a membrane was sufficient to enrich the cell population.

The effectiveness of phototherapy for surface decontamination against SARS-Cov-2. A systematic review.

COVID-19 appeared in December 2019, needing efforts of science. Besides, a range of light therapies (photodynamic therapy, ultraviolet [UV], laser) has shown scientific alternatives to conventional decontamination therapies. Investigating the efficacy of light-based therapies for environment decontamination against SARS-CoV2, a PRISMA systematic review of Phototherapies against SARS-CoV or MERS-CoV species discussing changes in viral RT-PCR was done. After searching MEDLINE/PubMed, EMBASE, and Literatura Latino-Americana e do Caribe em Ciências da Saúde we have found studies about cell cultures irradiation (18), blood components irradiation (10), N95 masks decontamination (03), inanimate surface decontamination (03), aerosols decontamination (03), hospital rooms irradiation (01) with PDT, LED, and UV therapy. The best quality results showed an effective low time and dose UV irradiation for environments and inanimate surfaces without human persons as long as the devices have safety elements dependent on the surfaces, viral charge, humidity, radiant exposure. To interpersonal contamination in humans, PDT or LED therapy seems very promising and are encouraged.

Serum and salivary cytokines in patients with oral lichen planus treated with Photobiomodulation.

OBJECTIVES: To evaluate the serum and salivary levels of IL-1ß, IL-6, IL-17A, TNF-α, IL-4, and IL-10 in patients with oral lichen planus (OLP) treated with Photobiomodulation (PBM) and clobetasol propionate 0.05%. MATERIAL AND METHODS: Thirty-four OLP patients were randomized into two groups: Control (clobetasol propionate 0.05%) and PBM (660 nm, 100 mW, 177 J/cm2 , 5 s, 0.5 J per point). Serum and saliva were collected at baseline and at the end of treatment (after 30 days) and evaluated using ELISA. The cytokine results were correlated with pain, clinical subtypes, and clinical scores of OLP. RESULTS: IL-1ß, IL-6, IL-17A, TNF-α, and IL-4 levels were higher in saliva in relation to serum. IL-1ß was the most concentrated cytokine in saliva, and a positive correlation with the severity of OLP was noticed. After treatment with corticosteroid, IL-1ß in saliva decreased significantly. No modulation of all cytokines was observed after PBM. CONCLUSION: IL-1ß appears to be an important cytokine involved in OLP pathogenesis. In addition, the mechanisms of action of PBM do not seem to be linked to the modulation of pro or anti-inflammatory cytokines at the end of treatment. It is possible that this events occurred early during treatment.

Influence of conditioned medium from squamous cell carcinoma of the tongue on lymphoblasts and peripheral blood mononuclear cells.

BACKGROUND: Squamous cell carcinoma (SCC) is the most common malignant neoplasm of the oral cavity and is associated with high morbidity and mortality. Attention has been given to the role of inflammatory cells in carcinogenesis because of the ability of cancer cells to subvert the immune response. However, little is known about how molecules from neoplastic cells interact with lymphoblasts and circulating immune cells. This study aimed to understand the mechanisms by which SCC cells modulate the immune response by analyzing the influence of conditioned medium derived from SCC cell lines on immune cells. METHODS: Lymphoblastic cells (CEM) and peripheral blood mononuclear cells (PBMC) were cultured in a conditioned medium derived from squamous cell carcinoma cells (SCC9 or SCC4) and analyzed for cell viability, CD4/CD8/FOXP3 profile by flow cytometry, and chemokine levels. RESULTS: Conditioned medium derived from SCC4 and SCC9 presented higher concentrations of IL-6 and IL-8 than IL-1ß, IL-10, and IFN-γ. CEM and PBMCs when cultured with conditioned medium derived from SCC4 and SCC9 reduced IL-1ß, IL-8, and IFN-γ concentrations. Conditioned medium from SCC4 increased CD4+ population in both CEM and PBMCs, while in conditioned medium from SCC9 it occurred only in PBMCs. PBMCs when cultured with both conditioned mediums increased CD8+ /FOXP3+ cells. CEM cells when cultured with conditioned medium derived from SCC4 and SCC9 reduced. CONCLUSION: Collectively, our results suggest that the products derived from squamous cell carcinoma on inflammatory cells can promote an immunosuppressed environment by reducing cell viability, changing cytokine expression, and altering the cell immunoprofile.

Responses of melanoma cells to photobiomodulation depend on cell pigmentation and light parameters.

Melanoma is a highly aggressive skin cancer that requires new approaches for its management. Low-level laser therapy, currently named photobiomodulation therapy (PBM), has been used to improve different conditions but its effects and safe use on melanoma remain unexplored. Herein, we investigated the PBM impact on melanoma cells differing by pigmentation using near-infrared (NIR) and red lasers in vitro. In vivo, we evaluated the effects of the red laser on melanoma-bearing mice. Amelanotic (SK-MEL-37) and melanotic (B16F10) cells were exposed in vitro to a NIR (780 nm, 40 mW) or a red laser (660 nm, 40 mW) in 3 different light doses: 30, 90, and 150 J/cm2 and responses were assessed regarding mitochondrial activity, invasiveness, migration, and VEGF production. In vivo, melanoma-bearing mice received the red laser delivering 150 J/cm2 directly to the tumor on 3 consecutive days. Mice were monitored for 15 days regarding tumor progression and mouse survival. We noticed that amelanotic cells were unresponsive to NIR light. In contrast, NIR irradiation at 30 J/cm2 promoted an increase in the invasiveness of pigmented cells, even though all light doses have inhibited cell migration. Regarding the red laser on pigmented cells, the highest light dose (150 J/cm2) decreased the VEGF production and migration. In vivo, melanoma-bearing mice treated with red laser showed smaller tumor volume and longer survival than controls. We conclude that PBM appears to be safe for amelanotic non-pigmented melanoma but triggers different responses in melanotic pigmented cells depending on light parameters. Additionally, a high dose of red laser impairs the invasive behavior of melanoma cells, probably due to the decrease in VEGF synthesis, which may have contributed to tumor arrest and increased mouse survival. These findings suggest that red laser therapy could be a new ally in the supportive care of melanoma patients.

Effects of 5-ALA mediated photodynamic therapy in oral cancer stem cells.

The aim of the present study was to investigate the effects of PDT using the photosensitizer 5-aminoulevulinic acid (5-ALA) in oral squamous cell carcinoma (OSCC) behavior, mainly regarding its role on the cancer stem cell (CSC) phenotypes and in maintenance of the stem cell properties. Two OSCC cell lines were used and divided in the groups: Control, 5-ALA, LED 6 J/cm2 and PDT. MTT and Neutral red assays were used to access cellular viability, cell migration was evaluated by the wound healing assay. The stem cell phenotype was analyzed by flow cytometry to evaluate the CD44high/ESAhigh, CD44high/ESAlow and CD44low populations, by the clonogenic and tumor sphere formation assays as well as by RT-qPCR. The presence of Protoporphyrin IX in each CSC fraction was evaluated by flow cytometry. The OSCC cell lines showed a significant decrease in cell viability and migration after PDT. The percentage of CD44high/ESAhigh cells decreased after PDT, which was associated with an increase in the CD44low cells and with a functional decrease in the colony and sphere formation capacity. CD44high/ESAhigh cells showed increased PpIX, which contributed for their greater sensitivity to PDT. INV gene increased significantly after PDT, indicating cellular differentiation. Altogether, our results demonstrate that 5-ALA mediated PDT decreases not only the fraction of oral CSC but also their functional capabilities, inducing their differentiation.

Mechanisms of immune evasion by head and neck cancer stem cells.

Different mechanisms are involved in immune escape surveillance driven by Oral and Head and Neck Cancer Stem Cells (HNCSCs). The purpose of this review is to show the most current knowledge regarding the main impact of HNCSCs on tumor evasion through immunosuppression, CSCs phenotypes and environmental signals, highlighting strategies to overcome immune evasion. The main results drive the participation of cell surface receptors and secreted products and ligands, the crosstalk between cells, and genetic regulation. The reduction in CD8+ T cell recruitment and decreased effector of anti-PD-1 therapy by cells expressing BMI1 is a key event; Natural Killer cell ligands and cytokines needed for its activation and expansion are crucial to control tumor growth and to target CSCs by immunotherapy; CSCs expressing ALDH1 are related to increased expression of PD-L1, with a positive link between DNMT3b expression; CD276 expression in CSCs can act as a checkpoint inhibitor and together with Activator Protein 1 (AP-1) activation, they create continuous positive feedback that enables immune evasion by suppressing CD8+ T cells and prevent immune cell infiltration in head and neck cancer. These data demonstrate the relevance of the better understanding of the interaction between HNCSCs and immune cells in the tumor microenvironment. The ultimate clinical implication is to ground the choice of optimized targets and improve immune recognition for ongoing treatments as well as the response to approved immunotherapies.