Perineural invasion is a pathologic process of neoplastic dissemination along and invading into the nerves. Perineural invasion is associated with aggressive disease and a greater likelihood of poor outcomes. In this study, 3 of 9 patients with cutaneous squamous cell carcinoma and perineural invasion exhibited poor clinical outcomes. Tumors from these patients expressed high levels of MAGE-A3, a cancer testis antigen that may contribute to key processes of tumor development. In addition to perineural invasion, the tumors exhibited poor differentiation and deep invasion and were subsequently classified as Brigham and Women's Hospital tumor stage 3. Cyclin E, A and B mRNA levels were increased in these tumors compared with normal skin tissues (102.93±15.03 vs. 27.15±4.59, 36.83±19.41 vs. 11.59±5.83, 343.77±86.49 vs. 95.65±29.25, respectively; p<0.05). A431 cutaneous squamous cell carcinoma cells pretreated with MAGE-A3 antibody exhibited a decreased percentage S-phase cells (14.13±2.8% vs. 33.97±1.1%; p<0.05) and reduced closure in scratch assays (43.88±5.49% vs. 61.17±3.97%; p = 0.0058). In a syngeneic animal model of squamous cell carcinoma, immunoblots revealed overexpression of MAGE-A3 and cyclin E, A, and B protein in tumors at 6 weeks. However, knockout of MAGE-A3 expression caused a reduction in tumor growth (mean tumor volume 155.3 mm3 vs. 3.2 mm3) compared with parental cells. These results suggest that MAGE-A3 is a key mediator in cancer progression. Moreover, elevated collagen XI and matrix metalloproteases 3, 10, 11, and 13 mRNA levels were observed in poorly differentiated cutaneous squamous cell carcinoma with perineural invasion compared with normal skin tissue (1132.56±882.7 vs. 107.62±183.62, 1118.15±1109.49 vs. 9.5±5, 2603.87±2385.26 vs. 5.29±3, 957.95±627.14 vs. 400.42±967.66, 1149.13±832.18 vs. 19.41±35.62, respectively; p<0.05). In summary, this study highlights the potential prognostic value of MAGE-A3 in clinical outcomes of cutaneous squamous cell carcinoma patients.

Introduction: Hypokalemia in primary aldosteronism (PA) patients correlates with higher levels of cardiovascular events and altered left ventricular geometry. However, the influence of aldosterone on microvascular endothelial function and the effect of hypokalemia on the vascular structure still remain unclear. Objectives: We investigated the peripheral arterial functions, including the endothelial function of microvasculature and arterial stiffness in PA and essential hypertension (EH) patients, and the correlation between hypokalemia and peripheral arterial function among PA patients. Methods: Twenty patients diagnosed as EH and 37 patients with PA were enrolled in this study. Reactive hyperemia index (RHI) and the augmentation index (AI) were obtained by non-invasive peripheral arterial tonometry. Results: Twenty EH patients and a total of 37 PA patients, including 21 patients with normokalemia and 16 patients with hypokalemia, were enrolled and divided into groups 1, 2 and 3 respectively. PA patients had significantly higher AI ( p =0.024) but not RHI than EH patients. RHI showed no difference between groups 1, 2 and 3. Group 3 had higher AI than either group 1 or group 2. In the whole study population, serum potassium level, after multivariate regression analysis testing, was the only factor associated with AI (ß= −0.102; p =0.002). In PA patients, serum potassium level was the only significant factor correlated with AI. (r= −0.458; p =0.004) Conclusions: PA patients had higher arterial stiffness but comparable microvascular endothelial function to EH patients. Hypokalemia correlated with arterial stiffness but not microvascular endothelial function in PA patients.

Background Primary aldosteronism (PA) is one of the major etiologies for secondary hypertension featuring more prominent left ventricular hypertrophy. The purpose of the study was to investigate the predictive factors of left ventricular mass index (LVMI) regression in patients with PA after adrenalectomy. Methods We prospectively analyzed 30 patients with aldosterone-producing adenoma (APA) who received adrenalectomy from October 2006 to September 2008. Echocardiography was performed preoperation and 1 year after operation. Results Thirty patients with aldosterone-producing adenoma undergoing adrenalectomy were enrolled. In a 1-year follow-up, LVMI decreased significantly by an average of 18.6%. Net LVMI decrease (ΔLVMI) was associated with preoperative LVMI, preoperative serum potassium level, baseline systolic blood pressure (SBP), baseline diastolic blood pressure, net SBP decrease (ΔSBP), net diastolic blood pressure decrease, preoperative/postoperative change of log-transformed plasma aldosterone concentration, preoperative/postoperative change of log-transformed plasma renin activity, and preoperative/postoperative change of serum potassium level (Δserum potassium level). In a multiple regression analysis, preoperative LVMI (β = −0.287, P = 0.049), ΔSBP (β = 0.518, P = 0.01), and Δserum potassium level (β = −20.471, P = 0.014) were significantly correlated with ΔLVMI. Conclusions The LVMI in patients with PA regressed significantly after adrenalectomy. Preoperative LVMI, ΔSBP, and Δserum potassium levels are independent factors associated with the degree of LVMI regression.

BACKGROUND:Galectin-3 (Gal-3) shows the ability of survival prediction in heart failure (HF) patients. However, Gal-3 is strongly associated with serum markers of cardiac extracellular matrix (ECM) turnover. The aim of this study is to compare the impact of Gal-3 and serum markers of cardiac ECM turnover on prognostic prediction of chronic systolic HF patients.METHODS:Serum Gal-3, brain natriuretic peptide (BNP), extracellular matrix including type I and III aminoterminal propeptide of procollagen (PINP and PIIINP), matrix metalloproteinase-2, 9 (MMP-2, 9), and tissue inhibitor of metalloproteinase-1 (TIMP-1) were analyzed. Cox regression analysis was used for survival analysis.RESULTS:A total of 105 (81 male) patients were enrolled. During 980±346 days follow-up, 17 patients died and 36 episodes of HF admission happened. Mortality of these patients was significantly associated with the log PIIINP (β= 15.380; P=0.042), log TIMP-1(β= 44.530; P=0.003), log MMP-2 (β= 554.336; P<0.001), log BNP (β= 28.273; P=0.034). Log Gal-3 (β= 7.484; P=0.066) is borderline associated with mortality. Mortality or first HF admission of these patients was significantly associated with the log TIMP-1(β= 16.496; P=0.006), log MMP-2 (β= 221.864; P<0.001), log BNP (β= 5.999; P=0.034). Log Gal-3 (β= 4.486; P=0.095) only showed borderline significance. In several models adjusting clinical parameters, log MMP-2 was significantly associated with clinical outcome. In contrast, log Gal-3 was not.CONCLUSION:The prognostic strength of MMP-2 to clinical outcome prediction in HF patients is stronger than Gal-3.

Low-power laser–activated endogenous latent transforming growth factor–β1 (LTGF-β1) directs resident dental stem cell differentiation to promote dentin regeneration.

Eur J Clin Invest 2012; 42 (10): 1079–1086AbstractObjectives The goal of this study was designed to assess prognostic values of simultaneous measurement of adipocytokines in systolic heart failure (HF) patients.Methods Patients with HF manifestations and left ventricular ejection fraction (LVEF) ≤ 50% were selected in this study. Gender, age, medications and serum biochemical data were recorded upon admissions. Adipocytokines including adiponectin, leptin, resistin, visfatin and retinol binding protein‐4 were measured.Results A total of 108 (83 males and 25 females) patients were enroled. The age was 62 ± 15 years and mean LVEF was 35%. Twenty patients died during 776 ± 323 days follow‐up. In univariate analysis, mortality was found to be associated with the log‐transformed values of serum resistin (β = 5·616, P = 0·04), log‐transformed values of serum adiponectin (β = 4·377, P = 0·038), age (β = 1·071, P < 0·001), NTHA functional status (β = 3·752, P = 0·001) and body mass index (β = 0·858, P = 0·012). Patients with higher level of serum resistin were associated with higher mortality (P = 0·012). In multivariate analysis, mortality is associated with log‐transformed values of serum resistin (β = 3·666, P = 0·045), age (β = 1·044, P = 0·017) and NTHA functional status (β = 2·541, P = 0·025).Conclusions Serum resistin level was associated with higher mortality in systolic HF patients even after adjusting clinical parameters. Resistin may be an informative risk marker for systolic HF patients.

OBJECTIVEThe purpose of this study was to investigate the effect of 810-nm low level laser therapy (LLLT) on dendritic cells (DC) in vitro.BACKGROUND DATALLLT can enhance wound healing and increase cell proliferation and survival, and is used to treat inflammatory conditions. However there are reports that LLLT can stimulate leukocytes and could therefore be pro-inflammatory. Recently, DC have been found to play an important role in inflammation and immune response.METHODSMurine bone-marrow-derived DC were isolated, stimulated with lipopolysaccharide (LPS) or CpG oligodeoxynucleotide and treated with 810-nm laser, using fluences of 0.3, 3, and 30  J/cm(2) delivered at irradiances of 1, 10, and 100  mW/cm(2) respectively. Confocal microscopy, flow cytometry for DC markers, viability using propidium iodide, enzyme-linked immunosorbent assays (ELISA) for secreted interleukin-12 (IL-12), and bioluminescence measurements in cells transduced with a reporter for toll-like receptor (TLR)-9/nuclear factor kappa B (NF-κB) activation, were performed.RESULTSLLLT changed the morphology of LPS-stimulated DC, increased their viability, and altered the balance of DC activation markers (major histocompatibility complex [MHC] class 2 up and CD86 down). LLLT reduced IL-12 secretion from DC stimulated by either LPS or CpG. LLLT reduced NF-κB activation in reporter cells stimulated with CpG. There was no obvious light dose response observed.CONCLUSIONSTaken together, these data suggest that 810-nm LLLT has an anti-inflammatory effect on activated DC, possibly mediated by cyclic adenosine monophosphate (cAMP) and reduced NF-κB signaling.

BACKGROUND:Despite over forty years of investigation on low-level light therapy (LLLT), the fundamental mechanisms underlying photobiomodulation at a cellular level remain unclear.METHODOLOGY/PRINCIPAL FINDINGS:In this study, we isolated murine embryonic fibroblasts (MEF) from transgenic NF-kB luciferase reporter mice and studied their response to 810 nm laser radiation. Significant activation of NF-kB was observed at fluences higher than 0.003 J/cm(2) and was confirmed by Western blot analysis. NF-kB was activated earlier (1 hour) by LLLT compared to conventional lipopolysaccharide treatment. We also observed that LLLT induced intracellular reactive oxygen species (ROS) production similar to mitochondrial inhibitors, such as antimycin A, rotenone and paraquat. Furthermore, we observed similar NF-kB activation with these mitochondrial inhibitors. These results, together with inhibition of laser induced NF-kB activation by antioxidants, suggests that ROS play an important role in the laser induced NF-kB signaling pathways. However, LLLT, unlike mitochondrial inhibitors, induced increased cellular ATP levels, which indicates that LLLT also upregulates mitochondrial respiration.CONCLUSION:We conclude that LLLT not only enhances mitochondrial respiration, but also activates the redox-sensitive NFkB signaling via generation of ROS. Expression of anti-apoptosis and pro-survival genes responsive to NFkB could explain many clinical effects of LLLT.

Background Primary aldosteronism (PA) is the most frequent cause of secondary hypertension, and is associated with more prominent vascular stiffness and atherosclerosis. Current studies have implied that access aldosterone induced by PA could lead to increase vascular stiffness and carotid intima media thickness (CIMT). Though aldosterone producing adenoma (APA) is the most common subtype that can be cured by adrenalectomy, the post-surgical vascular changes remains unclear. Method We prospectively analyzed 20 patients with APA that received adrenalectomy from October 2006 to Dec 2008and 21 patients with essential hypertension (EH) were enrolled as the control group. CIMT measurement by B-mode ultrasound of the distal right common carotid arteries and pulse wave velocity (PWV) measurement including brachial-ankle PWV (baPWV) and heart-ankle PWV (haPWV) were performed in both groups. The follow-up measurements were performed one year after adrenalectomy in APA group. Results APA patients had significantly higher diastolic blood pressure, plasma aldosterone concentration (PAC) and aldosterone-renin ratio (ARR), but lower serum potassium level and plasma renin activity (PRA) than EH patients.APA patients had significantly higher CIMT, greater baPWV and haPWVthan EH patients (both p < 0.05). One year afteradrenalectomy, CIMT and PWV level decreased significantly (0.64 ± 0.13 to 0.59 ± 0.14 mm for CIMT, 1589 ± 296 to 1463 ± 188 cm/s for baPWV, 1095 ± 150 to 1017 ± 109 cm/s for haPWV, all p < 0.05). Conclusion The results of this study showed that APA patients have higher degree of early atherosclerosis and vascular stiffness. More importantly, this study is the first human trial demonstrating that adrenalectomy not only corrects the high blood pressure and biochemical data but also reverse adverse vascular change in APA patients.

Context: Primary aldosteronism (PA) is the most frequent cause of secondary hypertension, and is associated with more prominent vascular stiffness and atherosclerosis. However, the effect of adrenalectomy on reversibility of vascular damage is unclear. Objective: Our objective was to investigate the vascular changes and possibility of reversibility after adrenalectomy in PA patients.Methods: We prospectively analyzed 20 patients with aldosterone producing adenoma (APA) that received adrenalectomy from October 2006 to December 2008 and 21 patients with essential hypertension (EH) were enrolled as the control group. Carotid intima media thickness (CIMT) measurement by B-mode ultrasound of the right common carotid arteries and pulse wave velocity (PWV) measurement including brachial-ankle PWV (baPWV) and heart-ankle PWV (haPWV) were performed in both groups. The follow-up measurements were performed one-year after adrenalectomy in APA group.Results: APA patients had significantly higher diastolic blood pressure, plasma aldosterone concentration (PAC) and aldosterone-renin ratio (ARR), but lower serum potassium level and plasma renin activity (PRA) than EH patients. APA patients had significantly higher CIMT (0.64 +/- 0.13 vs. 0.53 +/- 0.10 mm, p = 0.006), higher baPWV (1589 +/- 296 vs. 1405 +/- 187 cm/s, p = 0.024) and haPWV (1095 +/- 150 vs. 987 +/- 114 cm/s, p = 0.013) comparing with EH patients. One-year after adrenalectomy, CIMT reduced significantly from 0.64 +/- 0.13 mm to 0.59 +/- 0.14 mm (p = 0.014), and baPWV and haPWV also showed significant reduction (baPWV, 1589 +/- 296 to 1463 +/- 188 cm/s, p = 0.035; haPWV, 1095 +/- 150 to 1017 +/- 109 cm/s, p = 0.019).Conclusion: APA patients have higher degree of early atherosclerosis and vascular stiffness. Adrenalectomy not only corrects the high blood pressure and biochemical parameters but also reverse adverse vascular change in APA patients. (C) 2012 Elsevier Ireland Ltd. All rights reserved.

Discoveries are rapidly being made in multiple laboratories that shed "light" on the fundamental molecular and cellular mechanisms underlying the use of low level light therapy (LLLT) in vitro, in animal models and in clinical practice. Increases in cellular levels of respiration, in cytochrome c oxidase activity, in ATP levels and in cyclic AMP have been found. Increased expression of reactive oxygen species and release of nitric oxide have also been shown. In order for these molecular changes to have a major effect on cell behavior, it is likely that various transcription factors will be activated, possibly via different signal transduction pathways. In this report we compare and contrast the effects of LLLT in vitro on murine embryonic fibroblasts, primary cortical neurons, cardiomyocytes and bone-marrow derived dendritic cells. We also examined two human cell lines, HeLa cancer cells and HaCaT keratinocytes. The effects of 810-nm near-infra-red light delivered at low and high fluences were addressed. Reactive oxygen species generation, transcription factor activation and ATP increases are reported. The data has led to the hypothesis that cells with a high level of mitochondrial activity (mitochondrial membrane potential) have a higher response to light than cells with low mitochondrial activity.

OBJECTIVE The purpose of this case report was to evaluate the efficacy of photodynamic therapy (PDT) for a patient with primary cutaneous adenoid cystic carcinoma (PCACC). MATERIALS AND METHODS A 53-year-old female patient presented with a lesion at the pre-auricular area. After performing a local resection along a margin of 0.5 cm beyond the surrounding tissue, PCACC was clearly revealed based on the histological features and the exclusion of differentiation of other metastasis in the skin. The patient was then treated repetitively with 5-aminolevulinic acid (ALA)-PDT. After the final PDT treatment, the control pathological biopsy at the same location demonstrated no tumor cell in the specimen. RESULTS The 40-month follow-up diagnosis showed no evidence of recurrence or metastasis observed on the treated area. CONCLUSION ALA-PDT was a safe and effective therapy for obtaining good cosmetic results and reducing recurrence in this case report.

There are many reports showing that low-level light/laser therapy (LLLT) can enhance wound healing, upregulate cell proliferation and has anti-apoptotic effects by activating intracellular protective genes. In the field of immune response study, it is not known with any certainty whether light/laser is pro-inflammatory or anti-inflammatory. Increasingly in recent times dendritic cells have been found to play an important role in inflammation and the immunological response. In this study, we try to look at the impact of low level near infrared light (810-nm) on murine bone-marrow derived dendritic cells. Changes in surface markers, including MHC II, CD80 and CD11c and the secretion of interleukins induced by light may provide additional evidence to reveal the mystery of how light affects the maturation of dendritic cells as well how these light-induced mature dendritic cells would affect the activation of adaptive immune response.

The use of low levels of visible or near infrared light for reducing pain, inflammation and edema, promoting healing of wounds, deeper tissues and nerves, and preventing tissue damage and death has been known for almost forty years since the invention of lasers. Originally thought to be a peculiar property of laser light (soft or cold lasers), the subject has now broadened to include photobiomodulation and photobiostimulation using noncoherent light. Despite many reports of positive findings from experiments conducted in vitro, in animal models and in randomized controlled clinical trials, low level light therapy (LLLT) remains controversial. This controversy is likely due to two main reasons; first, the biochemical mechanisms underlying the positive effects are incompletely understood, and second, the complexity of rationally choosing among a large number of illumination parameters such as wavelength, fluence, power density, pulse structure, and treatment timing has led to the publication of a number of negative studies as well as many positive ones. In particular, a biphasic dose response has been frequently observed where low levels of light have a much better effect than higher levels. This chapter will cover some of the proposed cellular chromophores responsible for the effect of visible light on mammalian cells, including cytochrome c oxidase (with absorption peaks in the near infrared region of the spectrum) and photoactive porphyrins and flavins. Mitochondria are thought to be a likely site for the initial effects of light and the involvement of nitric oxide had been postulated. Activation of the respiratory chain leads to increased ATP production, modulation of reactive oxygen species, and activation of transcription factors. These effects in turn lead to increased cell proliferation and migration (particularly by fibroblasts), modulation in levels of cytokines, growth factors and inflammatory mediators, prevention of cell death by anti-apoptotic signaling, and increased tissue oxygenation. The results of these biochemical and cellular changes in animals and patients include such benefits as increased healing in chronic wounds, improvements in sports injuries and carpal tunnel syndrome, pain reduction in arthritis and neuropathies, and amelioration of damage after heart attacks, stroke, nerve injury, and retinal toxicity.

BACKGROUND/PURPOSE:Based on the observation that increasing skin temperature could improve 5-aminolevulinic acid (ALA) penetration and accumulation of protoporphyrin IX (PpIX) in the ALA-based photodynamic therapy (PDT), this study was designed to investigate how temperature change varied the therapeutic effect of ALA-based PDT in vitro.METHODS:HaCat cells were cultured with or without ALA at various temperatures. ALA uptake and PpIX accumulation were analyzed before laser irradiation as the baseline. After irradiation, cell death and cytokine secretions in the media, including interleukin (IL)-1alpha, tumor necrosis factor (TNF)-alpha and basic fibroblast growth factor (bFGF) were assayed, and the morphological changes were recorded.RESULTS:With increasing temperature, the amount of ALA uptake, intracellular PpIX concentration and cell death increased in both the PDT and the non-PDT groups. Secretions of IL-1alpha, TNF-alpha and bFGF also increased and reached a peak around 44-47 degrees C and then declined at a higher temperature. This biphasic response might be due to protein thermolysis that occurs when cells reach beyond thermal tolerance.CONCLUSIONS:Elevating temperature could augment photodynamic reactions to a certain extent, but adverse effects occurred when cells were overheated.

Low level laser (or light) therapy (LLLT) has been clinically applied for many indications in medicine that require the following processes: protection from cell and tissue death, stimulation of healing and repair of injuries, and reduction of pain, swelling and inflammation. One area that is attracting growing interest is the use of transcranial LLLT to treat stroke and traumatic brain injury (TBI). The fact that near-infrared light can penetrate into the brain would allow non-invasive treatment to be carried out with a low likelihood of treatment-related adverse events. LLLT may have beneficial effects in the acute treatment of brain damage injury by increasing respiration in the mitochondria, causing activation of transcription factors, reducing key inflammatory mediators, and inhibiting apoptosis. We tested LLLT in a mouse model of TBI produced by a controlled weight drop onto the skull. Mice received a single treatment with 660-nm, 810-nm or 980-nm laser (36 J/cm2) four hours post-injury and were followed up by neurological performance testing for 4 weeks. Mice with moderate to severe TBI treated with 660- nm and 810-nm laser had a significant improvement in neurological score over the course of the follow-up and histological examination of the brains at sacrifice revealed less lesion area compared to untreated controls. Further studies are underway.

The use of low levels of visible or near infrared light for reducing pain, inflammation and edema, promoting healing of wounds, deeper tissues and nerves, and preventing cell death and tissue damage has been known for over forty years since the invention of lasers. Despite many reports of positive findings from experiments conducted in vitro, in animal models and in randomized controlled clinical trials, LLLT remains controversial in mainstream medicine. The biochemical mechanisms underlying the positive effects are incompletely understood, and the complexity of rationally choosing amongst a large number of illumination parameters such as wavelength, fluence, power density, pulse structure and treatment timing has led to the publication of a number of negative studies as well as many positive ones. A biphasic dose response has been frequently observed where low levels of light have a much better effect on stimulating and repairing tissues than higher levels of light. The so-called Arndt-Schulz curve is frequently used to describe this biphasic dose response. This review will cover the molecular and cellular mechanisms in LLLT, and describe some of our recent results in vitro and in vivo that provide scientific explanations for this biphasic dose response.

This review will focus on the role of reactive oxygen species in the cellular and tissue effects of low level light therapy (LLLT). Coincidentally with the increase in electron transport and in ATP, there has also been observed by intracellular fluorescent probes and electron spin resonance an increase in intracellular reactive oxygen species (ROS) such as superoxide, hydrogen peroxide, singlet oxygen and hydroxyl radical. ROS scavengers, antioxidants and ROS quenchers block many LLLT processes. It has been proposed that light between 400-500- nm may produce ROS by a photosensitization process involving flavins, while longer wavelengths may directly produce ROS from the mitochondria. Several redox-sensitive transcription factors are known such as NF-kB and AP1, that are able to initiate transcription of genes involved in protective responses to oxidative stress. It may be the case that LLLT can be pro-oxidant in the short-term, but anti-oxidant in the long-term.