Discrimination of non-melanoma skin lesions from non-tumor human skin tissues in vivo using Raman spectroscopy and multivariate statistics.

BACKGROUND AND OBJECTIVE: Raman spectroscopy was used to discriminate human non-melanoma skin lesions from non-tumor tissues in vivo. This work proposed the discrimination between non-melanoma (basal cell carcinoma, BCC; squamous cell carcinoma, SCC) and pre-cancerous lesions (actinic keratosis, AK) from benign lesions and normal (non-tumor group, NT) tissues, using near-infrared Raman spectroscopy with a Raman probe. MATERIALS AND METHODS: Prior to surgery, the spectra of suspicious lesions were obtained in situ. The spectra of adjacent, clinically normal skin were also obtained. Lesions were resectioned and submitted for histopathology. The Raman spectra were measured using a Raman spectrometer (830 nm). Two types of discrimination models were developed to distinguish the different histopathological groups. The principal components analysis discriminant analysis (PCA/DA) and the partial least squares discriminant analysis (PLS/DA) were based on Euclidean, quadratic and Mahalanobis distances. RESULTS: PCA and PLS spectral vectors showed spectral features of skin constituents, such as lipids (between 1,250 cm(-1) and 1,300 cm(-1) and at 1,450 cm(-1)) and proteins (between 870 cm(-1) and 940 cm(-1), 1,240 cm(-1) and 1,271 cm(-1), and at 1,000 cm(-1) and 1,450 cm(-1)). Despite the small spectral differences between malignant lesions and benign tissues, the algorithms discriminated the spectra of non-melanoma skin and pre-cancerous lesions from benign and normal tissues, with an overall accuracy of 82.8% and 91.9%, respectively. CONCLUSION: PCA and PLS could discriminate Raman spectra of skin tissues, opening the way for an in vivo optical diagnosis.

Identification of different forms of cocaine and substances used in adulteration using near-infrared Raman spectroscopy and infrared absorption spectroscopy.

Identification of cocaine and subsequent quantification immediately after seizure are problems for the police in developing countries such as Brazil. This work proposes a comparison between the Raman and FT-IR techniques as methods to identify cocaine, the adulterants used to increase volume, and possible degradation products in samples seized by the police. Near-infrared Raman spectra (785 nm excitation, 10 sec exposure time) and FT-IR-ATR spectra were obtained from different samples of street cocaine and some substances commonly used as adulterants. Freebase powder, hydrochloride powder, and crack rock can be distinguished by both Raman and FT-IR spectroscopies, revealing differences in their chemical structure. Most of the samples showed characteristic peaks of degradation products such as benzoylecgonine and benzoic acid, and some presented evidence of adulteration with aluminum sulfate and sodium carbonate. Raman spectroscopy is better than FT-IR for identifying benzoic acid and inorganic adulterants in cocaine.

The effects of transcranial LED therapy (TCLT) on cerebral blood flow in the elderly women.

During aging processes, there is a range of functional changes, where we can highlight the disease related to the central nervous system, such as Alzheimer disease and others forms of dementia. This study investigated the effects of transcranial light emitting diode (LED) on cerebral blood flow in healthy elderly women analyzed by transcranial Doppler ultrasound (TCD) of the right and left middle cerebral artery and basilar artery. Twenty-five noninstitutionalized elderly women (mean age 72 years old), with a cognitive status >24, were assessed using transcranial Doppler ultrasound on two separate occasions: pre-irradiation and post-transcranial LED therapy (TCLT). Prior to this, they answered two questionnaires: the perceived stress scale and the general health questionnaire. TCLT (627 nm, 70 mW/cm(2), 10 J/cm(2)) was performed at four points of the frontal and parietal region for 30 s each, totaling 120 s two times per week for 4 weeks. Paired t-test results showed that there was a significant improvement after TCLT with increase in the systolic and diastolic velocity of the left middle cerebral artery (25 and 30%, respectively) and basilar artery (up to 17 and 25%), as well as a decrease in the pulsatility index and resistance index values of the three cerebral arteries analyzed (p < 0.05). TCD parameters showed improvement in the blood flow on the arteries analyzed. TCLT promoted a blood and vasomotor behavior of the basilar and middle cerebral arteries in healthy elderly women.

Near-infrared Raman spectroscopy to detect anti-Toxoplasma gondii antibody in blood sera of domestic cats: quantitative analysis based on partial least-squares multivariate statistics.

Toxoplasmosis is an important zoonosis in public health because domestic cats are the main agents responsible for the transmission of this disease in Brazil. We investigate a method for diagnosing toxoplasmosis based on Raman spectroscopy. Dispersive near-infrared Raman spectra are used to quantify anti-Toxoplasma gondii (IgG) antibodies in blood sera from domestic cats. An 830-nm laser is used for sample excitation, and a dispersive spectrometer is used to detect the Raman scattering. A serological test is performed in all serum samples by the enzyme-linked immunosorbent assay (ELISA) for validation. Raman spectra are taken from 59 blood serum samples and a quantification model is implemented based on partial least squares (PLS) to quantify the sample's serology by Raman spectra compared to the results provided by the ELISA test. Based on the serological values provided by the Raman/PLS model, diagnostic parameters such as sensitivity, specificity, accuracy, positive prediction values, and negative prediction values are calculated to discriminate negative from positive samples, obtaining 100, 80, 90, 83.3, and 100%, respectively. Raman spectroscopy, associated with the PLS, is promising as a serological assay for toxoplasmosis, enabling fast and sensitive diagnosis.

Laser biomodulation on L 929 cell culture.

OBJECTIVE: The aim of the present study was to analyze the effects of photobiomodulation using a 904-nm diode laser at two energy densities (6 J/cm(2) and 50 mJ/cm(2)) on L929 fibroblast cells. BACKGROUND: Low-power laser irradiation (LPLI) is a non-pharmacological resource that induces important in vitro photobiomodulation on cell cultures and tissues. METHODS: Irradiation was performed for three days at 24-h intervals. After each interval, the cells were stained with MitoTracker Orange and DioC6 dyes to assess the photobiomodulatory effects of irradiation on mitochondrial activity and changes in the endoplasmic reticulum. The MTT assay [3-(4.5-dimethylthiazol-2-yl)-2.5 diphenyltetrazolium bromide] was used to evaluate cell proliferation. RESULTS AND CONCLUSIONS: The fluorescence microscopy assessment of mitochondria and endoplasmic reticulum in cells irradiated with 6 J/cm(2) and 50 mJ/cm(2) demonstrated intense mitochondrial activity, which was confirmed by DioC6 staining. Reticular activity was observed stemming from increased protein synthesis. Photobiomodulation with 50 mJ/cm(2) was slightly higher than with 6 J/cm(2), as demonstrated by fluorescence microscopy results. Photobiomodulation was also time-dependent, with better results 72-h after irradiation.

Raman spectroscopy study of atherosclerosis in human carotid artery.

Fourier-transform (FT)-Raman spectroscopy has been used for identification and evaluation of human artherosclerotic lesions, providing biochemical information on arteries. In this work, fragments of human carotid arteries postmortem were analyzed using a FT-Raman spectrometer operating at an excitation wavelength of 1064 nm, power of 200 mW, and spectral resolution of 4 cm(-1). A total of 75 carotid fragments were spectroscopically scanned and FT-Raman results were compared with histopathology. Discriminant analysis using Mahalanobis distance was applied over principal components scores for tissue classification into three categories: nonatherosclerotic, atherosclerotic plaque without calcification and with calcification. Nonatherosclerotic artery, atherosclerotic plaque, and calcified plaque exhibit spectral signatures related to biochemicals presented in each tissue type, such as bands of collagen and elastin (proteins), cholesterol and its esters, and calcium hydroxyapatite and carbonate apatite, respectively. Spectra of nonatherosclerotic artery were then classified into two groups: normal and discrete diffuse thickening of the intima layer (first group) and moderate and intense diffuse thickening of the intima layer (second group). FT-Raman could identify and classify the tissues found in the atherosclerotic process in human carotid in vitro and had the ability to identify alterations to the diffuse thickening of the intima layer and classify it depending on the intensity of the thickening.

Resposta in vitro de células gliossarcoma 9L após terapia fotodinâmica utilizando imagem de bioluminescência / In vitro gliosarcoma 9L cells response after photodynamic therapy using bioluminescence imaging

Objetivos: O presente estudo tem o objetivo de demonstrar os resultados iniciais da utilização da técnica deimagem de bioluminescência como método de monitoramento do tratamento de células de gliossarcoma de rato 9L após Terapia Fotodinâmica (TFD) utilizando o ácido aminolevulínico (ALA) como agente fotossensibilizante. Métodos: Para o presente estudo, células 9L foram transfectadas com um plasmídeo contendo o gene da luciferase, permitindo que essa linhagem celular produzisse a proteína luciferase, um dos substratos necessários para a reação de bioluminescência. No presente estudo, a TFD foi realizada utilizando diferentes doses de luz e de ácido aminolevulínico. Para validar a técnica de imagem de bioluminescência como método para o monitoramento da resposta de células tumorais e para a verificação da correlação entre o sinal luminoso e o grau de morte celular após TFD, foi utilizado o ensaio de viabilidade celular com Sulforodamina B (SRB). Resultados: Os resultados destetrabalho mostram alta correlação entre o número de células e o sinal de bioluminescência (R2 = 0,996). Os ensaios de viabilidade celular utilizando a técnica SRB, mostraram excelente correlação entre o número relativo de células sobreviventes após TFD e o sinal de bioluminescência, mostrando que a técnica de imagem de bioluminescência pode ser utilizada para o monitoramento da resposta tumoral após o tratamento de células tumorais com TFD. Os resultados do tratamento das células com terapia fotodinâmica mostram que a taxa de indução de morte celular varia de acordo com a dose de luz e fotossensibilizante empregada durante o tratamento. Nesse sentido, doses maiores de TFD resultaram em níveis mais elevados de indução de morte celular, com efeitos mais prolongados observados através do sinal bioluminescente decorridos 48 horas após a TFD. Conclusão: Este estudo demonstra que a técnica de imagem de bioluminescência pode ser utilizada para o estudo dos efeitos da terapia fotodinâmica em células tumorais. Trabalhos em animais estão em andamento no presente momento para validar a técnica de imagem de bioluminescência após TFD in vivo.

Dentin evaluation after Nd:YAG laser irradiation using short and long pulses.

OBJECTIVE: Several reports have demonstrated the advantages of using the Nd:YAG laser to reduce dentin permeability by melting the dentin surface. A comparative study using different pulse durations can be useful to obtain further information about the laser-hard tissue interaction. MATERIALS AND METHODS: The present study pursues the evaluation of the morphological and chemical changes in human dentin surface resulting from Nd:YAG laser (lambda = 1064 nm) irradiation, with a total energy of 0.9 J distributed in 1, 2, 3, and 6 pulses with different pulse durations to promote surface melting and dentinal tubule occlusion. After irradiation, the samples were submitted to scanning electron microscopy (SEM) analysis for morphological study and energy-dispersive spectrometry (EDS) analysis for evaluation of the concentration of calcium and phosphorous in the melted layer. RESULTS: SEM analysis of the irradiated dentin surface showed surface structural changes due to laser irradiation, where the morphological changes are dependent on the laser pulse duration. EDS analysis showed an increase of calcium and phosphorous concentrations after Nd:YAG laser exposure, but no correlation with the number of pulses or pulse duration was found. CONCLUSION: Our results suggest that longer interaction times resulted in more evident effects with more melted substrate than shorter pulses, and in both cases the resultant melted layer contains a greater concentration of inorganic substances than non-irradiated dentin.

Effect of low-power GaAlAs laser (660 nm) on bone structure and cell activity: an experimental animal study.

Low-level laser therapy (LLLT) is increasingly being used in the regeneration of soft tissue. In the regeneration of hard tissue, it has already been shown that the biomodulation effect of lasers repairs bones more quickly. We studied the activity in bone cells after LLLT close to the site of the bone injury. The femurs of 48 rats were perforated (24 in the irradiated group and 24 in the control group) and the irradiated group was treated with a GaAlAs laser of 660 nm, 10 J/cm2 of radiant exposure on the 2nd, 4th, 6th and 8th days after surgery (DAS). We carried out histomorphometry analysis of the bone. We found that activity was higher in the irradiated group than in the control group: (a) bone volume at 5 DAS (p=0.035); (b) osteoblast surface at 15 DAS (p=0.0002); (c) mineral apposition rate at 15 and 25 DAS (p=0.0008 and 0.006); (d) osteoclast surface at 5 DAS and 25 DAS (p=0.049 and p=0.0028); and (e) eroded surface ( p=0.0032). We concluded that LLLT increases the activity in bone cells (resorption and formation) around the site of the repair without changing the bone structure.

Optothermal transfer simulation in laser-irradiated human dentin.

Laser technology has been studied as a potential replacement to the conventional dental drill. However, to prevent pulpal cell damage, information related to the safety parameters using high-power lasers in oral mineralized tissues is needed. In this study, the heat distribution profiles at the surface and subsurface regions of human dentine samples irradiated with a Nd:YAG laser were simulated using Crank-Nicolson's finite difference method for different laser energies and pulse durations. Heat distribution throughout the dentin layer, from the external dentin surface to the pulp chamber wall, were calculated in each case, to investigate the details of pulsed laser-hard dental tissue interactions. The results showed that the final temperature at the pulp chamber wall and at the dentin surface are strongly dependent on the pulse duration, exposure time, and the energy contained in each pulse.

Use of near-infrared raman spectroscopy to detect IgG and IgM antibodies against Toxoplasma gondii in serum samples of domestic cats.

Near-infrared Raman spectroscopy (NIRS) is a particularly promising technique that is being used in recent years formany biomedical applications. Optical spectroscopy has gained increasing prominence as a tool for quantitative analysis of biological samples, clinical diagnostic, concentration measurements of blood metabolites and therapeutic drugs, and analysis of the chemical composition of human tissues. Toxoplasmosis is an important zoonosis in public health, and domestic cats are the most important transmitters of the disease. This disease can be detected by several serological tests, which usually have a high cost and require a long time. The goal of this work was to investigate a new method to diagnosis Toxoplasma gondii infections using NIRS. In order to confirm antibody detection, 24 cat blood serum samples were analyzed by the Raman spectra, from which 23 presented positive serology to toxoplasmosis and one was a reference negative serum. Characteristic Raman peaks allowed differentiation between negative and positive sera, confirming the possibility of antibody detection by Raman spectroscopy. These results give the first evidence that this technique can be useful to quantify antibodies in cat sera.

Correlation between near-infrared Raman spectroscopy and the histopathological analysis of atherosclerosis in human coronary arteries.

BACKGROUND AND OBJECTIVES: Modern diagnostic methods such as near-infrared Raman spectroscopy (NIRS) allow quantification and evaluation of human atherosclerotic lesions, which can be useful in diagnosing coronary artery disease. The objective of the present study is to obtain feasible diagnostic information to detect atheromatous plaque using NIRS combined with discriminant analysis. STUDY DESIGN/MATERIAL AND METHODS: An 830 nm Ti: sapphire laser pumped by an argon laser provides near-infrared excitation. A spectrograph disperses light scattered from arterial tissue and a liquid-nitrogen cooled CCD detects the Raman spectra. A total of 111 arterial fragments were scanned and Raman results were compared with histopathology. Principal components analysis (PCA) and Mahalanobis distance (m-distance) were used to model an algorithm for tissue classification into three categories: non-atherosclerotic (NA), non-calcified (NC), and calcified (C) using Raman spectra. Spectra were randomly separated into training and prospective groups. RESULTS: It has been found that, for the NA tissue, the algorithm has sensitivity of 84 and 78% and specificity of 91 and 93% for training and prospective groups, respectively. For the NC tissue the algorithm has sensitivity of 88 and 90% and specificity of 88 and 83%. For the C tissue both sensitivity and specificity were maximum, 100%. CONCLUSIONS: An algorithm using PCA and discriminant analysis based on m-distance has been developed and successfully applied to diagnose coronary artery disease by NIRS obtaining good sensitivity and specificity for each tissue category.

Sensor de Pressäo a fibra óptica de média birrefringência / Pressure sensor and medium birefringence optical fiber

Neste trabalho são apresentados e discutidos os resultados experimentais de um sistema para medida e peso para aplicações ergométricas, utilizando fibra óptica de média birrefrigência. O sensor de 10cm de comprimento contém uma fibra óptica monomodo com comprimento de onda de corte de 1,24 mum, operando com um laser semicondutor emitindo em 1,3 mum. Os resultados experimentais mostram que o sistema responde sem redundância na faixa de 0 a 130 Kg, com sensibiidade de aferir variações de dezenas de gramas.

- ln this work the experimental results for a weighing sensor for ergometric applications using medium birefringence optical fíber are presented. The sensor is 1 Ocm long and it has a singlemode optical fiber inside, with 1.24 cutoff wavelength, and a semiconductor laser emitting at 1.3 pm. The experimental results show that the system works without redundancy from O to 130 kg, with a sensibility good enough to detect variations of order of O.O 1 Kg.

Cateter óptico para aplicaçöes espectroscópicas / Optic catheter for spectroscopic applications

Neste trabalho é apresentada a caracterização de cateteres utilizando fibras ópticas para aplicações de espectroscopia in vivo. Estes cateteres apresentam a configuração de seis fibras em torno de uma fibra central de mesmo diâmetro, sendo estas fibras do tipo multimodo com diâmetro de núcleo de 100 mum. A eficiência de coleta de radiação para este tipo de cateter é da ordem de 0,05 por cento, variando em função da distância ao tecido e da distribuição de intensidade de sinal retroespalhado na superfície do mesmo.

ln this work we describe the characterization of fiber optic catheters for in vivo spectroscopy applications. This type of catheters have the geometry of one central fiber sun-ounded by six 100 µm diameter multimode fibers. The collection efficiency of these type of catheter is about 0.05% and it changes with the distance to the tissue surface and the signal intensity distribution on it.

Estudo da eficiência de captaçäo de energia entre cateter momofibra e polifibra / Study about the efficiency of energy uptake between mutifiber and single fiber catheter

Neste trabalho é realizado o estudo comparativo entre o cateter convencional tipo polifibras (6x1), e uma nova configuração do mesmo dispositivo utilizando apenas uma fibra. Os resultados demostram que esta nova configuração apresenta melhor eficiência da coleta de radiação do sinal retroespalhado pela amostra.

This work aims the comparative study between a standard multifiber catheter (6x1), and a new configuration of the same device using only single fiber. The results show that this new configuration has better collection efficiency for the backscattering signal from the sample.

Filtragem de sinais Raman de tecidos biológicos para reduçäo de ruído e background / Filter of Raman signals of biological tissue for background and noise reduction

A redução do tempo de aquisição do sinal e processamento de espectros Raman obtidos à partir de tecidos biológicos é um passo importante para se tornar os sistemas laboratoriais uma ferramenta clínica. Neste trabalho é apresentado a extração de fluorescência e ruído de um espectro Raman de uma coronária humana.