The target-specific drug release which is depending on the pH of the medium appears precious approach for increasing of drug selectivity and further efficacy in photodynamic diagnosis (PDD) and therapy (PDT) of cancer. A strongly hydrophobic Zn (II) phthalocyanine (ZnPc) along with its liposomal formulates belongs to the clinically approved phthalocyanines for cancer photodiagnosis PDD and PDT. Nevertheless closed to the ideal photo-physicochemical properties the planar molecular structure of ZnPc easily forms aggregates which finally limit the phototoxic effect. Different strategies are on hand, all aiming to minimize the formation of photo-inactive associates. At present time, the polymeric carrier systems are still under development as carrier systems for biomedical applications. The present work evaluated the efficiency of two recently proposed graft - copolymers namely polyimide (PI)-graft-polymethacrylic acid (PAT2 and PAT3) as vehicles of hydrophobic ZnPc. The potential of the proposed polymeric brushes with great number of carboxylic groups, for conformational changes in different pH media, can be used for ZnPc release as studied by fluorescence spectroscopy. Complexes developed were investigated in vivo on rat model system of gastric carcinoma lesions for photodiagnosis and photodynamic treatment of neoplasia and compared with the application of ZnPcs solely. The contrast achieved of the fluorescence detection using polymer brushes - phthalocyanine conjugate is about twice higher that standard applied photosensitizer using same protocol and doses of application. The photodynamic treatment efficacy was also significantly improved allowing to receive full response using doses of 0,5-1 mg/kg of the conjugates. |To obtain same efficiency for the ZnPc PDT in the gastric lesions treated drug doses of 1,5-2 mg/kg had to be applied. Initially detected fluorescence signal and photo-bleaching dynamics were used for the monitoring of the processes of accumulation and photodynamic treatment processes.
The early stages of gastric cancer are not recognized due to its asymptomatic development. Detection of patho- logical changes can be achieved using the photodynamic diagnostics, but this approach provides rather low tumor selectivity. Additional opportunities to improve the detection of precancerous stages may be provided by ana- lyzing blood circulation in the vessels of the stomach. We consider the possibilities of multiresolution analysis in recognizing changes in blood circulation in rats with different levels of the fluorescent signal in the stomach associated with precancerous stages.
Combined spectroscopic and biochemical measurements were used to improve diagnostic accuracy and to evaluate gastrointestinal tract (GIT) neoplasia development parameters noninvasively. Experiments were performed in mongrel male rats divided into 2 groups – control and experimental. To induce gastric cancer, the rats underwent to chronic stress (overpopulation during 9 months) and diet including the daily using of mtoluidine (25 μg/kg weight) in food and water with a solution of sodium nitrite (0.2%) for 9 months. We studied effectiveness of 5-ALA/PpIX fluorescent analysis of gastric carcinoma and biochemical stress-corresponding indices detection for early diagnosis of primary gastric tumours and their metastatic spreading in liver. Affected by precancerous and cancerous alterations mucosa reveal red fluorescence, related to the accumulation of 5- ALA/PpIX. Liver tissues investigated also presented increase of the red fluorescence, which was used as an indicator for possible pathologic process detection there. The histological examination revealed liver metastases in 67.8 % of the rats with gastric cancer. Biochemical indicators detected malignant alterations presence in GIT, and fluorescent observation addressed the exact area and borders of neoplastic lesions. 5-ALA/PpIX fluorescence detection allow to find and precisely map premalignant and malignant areas of gastric mucosa and liver metastases of stress-induced gastric heterogeneous adenocarcinoma and biochemical evaluation of stressrelated compounds increased the efficiency of such diagnosis and reveal information about the dynamics of lesions development. Diagnostic accuracy achieved using fluorescent detection reaches 93% for gastric carcinoma, and 87% for pre-cancerous mucosa alterations observed.
Transcranical photobiomodulation (tPBM, 1267 nm, 32 J/cm2) is effective non-invasive method for clearance of beta-amyloid from the brain in mice with Alzheimer’s disease and for improvemen of their neurological status.
Gastrointestinal cancerous lesions were induced in laboratorial animals using prolonged exposure to social (overpopulation) and chemical stress (nitrosamine diet), mimicking typical stress factors for humans in the big cities. Twenty laboratorial rats with developed stress-induced neoplasia in gastrointestinal tract were used. Three of them formed control group - without application of exogenous contrast fluorescent marker. Exogenous fluorescent spectroscopic measurements were used to evaluate gastrointestinal tract (GIT) neoplasia development noninvasively using excitation at 405 nm and emission was detected in the region of 500-850 nm using microspectrometer. We used as exogenous fluorescent marker protoporphyrin IX applying its precursor delta aminolevulinic acid in a dose of 20mg/kg, in laboratorial rats with stress-induced neoplasia. Spectroscopic analysis of primary gastric tumours and their metastases spreading in rats’ liver was carried out on animals in vivo, as excitation and emission light were delivered by fiberoptic probe 6+1 to the organs investigated during open surgery procedure. Biochemical indicators detected malignant alterations presence in GIT were also evaluated and used as complementary indices for lesions’ growth. Fluorescent observation using exogenous fluorophore addressed the exact area and borders of neoplastic lesions in stomach and liver. In some cases in inflammatory areas significant accumulation of delta-ALA had place, which lead to false-positive fluorescent signal of protoporphyrin IX in these tissues. Photodiagnostics accuracy of 93% was reached for cancerous cases and 87% for gastric pre-cancer lesions evaluation. Very good correlation between fluorescence data and histology examination of the lesions investigated was achieved as well.
Gastrointestinal tract neoplasia is on the third place of death statistics among oncological diseases, which is associated with the difficulty of early diagnosis of this disease. This study aims to evaluate applicability of delta-aminolevulinic acid / protoporphyrin IX photosensitizer (5-ALA / PpIX) as diagnosing agent for detection of small and large intestine neoplasia. The experiments were performed in vivo on white laboratorial rats. The animals were divided into 2 groups: control and experimental (overpopulation social stress and a diet, including nitrites and amines) ones. The results of the studies, averaged for each type of tissue, clearly show significant differences between normal tissues and malignant intestine tissues of the rats useful for fluorescent-based discrimination of the tissue state. The fluorescence from malignant areas in small and large intestine represented bright red fluorescence with specific maxima at 635 and 704 nm, typical for PpIX, due to the high accumulation of this photosensitizer in the affected tissues. The accumulation of protoporphyrin IX in the intestine cancerous tissues was very well pronounced. A contrast achieved between normal and abnormal tissue sites at 635 nm fluorescence emission was more than an order of magnitude in the case of large intestine neoplasia, which allows using the 5-ALA/PpIX as a sensitive indicator of gastrointestinal carcinoma.
Gastric cancer often appears without preliminary symptoms that could be detected and recognized at early stages. Its diagnostic is sometime provided based on photodynamic techniques, although the latter are restricted by quite poor tumor selectivity. Due to this, additional experimental and data processing tools are required to improve diagnostic abilities. In this study we analyze how complexity measures can detect early changes in the normal gastric microcirculation. We apply a recently developed approach based on the multiscale entropy and show that the estimated measures of SampEn related to distinct ranges of scales enable the detection of early gastric cancer that is in accordance with the results of histological analysis.
Here in ex vivo (confocal imaging) and in vivo (optical coherent tomography) experiments on adult mice, we clearly demonstrate that the meningeal lymphatic drainage is an important mechanism for the brain clearing that might be also pathway for the brain clearing from metabolites and waste products that requests further detailed studies. Photodynamic effects stimulate the meningeal/cervical drainage opening new ideas for development of novel methods of activation of brain clearing that might be progressive strategies in therapy of neurovegetative diseases.
In our experiments on rats we performed visualization of brain clearing from substances injected into the brain parenchyma using optical coherent tomography and fluorescence microscopy. Our results uncovered that the meningeal lymphatic system play an important role in the brain clearing processes, where the deep cervical lymph node is the first anatomical station of cerebral fluid exist from the brain. These data shed light on the novel mechanisms underlying drainage and clearing function of the brain.
Gastric cancer is the main cause of death among oncological diseases of the gastrointestinal tract, which is associated with the difficulty of early diagnosis of this disease. This study aims to develop a new method for diagnosing gastric cancer using the delta-aminolevulinic acid / protoporphyrin IX photosensitizer (5-ALA / PpIX). The experiments were performed on white rats of average weight 250 grams. The animals were divided into 2 groups: control and experimental (stress + diet, including nitrites and carcinogenic amines) ones. The results of the studies, averaged for each type of tissue, clearly show significant differences between normal, precancerous and malignant changes in the stomach tissues of rats on the background of application of photodynamic diagnostics. The fluorescence from malignant neoplasms in the organ was maximal at 635 nm, which corresponds to the maximum emission of protoporphyrin due to the high accumulation of this photosensitizer in the affected tissues. The most important result of the study was the detection of precancerous changes (atrophic gastritis, confirmed by histological methods), using photodynamic diagnostics with photosensitizer 5-ALA / PpIX. Intensity of fluorescence in case of precancerous changes occupied an intermediate value between that in normal and in malignant mucosa of the stomach.
Stroke and traumatic brain injury are often associated with formation of brain edema, which is a potentially fatal pathological state provoking extensive accumulation of fluid in the brain tissues resulting in elevation of intracranial pressure and leading to impaired nerve function. There is only symptomatic anti-edema therapy is currently available. Therefore, the development of novel strategies to remove edema fluid is required. The brain edema is classified as vasogenic or cytotoxic edema, which associated with excess accumulation of fluid (edema) around capillaries resulting from disruption of the blood-brain barrier (BBB) or intracellular spaces (cell swelling) of the brain, respectively. In this brief review, we discuss possible mechanisms underlying brain edema formation and new strategies in development of novel of anti-edema drugs.
Here we present preliminary experimental data suggesting about involvement of the meningeal and cervical lymphatics in neurorehabilitation. Using model of hemorrhagic stroke, immunohistochemical analysis and atomic absorption spectroscopy, we clearly demonstrate the lymphatic clearance from the blood after stroke via the meningeal lymphatic vessels with further accumulation of hemosiderin and iron (products of disaggregated hemoglobin) in the deep cervical node (dcLN). The optical coherent tomography (OCT) was used for in vivo monitoring of accumulation of gold nanorods (92 nm in diameter) in the dcLN after their injection into the cisterna magna with the aim of mimicking of the brain clearance from of blood. The both ex vivo and in vivo data show the lymphatic clearance from subjects (the blood/GNRs) injected into the subarachnoid space that might be an important mechanism of neurorehabilitation after the intracranial hemorrhages.
In the recent study delta-ALA/PpIX is used as fluorescent marker for tumor detection in stomach, small intestine and colon. 5-ALA was administered per os two hours before measurements at dose 20 mg/kg weight. Twenty laboratorial rats with developed neoplasia in gastrointestinal tract were used. Three of them formed control group - without application of exogenous 5-ALA/PpIX, as a contrast fluorescent marker. Their spectral properties were evaluated and used for correction of endogenous sources of fluorescence from stomach and intestine lesions detected. Macroscopic fluorescence spectroscopy parameters of gastrointestinal tissues of the other 17 animals, which were sensitized with 5- ALA?PpIX, were obtained on excitation at 405 nm. Excitation and emission light was delivered by fiberoptic probe 6+1 to the organs investigated in vivo during open surgery procedure on the laboratorial animals used. The fluorescence detected from tumor sites has very complex spectral origins. It consists of autofluorescence, fluorescence from exogenous fluorophores and re-absorption from the chromophores accumulated in the tissue investigated. Spectral features observed during endoscopic investigations could be distinct as the next regions: 450-630 nm region, where tissue autofluorescence is observed; 630-710 nm region, where fluorescence of PpIX is clearly pronounced; 530-580 nm region, where minima in the autofluorescence signal are observed, related to re-absorption of oxy-hemoglobin in this spectral area. The contrast achieved in normal vs. neoplastic gastrointestinal tissues was evaluated. Influence of endogenous 5-ALA/PpIX accumulation was taken into account; as well the role of inflammatory processes on the PpIX distribution and accumulation into the body was evaluated. Dimensionless ratio R=I635/I560 is evaluated as highly informative one for development of a simple algorithm for differentiation, which could has high diagnostic accuracy for evaluation of tumor vs. inflammatory vs. normal mucosa, using 1-D spectral results. Very good correlation between fluorescence data and histology examination of the lesions investigated was achieved as well.
In our study aluminium–phthalocyanine (AlPc) compounds were applied for exogenous fluorescent diagnostics of adenocarcinoma of stomach in laboratorial animals (male adult rats (n=50)) after application of an experimental model of adenocarcinoma formation with metastasis. The neoplastic lesions were developed under the influence of social stress, as well as chemical stress using nitrosamines during 9 months period of application. Significant fluorescence signal in the region of 670-700 nm was observed in the neoplastic lesions, which absent in the normal mucosa investigated, related to the fluorescence of accumulated in the tumour area phthalocyanines. Autofluorescence background covered region at 450-650 nm with a maximum of 480-520 nm and consists mainly of protein cross-links and co-enzymes - NADH and flavin signals. Endogenous porphyrins fluorescence was also observed in the lesions with a maximum at 630-640 nm. Other organs (liver, small bowel, lungs) were also investigated for a presence of metastases. Histology examination of the lesions was used as a “gold standard” for comparison of the spectral data. Visually, a presence of accumulation sites of AlPc in a form of bright pink patches after excitation at 405 nm was observed, vs. healthy tissue, which remained blue-violet due to autofluorescence signal. Thus allow to use the AlPc fluorescence discrimination not only in spectroscopic mode of detection, but for imaging of the lesions investigated, which is more preferable in the case of clinical applications during endoscopic observations in humans.
Here we studied the role of cerebral lymphatic system in the brain clearing using intraparenchymal injection of Evans Blue and gold nanorods assessed by optical coherent tomography and fluorescence microscopy. Our data clearly show that the cerebral lymphatic system plays an important role in the brain cleaning via meningeal lymphatic vessels but not cerebral veins. Meningeal lymphatic vessels transport fluid from the brain into the deep cervical node, which is the first anatomical “station” for lymph outflow from the brain. The lymphatic processes underlying brain clearing are more slowly vs. peripheral lymphatics. These results shed light on the lymphatic mechanisms responsible for brain clearing as well as interaction between the intra- and extracranial lymphatic compartment.
The development of novel technologies for the imaging of meningeal lymphatic vessels is one of the amazing trends of biophotonics thanks to discovery of brain lymphatics over several years ago. However, there is the limited technologies exist for the study of lymphatics in vivo because lymphatic vessels are transparent with a low speed flow of lymph. Here we demonstrate the successful application of fluorescent microscopy for the imaging of lymphatic system in the mouse brain in vivo.
The main research objective is the development of innovative optical technologies for sensitive diagnosis of early stages of development of stomach cancer and monitoring of stress-induced appearance and development of tumors of the gastrointestinal tract by applying endogenous and exogenous fluorescence spectroscopy modalities.
Different mechanisms solely and in combination for evaluation of the joint impact of bioenvironmental factors (stress, Helicobacter pillory, exo-toxins in the food, water, soil and air) were applied to induce gastrointestinal tract (GIT) neoplasia in rats. The transformation of damaged areas of the stomach mucosa into malignancies in all parts of gastrointestinal tract were detected using exogenous fluorescence of photosensitizers - 5-aminolevulinic acid (5-ALA) and aluminum phthalocyanine (Al-Pc). Fluorescent mapping of different organs (liver, spleen, lungs, brain) also was developed – to evaluate the distribution of the photosensitizers in the whole body on the second hour after photosensitizer application by intravenous injection. Fiber-optic probe was used to measure the organs investigated. Fluorescence spectra were detected by microspectrometer USB4000 (OceanOptics Inc., USA), and FS405 LED source on 405 nm was used as excitation source for both types of photosensitizers applied.
Diagnostically-important parameters of oximetry, optical coherence tomography and speckle-imaging of the microcirculation of the stomach were also evaluated, to evaluate changes in the blood flow and vascular architecture, during the formation of the initial phases of the neoplasm development.
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