PEDRO ARTHUR AUGUSTO DE CASTRO
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Resumo IPEN-doc 29573 FTIR imaging on glass substrates evaluation of histological skin burn injuries specimens treated by femtosecond laser pulses2022 - ZEZELL, DENISE; CASTRO, PEDRO; DEL-VALLE, MATHEUS; CAMILLO-SILVA, CARLOS; SAMAD, RICARDO; DE ROSSI, WAGNER; SANTOS, MOISESBurn injuries continue to be one of the leading causes of unintentional death and injury in low- and middle-income countries [1]. Burns are considered an important public health problem, because in addition to physical problems that can lead the patient to death, they cause psychological and social damage. An estimated 180,000 deaths every year are caused by burns [2]. The use of infrared (IR) spectroscopy for studying biological specimens is nowadays a wide and active area of research. The IR microspectroscopy has proved to be an ideal tool for investigating the biochemical composition of biological samples at the microscopic scale, as well as its fast, sensitive, and label-free nature [3]. IR image spectral histopathology has shown great promise as an important diagnostic tool, with the potential to complement current pathological methods, reducing subjectivity in biopsy samples analysis. However, the use of IR transmissive substrates which are both fragile and prohibitively very expensive, hinder the clinical translation. The goal of this study is to evaluate the potential of discriminating healing process, in burned skin specimens treated with ultrashort pulses laser 3 days after the burn. This study is considering a previous paper [4], in which it analyzed only micro-ATR-FTIR spectra of a frozen sample point. The specimens were obtained from third degree burn wound. The wounds treatment were performed three days after the burn, and the animals were sacrificed 3 and 14 days post-treatment. Using coverslipped H&E stained tissue on glass from previous histopathological analysis and applying the analytical techniques PCA and K-means on N−H, O−H, and C−H stretching regions occurring at 2500−3800 cm−1 (high wavenumber region), were possible to discriminate burned epidermal and dermal regions from irradiated in same regions on sample. In the figures is shown the average spectrum at (a) day 3 and (b) day 14. , in both there were increase of burned+laser treated bands. The great potential of this study was to analyse coverslipped H&E stained tissue on glass, without compromising the histopathologist practices and contribute for clinical translation.Tese IPEN-doc 29223 Chemometrics of biological tissues using infrared spectroscopy2022 - CASTRO, PEDRO A.A. deVibrational spectroscopy techniques have been considered for the diagnosis of diseases. In particular, Fourier transforms infrared spectroscopy (FTIR) is an optical method widely adopted in biomedical research. This is because FTIR images allow the collection of various molecular information enabling a correlation between spectral characteristics and structural changes. Incorporating qualitative and quantitative aspects is essential to implementing a chemometric procedure that transcribes spectral variance into meaningful information. In the case of burns, the degree of tissue damage will strongly depend on several factors, such as severity, depth, and size, which influence the healing process. Laser scar treatment has been used for decades for post-healing remodeling. However, conventional laser techniques may not wholly normalize mature scar tissue, and a preventive approach to laser exposure during wound healing has emerged. Early intervention has already been investigated in several clinical studies, but there is no consensus. Therefore, the objectives of this study were; to investigate FTIR in biological tissue analysis to provide better chemometric strategies; to study the feasibility of FTIR to establish a profile of the skin and bone tissue, and to evaluate the effect of laser on wound healing and the development of a new chemometric methodology for automatic digital dewaxing. To this end, thermal injuries were performed on the back of Wistar rats by applying water vapor at 90ºC for 12s. Subsequently, on day three post-burn, laser irradiation was performed. In the laser procedure, 3 protocols were adopted: 1) Er:glass (SellasEvo®) with 3025FPA, 100mJ, 3s; 2) LLT with red laser with = 660nm, power 40mW; 3) Combination of the two lasers with the previous parameters. Then, a histological and chemometric analysis was performed. For the chemometric evaluation of the data, an analysis protocol was developed based on a pre-processing using OCTAVVS followed by implementing a baseline, threshold, and application of PCA. From that point, the images are classified, they can be normalized and go through the Fuzzy C-means (FCM) technique to determine the spectral differences. All-laser groups demonstrate a shrinkage of the wound region in the histopathological findings. Er:Glass, showed a smaller retraction area and less epithelium recovery. The PBM group has a dense crust. Both treatments demonstrate intense shrinkage and more consistent recovery of the epidermis. In the FCM, three trends occur 1) in the range of 900-950 cm-1 it has a significantly reduced amplitude in the control group; 2) in the range of 1200-1250 cm-1 the control group has a similar amplitude demonstrating bands of sensitive markers; and 3) the amide regions provide various spectral markers. The control group, as expected, remains with minimal changes in this FTIR region. On the other hand, in the PBM group, there is a shift from 1660 to 1679 cm-1, which may correspond to changes in the molecular structure of a protein in the context of type I collagen. FTIR is a valuable alternative for tissue research on a biochemical level through fingerprints that can be translated into morphological features.Artigo IPEN-doc 28104 Healing status of bum wound healing2021 - CASTRO, PEDRO; ZORN, TELMA; ZEZELL, DENISEThe purpose of this study is to use infrared spectroscopy (FTIR) for monitoring biological changes in burned skin. Wistar rats dorsum samples were compared to healthy group samples at 7, 14, 21 days after burn. Proteins changes of burn wounds were monitored by area under the curve (AUC) of bands at 1630 cm 1 , 1543 cm 1 and 1743 cm 1 . Kruskal-Wallis normality tests, unpaired t test with Welch's correction were used to evaluate the differences between AUC. These bands suggest association between collagen activity during wound healing stages. Our result indicates progressive recovery of 7,14 and 21 days tissues when compared with the healthy group.Artigo IPEN-doc 27120 Monitoring the progress and healing status of burn wounds using infrared spectroscopy2020 - CASTRO, PEDRO A.A.; LIMA, CASSIO A.; MORAIS, MYCHEL R.P.T.; ZORN, TELMA M.T.; ZEZELL, DENISE M.Burns are one of the leading causes of morbidity worldwide and the most costly traumatic injuries. A better understanding of the molecular mechanisms in wound healing is required to accelerate tissue recovery and reduce the health economic impact. However, the standard techniques used to evaluate the biological events associated to wound repair are laborious, time-consuming, and/or require multiple assays/staining. Therefore, this study aims to evaluate the feasibility of Fourier transform infrared (FT-IR) spectroscopy to monitor the progress and healing status of burn wounds. Burn injuries were induced on Wistar rats by water vapor exposure and biopsied for further histopathological and spectroscopic evaluation at four time-points (3, 7, 14, and 21 days). Spectral data were preprocessed and compared by principal component analysis. Pairwise comparison of post-burn groups to each other revealed that metabolic activity induced by thermal injury decreases as the healing progresses. Higher amounts of carbohydrates, proteins, lipids, and nucleic acids were evidenced on days 3 and 7 compared to healthy skin and reduced amounts of these molecular structural units on days 14 and 21 postburn. FT-IR spectroscopy was used to determine the healing status of a wound based on the biochemical information retained by spectral signatures in each phase of healing. Our findings demonstrate that FT-IR spectroscopy can monitor the biological events triggered by burn trauma as well as to detect the wound status including full recovery based on the spectral changes associated to the biochemical events in each phase.Artigo IPEN-doc 22896 Multimodal evaluation of ultra-short laser pulses treatment for skin burn injuries2017 - SANTOS, MOISES O. dos; LATRIVE, ANNE; CASTRO, PEDRO A.A. de; ROSSI, WAGNER de; ZORN, TELMA M.T.; SAMAD, RICARDO E.; FREITAS, ANDERSON Z.; CESAR, CARLOS L.; VIEIRA JUNIOR, NILSON D.; ZEZELL, DENISE M.Thousands of people die every year from burn injuries. The aim of this study is to evaluate the feasibility of high intensity femtosecond lasers as an auxiliary treatment of skin burns. We used an in vivo animal model and monitored the healing process using 4 different imaging modalities: histology, Optical Coherence Tomography (OCT), Second Harmonic Generation (SHG), and Fourier Transform Infrared (FTIR) spectroscopy. 3 dorsal areas of 20 anesthetized Wistar rats were burned by water vapor exposure and subsequently treated either by classical surgical debridement, by laser ablation, or left without treatment. Skin burn tissues were noninvasively characterized by OCT images and biopsied for further histopathology analysis, SHG imaging and FTIR spectroscopy at 3, 5, 7 and 14 days after burn. The laser protocol was found as efficient as the classical treatment for promoting the healing process. The study concludes to the validation of femtosecond ultra-short pulses laser treatment for skinburns, with the advantage of minimizing operatory trauma.