Use of near-infrared Raman spectroscopy for identification of atherosclerotic plaques in the carotid artery.

OBJECTIVES: The aim of this work was to identify the presence of atherosclerotic plaque in the human carotid artery using near infrared Raman spectroscopy. BACKGROUND DATA: Atherosclerosis is the most common and serious pathology of the cardiovascular system. Raman spectroscopy is an analytical tool that can be used to gather information about both the morphology and chemical composition of tissues. METHODS: A Ti:sapphire laser operating at the near-infrared wavelength of 830 nm pumped by an argon laser was used for excitation of the samples, and the Raman scattering was detected by an optical spectrometer with a liquid-nitrogen-cooled CCD detector. Carotid artery samples were classified into five groups: normal, intimal thickening, fatty plaque, fibrous-fatty plaque, and fibrous-calcified plaque. RESULTS: It was observed that the Raman spectrum of atheromatous plaque was different that that of normal tissue. The spectra of atheromatous plaques had bands due to the presence of cholesterol and its esters, with major bands at 1439 and 1663 cm(1), respectively. In normal tissues a peak related to C-H bending appears at 1451 cm(1). Calcified atheromatous plaques had primary bands at 961 and 1071 cm(1), which were due to the presence of phosphate and carbonate in the accumulated calcium. Peaks were seen at 1451 and 1655 cm(1) in the non-atherosclerotic tissue, which were shifted to 1439 and 1663 cm(1) in the atherosclerotic plaque. CONCLUSIONS: Our results indicate that this technique could be used to detect the presence of atherosclerotic plaques in carotid arterial tissue.

The impact of photodynamic therapy on the viability of Streptococcus mutans in a planktonic culture.

OBJECTIVE: This study investigated the effect of photodynamic therapy (PDT) with toluidine blue O (TBO) and a light-emitting diode (LED) on the viability of Streptococcus mutans cells in a planktonic culture. BACKGROUND DATA: Growth of Streptococcus mutans is the first step in the development of tooth decay. The use of light and dyes promotes cellular death in a noninvasive way, reducing treatment time. METHODS: The LED used in this study had output power of 116 mW, its energy was 21 J, and the fluency was 2.18 J/cm(2). Samples were prepared and divided into five groups: (1) control group (-); (2) control group (+); (3) TBO; (4) LED; and (5) LED + TBO. RESULTS: One hundred percent of the bacteria were killed following irradiation with LED and TBO. The biofilm that formed on the glass surfaces was analyzed by SEM and colony count. CONCLUSIONS: It was demonstrated that PDT was efficient at killing microorganisms and preventing the formation of biofilms.

The effect of the association of NIR laser therapy BMPs, and guided bone regeneration on tibial fractures treated with wire osteosynthesis: Raman spectroscopy study.

Bone fractures are lesions of different etiology; may be associated or not to bone losses; and have different options for treatment, such as the use of biomaterials, guided bone regeneration, techniques considered effective on improving bone repair. Laser therapy has also been shown to improve bone healing on several models. The association of these three techniques has been well documented by our group using different models. This study aimed to assess, through Raman spectroscopy, the incorporation of calcium hydroxyapatite (CHA approximately 958 cm(-1)) on the repair of complete tibial fractures in rabbits treated with wire osteosynthesis (WO); treated or not with laser therapy; and associated or not with the use of BMPs and/or Guided Bone Regeneration. Complete tibial fractures were created in 12 animals that were divided into four groups: WO; WO+BMPs; WO+laser therapy; and WO+BMPs+laser therapy. Irradiation started immediately after surgery; was repeated at every other day during 2 weeks; and was carried out with lambda 790 nm laser light (4 J/cm(2) per point, 40 mW, phi approximately 0.5 cm(2), 16J per session). Animal death occurred after 30 days. Raman spectroscopy was performed at both the surface and the depth of the fracture site. Statistical analysis showed significant difference on the concentrations of CHA between surface and depth. The analysis in each of the areas showed at the depth of the fracture significant differences between all treatment groups (p<0.0001). Significant differences were also seen between WO+BMPs+laser therapy and WO (p<0.001) and WO+laser therapy (p<0.001). At the surface, significant difference was seen only between the treatment groups and the non-fractured subjects (p=0.0001). However, no significant difference was seen between the treatment groups (p=0.14). It is concluded that the use of NIR laser therapy associated to BMPs and GBR was effective in improving bone healing on the fractured bones as a result of the increasing deposition of CHA measured by Raman spectroscopy.

Effect of low-level laser therapy on hemorrhagic lesions induced by immune complex in rat lungs.

OBJECTIVE: The aim of this study was to investigate if low-level laser therapy (LLLT) can modulate formation of hemorrhagic lesions induced by immune complex. BACKGROUND DATA: There is a lack of information on LLLT effects in hemorrhagic injuries of high perfusion organs, and the relative efficacy of LLLT compared to anti-inflammatory drugs. METHODS: A controlled animal study was undertaken with 49 male Wistar rats randomly divided into seven groups. Bovine serum albumin (BSA) i.v. was injected through the trachea to induce an immune complex lung injury. The study compared the effect of irradiation by a 650-nm Ga-Al-As laser with LLLT doses of 2.6 Joules/cm(2) to celecoxib, dexamethasone, and control groups for hemorrhagic index (HI) and myeloperoxide activity (MPO) at 24 h after injury. RESULTS: The HI for the control group was 4.0 (95% CI, 3.7-4.3). Celecoxib, LLLT, and dexamethasone all induced significantly (p < 0.01) lower HI than control animals at 2.5 (95% CI, 1.9-3.1), 1.8 (95% CI, 1.2-2.4), and 1.5 (95% CI, 0.9-2.1), respectively, for all comparisons to control. Dexamethasone, but not celecoxib, induced a slightly, but significantly lower HI than LLLT (p = 0.04). MPO activity was significantly decreased in groups receiving celecoxib at 0.87 (95% CI, 0.63-1.11), dexamethasone at 0.50 (95% CI, 0.24-0.76), and LLLT at 0.7 (95% CI, 0.44-0.96) when compared to the control group, at 1.6 (95% CI, 1.34-1.96; p < 0.01), but there were no significant differences between any of the active treatments. CONCLUSION: LLLT at a dose of 2.6 Joules/cm(2) induces a reduction of HI levels and MPO activity in hemorrhagic injury that is not significantly different from celecoxib. Dexamethasone is slightly more effective than LLLT in reducing HI, but not MPO activity.

Low-level laser therapy can reduce lipopolysaccharide-induced contractile force dysfunction and TNF-alpha levels in rat diaphragm muscle.

Our objective was to investigate if low-level laser therapy (LLLT) could improve respiratory function and inhibit tumor necrosis factor (TNF-alpha) release into the diaphragm muscle of rats after an intravenous injection of lipopolysaccharide (LPS) (5 mg/kg). We randomly divided Wistar rats in a control group without LPS injection, and LPS groups receiving either (a) no therapy, (b) four sessions in 24 h with diode Ga-AsI-Al laser of 650 nm and a total dose of 5.2 J/cm2, or (c) an intravenous injection (1.25 mg/kg) of the TNF-alpha inhibitor chlorpromazine (CPZ). LPS injection reduced maximal force by electrical stimulation of diaphragm muscle from 24.15+/-0.87 N in controls, but the addition of LLLT partly inhibited this reduction (LPS only: 15.01+/-1.1 N vs LPS+LLLT: 18.84+/-0.73 N, P<0.05). In addition, this dose of LLLT and CPZ significantly (P<0.05 and P<0.01, respectively) reduced TNF-alpha concentrations in diaphragm muscle when compared to the untreated control group.

Low level laser therapy partially restores trachea muscle relaxation response in rats with tumor necrosis factor alpha-mediated smooth airway muscle dysfunction.

BACKGROUND AND OBJECTIVE: It is unknown if the decreased ability to relax airway smooth muscles in asthma and other inflammatory airways disorders can be influenced by low level laser therapy (LLLT) irradiation. To investigate if LLLT could reduce impairment in inflamed trachea smooth muscles (TSM) in rats. STUDY DESIGN/MATERIALS AND METHODS: Controlled rat study where trachea was dissected and mounted in an organ bath apparatus with or without a TNF-alpha solution. RESULTS: Low level laser therapy administered perpendicularly to a point in the middle of the dissected trachea with a wavelength of 655 nm and a dose of 2.6 J/cm(2), partially restored TSM relaxation response to isoproterenol. Tension reduction was 47.0 % (+/-2.85) in the laser-irradiated group compared to 22.0% (+/-2.21) in the control group (P < 0.01). Accumulation of cAMP was almost normalized after LLLT at 22.3 pmol/mg (+/-2.1) compared to 17.6 pmol/mg (+/-2.1) in the non-irradiated control group (P < 0.01). CONCLUSION: Low level laser therapy partially restores the normal relaxation response in inflamed TSM and normalizes accumulation of cAMP in the presence of isoproterenol.

Low-level laser therapy induces dose-dependent reduction of TNFalpha levels in acute inflammation.

OBJECTIVE: The aim of this study was to investigate if low-level laser therapy (LLLT) can modulate acute inflammation and tumor necrosis factor (TNFalpha) levels. BACKGROUND DATA: Drug therapy with TNFalpha-inhibitors has become standard treatment for rheumatoid arthritis, but it is unknown if LLLT can reduce or modulate TNFalpha levels in inflammatory disorders. METHODS: Two controlled animal studies were undertaken, with 35 male Wistar rats randomly divided into five groups each. Rabbit antiserum to ovalbumin was instilled intrabronchially in one of the lobes, followed by the intravenous injection of 10 mg of ovalbumin in 0.5 mL to induce acute lung injury. The first study served to define the time profile of TNFalpha activity for the first 4 h, while the second study compared three different LLLT doses to a control group and a chlorpromazine group at a timepoint where TNFalpha activity was increased. The rats in LLLT groups were irradiated within 5 min at the site of injury by a 650-nm Ga-Al-As laser. RESULTS: There was a time-lag before TNFalpha activity increased after BSA injection. TNFalpha levels increased from < or =6.9 (95% confidence interval [CI], 5.6-8.2) units/mL in the first 3 h to 62.1 (95% CI, 60.8-63.4) units/mL (p < 0.001) at 4 h. An LLLT dose of 0.11 Joules administered with a power density of 31.3 mW/cm(2) in 42 sec significantly reduced TNFalpha level to 50.2 (95% CI, 49.4-51.0), p < 0.01 units/mL versus control. Chlorpromazine reduced TNFalpha level to 45.3 (95% CI, 44.0-46.6) units/mL, p < 0.001 versus control. CONCLUSION: LLLT can reduce TNFalpha expression after acute immunocomplex lung injury in rats, but LLLT dose appears to be critical for reducing TNFalpha release.

Effect of LLLT Ga-Al-As (685 nm) on LPS-induced inflammation of the airway and lung in the rat.

The purpose of this study was to investigate the effect of low level laser therapy (LLLT) on male Wistar rat trachea hyperreactivity (RTHR), bronchoalveolar lavage (BAL) and lung neutrophils influx after Gram-negative bacterial lipopolyssacharide (LPS) intravenous injection. The RTHR, BAL and lung neutrophils influx were measured over different intervals of time (90 min, 6 h, 24 h and 48 h). The energy density (ED) that produced an anti-inflammatory effect was 2.5 J/cm(2), reducing the maximal contractile response and the sensibility of trachea rings to methacholine after LPS. The same ED produced an anti-inflammatory effect on BAL and lung neutrophils influx. The Celecoxib COX-2 inhibitor reduced RTHR and the number of cells in BAL and lung neutrophils influx of rats treated with LPS. Celecoxib and LLLT reduced the PGE(2) and TXA(2) levels in the BAL of LPS-treated rats. Our results demonstrate that LLLT produced anti-inflammatory effects on RTHR, BAL and lung neutrophils influx in association with inhibition of COX-2-derived metabolites.

Side-viewing fiberoptic catheter for biospectroscopy applications.

Utilization of fiberoptic catheters can turn the Raman and fluorescence spectroscopy systems into powerful bio-medical diagnostic probes. An in vivo bio-chemical diagnosis of some important organs like the esophagus, intestine, lung branches, artery, etc., can be possible by developing fiber-probes with good signal collection capabilities, a good flexibility to scan different spatial regions of the sample and less background signals generated in the probes themselves. An in vivo diagnosis of endoluminal inner walls utilizing front-viewing catheters (FVC) is very difficult because the internal diameter of these organs do not allow (excitation and collection) flexibility to access the different spatial regions of the sample. In this work we have developed, different side-viewing catheter (SVC) probes with a very small distal tip (semi sphere, phi approximately 1.5 mm) and micro mirrors allow beam steering of the excitation and collected radiation at a 90 degree angle. Preliminary results of spectroscopic applications have been presented. Reflectance, fluorescence and Raman scattering measurements have been used to compare the efficiency of SVC with traditional FVC probes. The results demonstrate that the SVC probes not only exhibit more flexibility but also similar spectral characteristics and signal collection efficiencies in comparison with conventional FVC probes.

Mitochondrial membrane potential after low-power laser irradiation.

We used the lipophilic cationic fluorescent dye 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethyl-benzimidazol-carbocyanine iodide (JC-1) to determine mitochondrial membrane potential (mdeltapsi) in Hep-2 cells after irradiation with low-power laser (lambda=635 nm). Through this methodology it was possible to analyze the variation on mitochondrial number and mdeltapsi, in cells irradiated for 100, 150 and 200 s with energy density of 100 mJ/cm(2). Our results show that JC-1 dye allows the identification of populations with different mitochondria morphology as well as the functionality of this organelle in the cells incubated for 1, 6 and 24 h, after irradiation with low-power laser.

Analysis of mitochondria, endoplasmic reticulum and actin filaments after PDT with AlPcS(4).

Photodynamic therapy (PDT) is a therapeutic modality for the treatment of tumors. This technique uses a visible light to activate a photosensitizer compounds, leading to a photo-oxidation process of biological tissue that can induce apoptosis or necrosis both in vivo and in vitro. However many of the cytotoxic effects remain an open question to be investigated. The cytotoxicity to specific cellular targets of classical photosensitizers used in the PDT in vitro has been analyzed in this work. The photosensitizing effects of Chloroaluminum Phthalocyanine Tetrasulfonate (AlPcS(4)) were studied on the mitochondria, cytoskeleton and endoplasmic reticulum of HeLa cells. The cells were irradiated with a diode laser (working at 670 nm; energy density of 4.5 J/cm(2 )and power density of 45 mW/cm(2)). The spectrofluorimetric analysis of the mitochondria showed changes in membrane potential. Cytoskeleton and endoplasmic reticulum showed basic alterations in distribution after PDT treatment, as an indicator of cellular death process.

Effect of GaInAIP (685 nm) radiation on lipopolyssacharide induced rat trachea hyperreactivity

O objetivo do presente estudo foi investigar o efeito do laser de baixa potência (LLLT) na hiper-reatividade da traquéia de ratos Wistar macho(RTHR) depois da administração de lipopolissacarídeo (LPS)

Using the laser-induced fluorescence spectroscopy in the differentiation between normal and neoplastichuman breast tissue.

This article reports results of the in vitro study for potential evaluation of the laser-induced fluorescence spectroscopy in the differentiation between normal and neoplastic human breast tissue. A coumarine dye laser pumped by nitrogen laser generated an excitation light centered at 458 nm. In order to collect the fluorescence signal was used an optical fiber catheter coupled to a spectrometer and CCD detector. Fluorescence spectra were recorded from normal and neoplastic (benign and malignant) human breast tissue, adding up 94 different areas. The discrimination between normal and neoplasm groups reach a sensitivity and specificity of 100%.

Espectrofluorímetro de multi-excitaçäo portátil para a rápida deteçäo de displasias / Spectrofluorimeter of portable multi-excitation for rapid detection of dysplasia

Neste trabalho é apresentado o projeto e desenvolvimento de um espectrofluorímetro para a obtenção de espectros de fluorescência e refletância difusa de tecidos biológicos em um tempo inferior a 1s. Para acessar a região da diagnose, o sistema utiliza um catéter a fibra óptica para excitação do tecido e captação da fluorescência emitida. O sistema é desenvolvido para aplicação em procedimentos clínicos in vivo, onde o tempo de realização do experimento é objetivo de extrema importância

Abstract- This work presents the project and development of a spectrofluorimeter in order to obtain the fluorescence spectra and diffuse reflectance from biological tissues with period of time below 1 s. The diagnose region is reached with an optical fiber catheter for tissue excitation and collection of the emitted fluorescence. This system is developed for in vivo clinicai applications, where duration of the experimental procedure is a very important parameter