FXR agonists INT-787 and OCA increase RECK and inhibit liver steatosis and inflammation in diet-induced ob/ob mouse model of NASH.

BACKGROUND AND AIMS: We have previously shown in a model of hepatic ischaemia/reperfusion injury that the farnesoid X receptor (FXR) agonist obeticholic acid (OCA) restores reversion-inducing-cysteine-rich protein with Kazal motifs (RECK), an inverse modulator of metalloproteases (MMPs) and inhibitor of the sheddases ADAM10 and ADAM17 involved in inflammation and fibrogenesis. Here, the effects of FXR agonists OCA and INT-787 on hepatic levels of RECK, MMPs, ADAM10 and ADAM17 were compared in a diet-induced ob/ob mouse model of non-alcoholic steatohepatitis (NASH). METHODS: Lep ob/ob NASH mice fed a high-fat diet (HFD) or control diet (CD) for 9 weeks (wks) were treated with OCA or INT-787 0.05% dosed via HFD admixture (30 mg/kg/day) or HFD for further 12 wks. Serum alanine transaminase (ALT) and inflammatory cytokines, liver RECK, MMP-2 and MMP-9 activity as well as ADAM10, ADAM17, collagen deposition (Sirius red), hepatic stellate cell activation (α-SMA) and pCK+ reactive biliary cells were quantified. RESULTS: Only INT-787 significantly reduced serum ALT, IL-1ß and TGF-ß. A downregulation of RECK expression and protein levels observed in HFD groups (at 9 and 21 wks) was counteracted by both OCA and INT-787. HFD induced a significant increase in liver MMP-2 and MMP-9; OCA administration reduced both MMP-2 and MMP-9 while INT-787 markedly reduced MMP-2 expression. OCA and INT-787 reduced both ADAM10 and ADAM17 expression and number of pCK+ cells. INT-787 was superior to OCA in decreasing collagen deposition and α-SMA levels. CONCLUSION: INT-787 is superior to OCA in controlling specific cell types and clinically relevant anti-inflammatory and antifibrotic molecular mechanisms in NASH.

Fatty Acids and Bilirubin as Intrinsic Autofluorescence Serum Biomarkers of Drug Action in a Rat Model of Liver Ischemia and Reperfusion.

The autofluorescence of specific fatty acids, retinoids, and bilirubin in crude serum can reflect changes in liver functional engagement in maintaining systemic metabolic homeostasis. The role of these fluorophores as intrinsic biomarkers of pharmacological actions has been investigated here in rats administered with obeticholic acid (OCA), a Farnesoid-X Receptor (FXR) agonist, proven to counteract the increase of serum bilirubin in hepatic ischemia/reperfusion (I/R) injury. Fluorescence spectroscopy has been applied to an assay serum collected from rats submitted to liver I/R (60/60 min ± OCA administration). The I/R group showed changes in the amplitude and profiles of emission spectra excited at 310 or 366 nm, indicating remarkable alterations in the retinoid and fluorescing fatty acid balance, with a particular increase in arachidonic acid. The I/R group also showed an increase in bilirubin AF, detected in the excitation spectra recorded at 570 nm. OCA greatly reversed the effects observed in the I/R group, confirmed by the biochemical analysis of bilirubin and fatty acids. These results are consistent with a relationship between OCA anti-inflammatory effects and the acknowledged roles of fatty acids as precursors of signaling agents mediating damaging responses to harmful stimuli, supporting serum autofluorescence analysis as a possible direct, real-time, cost-effective tool for pharmacological investigations.

Autofluorescent Biomolecules in Diptera: From Structure to Metabolism and Behavior.

Light-based phenomena in insects have long attracted researchers' attention. Surface color distribution patterns are commonly used for taxonomical purposes, while optically-active structures from Coleoptera cuticle or Lepidoptera wings have inspired technological applications, such as biosensors and energy accumulation devices. In Diptera, besides optically-based phenomena, biomolecules able to fluoresce can act as markers of bio-metabolic, structural and behavioral features. Resilin or chitinous compounds, with their respective blue or green-to-red autofluorescence (AF), are commonly related to biomechanical and structural properties, helpful to clarify the mechanisms underlying substrate adhesion of ectoparasites' leg appendages, or the antennal abilities in tuning sound detection. Metarhodopsin, a red fluorescing photoproduct of rhodopsin, allows to investigate visual mechanisms, whereas NAD(P)H and flavins, commonly relatable to energy metabolism, favor the investigation of sperm vitality. Lipofuscins are AF biomarkers of aging, as well as pteridines, which, similarly to kynurenines, are also exploited in metabolic investigations. Beside the knowledge available in Drosophila melanogaster, a widely used model to study also human disorder and disease mechanisms, here we review optically-based studies in other dipteran species, including mosquitoes and fruit flies, discussing future perspectives for targeted studies with various practical applications, including pest and vector control.

The Bright Side of the Tiger: Autofluorescence Patterns in Aedes albopictus (Diptera, Culicidae) Male and Female Mosquitoes.

Light-based events in insects deserve increasing attention for various reasons. Besides their roles in inter- and intra-specific visual communication, with biological, ecological and taxonomical implications, optical properties are also promising tools for the monitoring of insect pests and disease vectors. Among these is the Asian tiger mosquito, Aedes albopictus, a global arbovirus vector. Here we have focused on the autofluorescence characterization of Ae. albopictus adults using a combined imaging and spectrofluorometric approach. Imaging has evidenced that autofluorescence rises from specific body compartments, such as the head appendages, and the abdominal and leg scales. Spectrofluorometry has demonstrated that emission consists of a main band in the 410-600 nm region. The changes in the maximum peak position, between 430 nm and 500 nm, and in the spectral width, dependent on the target structure, indicate the presence, at variable degrees, of different fluorophores, likely resilin, chitin and melanins. The aim of this work has been to provide initial evidence on the so far largely unexplored autofluorescence of Ae. albopictus, to furnish new perspectives for the set-up of species- and sex-specific investigation of biological functions as well as of strategies for in-flight direct detection and surveillance of mosquito vectors.

Spectrofluorometric Analysis of Autofluorescing Components of Crude Serum from a Rat Liver Model of Ischemia and Reperfusion.

Autofluorescence (AF) of crude serum was investigated with reference to the potential of its intrinsic AF biomarkers for the noninvasive diagnosis of liver injury. Spectral parameters of pure compounds representing retinol (vitamin A) and fluorescing free fatty acids were characterized by spectrofluorometry, to assess spectral parameters for the subsequent AF analysis of serum, collected from rats undergoing liver ischemia/reperfusion (I/R). Differences in AF spectral profiles detected between control and I/R were due to the increase in the AF components representing fatty acids in I/R serum samples. No significant changes occurred for retinol levels, consistently with the literature reporting that constant retinol levels are commonly observed in the blood, except for malnutrition or chronic severe liver disease. Conversely, fatty acids, in particular arachidonic and linoleic acid and their derivatives, act as modulating agents in inflammation, representing both a protective and damaging response to stress stimuli. The biometabolic and pathophysiological meaning of serum components and the possibility of their direct detection by AF spectrofluorometry open up interesting perspectives for the development of AF serum analysis, as a direct, cost effective, supportive tool to assess liver injury and related systemic metabolic alterations, for applications in experimental biomedicine and foreseen translation to the clinics.

Ozone Treatment of Grapes During Withering for Amarone Wine: A Multimodal Imaging and Spectroscopic Analysis.

The production of Amarone wine is governed by a disciplinary guideline to preserve its typical features; however, postharvest infections by the fungus Botrytis cinerea (B. cinerea) not only represent a phytosanitary problem but also cause a significant loss of product. In this study, we tested a treatment with mild ozoniztion on grapes for Amarone wine production during withering in the fruttaio (the environment imposed by the disciplinary guideline) and evaluated the impact on berry features by a multimodal imaging approach. The results indicate that short and repeated treatments with low O3 concentrations speed up the naturally occurring berry withering, probably inducing a reorganization of the epicuticular wax layer, and inhibit the development of B. cinerea, blocking the fungus in an intermediate vegetative stage. This pilot study will pave the way to long-term research on Amarone wine obtained from O3-treated grapes.

Serum and Hepatic Autofluorescence as a Real-Time Diagnostic Tool for Early Cholestasis Assessment.

While it is well established that various factors can impair the production and flow of bile and lead to cholestatic disease in hepatic and extrahepatic sites, an enhanced assessment of the biomarkers of the underlying pathophysiological mechanisms is still needed to improve early diagnosis and therapeutic strategies. Hence, we investigated fluorescing endogenous biomolecules as possible intrinsic biomarkers of molecular and cellular changes in cholestasis. Spectroscopic autofluorescence (AF) analysis was performed using a fiber optic probe (366 nm excitation), under living conditions and in serum, on the livers of male Wistar rats submitted to bile duct ligation (BDL, 24, 48, and 72 h). Biomarkers of liver injury were assayed biochemically. In the serum, AF analysis distinctly detected increased bilirubin at 24 h BDL. A continuous, significant increase in red-fluorescing porphyrin derivatives indicated the subversion of heme metabolism, consistent with an almost twofold increase in the serum iron at 72 h BDL. In the liver, changes in the AF of NAD(P)H and flavins, as well as lipopigments, indicated the impairment of mitochondrial functionality, oxidative stress, and the accumulation of oxidative products. A serum/hepatic AF profile can be thus proposed as a supportive diagnostic tool for the in situ, real-time study of bio-metabolic alterations in bile duct ligation (BDL) in experimental hepatology, with the potential to eventually translate to clinical diagnosis.

Autofluorescence of liver tissue and bile: organ functionality monitoring during ischemia and reoxygenation.

BACKGROUND AND OBJECTIVE: Autofluorescence (AF) based optical biopsy of liver tissue is a powerful approach for the real-time diagnosis of its functionality. Since increasing attention is given to the bile production and composition to monitor the liver metabolic engagement in surgery and transplantation, we have investigated the bile AF properties as a potential, additional diagnostic parameter. STUDY DESIGN/MATERIALS AND METHODS: Spectrofluorometric analysis has been performed in real time on a rat liver model of warm ischemia and reperfusion-60 minutes partial portal vein and hepatic artery clamping and subsequent restoration of blood circulation-in comparison with sham operated rats. The AF spectra have been recorded through a single fiber optic probe (366 nm excitation) from both liver tissue and bile, collected from the cannulated bile duct, and analyzed by means of curve fitting procedures. Bile composition has been also analyzed through biochemical assays of bilirubin, total bile acids (TBA) and proteins. RESULTS: Both liver and bile AF signal amplitude and spectral shape undergo changes during induction of ischemia and subsequent reperfusion. The liver tissue response is mainly ascribable to changes in NAD(P)H and flavins and their redox state, largely dependent on oxygen supply, and to the decrease of both vitamin A and fatty acid AF contributions. During comparable times, sham operated rat livers undergo smaller alterations in AF spectral shape, indicating a continuous, slight increase in the oxidized state. Bile AF emission shows a region in the 510-600 nm range ascribable to bilirubin, and resulting from the contribution of two bands, centered at about 515-523 and 570 nm, consistently with its bichromophore nature. Variations in the balance between these two bands depend on the influence of microenvironment on bilirubin intramolecular interchromophore energy transfer efficiency and are likely indicating alteration in a bile composition. This event is supported also by changes observed in the 400-500 nm emission region, ascribable to other bile components. CONCLUSIONS: In parallel with the intratissue AF properties, mainly reflecting redox metabolic activities, the bile AF analysis can provide additional information to assess alterations and recovery in the balance of liver metabolic activities.

Autofluorescence properties of murine embryonic stem cells during spontaneous differentiation phases.

BACKGROUND AND OBJECTIVE: The autofluorescence (AF) analysis allows in vivo, real-time assessment of cell functional activities, depending on the presence of biomolecules strictly involved in metabolic reactions and acting as endogenous fluorophores. Pluripotent stem cells during differentiation are known to undergo changes in their morphofunctional properties, with particular reference to bioenergetic metabolic signatures involving endogenous fluorophores such as NAD(P)H, flavins, lipofuscin-like lipopigments. Since the development of regenerative therapies based on pluripotent cells requires a careful monitoring of the successful maturation into the desired phenotype, aim of our work is to evaluate the AF potential to assess the differentiation phases in a murine stem cell model. STUDY DESIGN/MATERIALS AND METHODS: Mouse embryonic stem cells (ESCs) maintained with and without leukemia inhibitory factor (LIF), embryoid bodies (EBs), and EB-derived cells undergoing spontaneous differentiation toward the hematopoietic lineage have been used as a sample models. Cell AF properties have been characterized upon 366-nm excitation, under living conditions and in the absence of exogenous markers. Imaging, microspectrofluorometric techniques, and spectral fitting analysis based on the spectral parameters of each endogenous fluorophore have been applied to estimate their contribution to the whole cell AF emission spectra. Specific cytochemical labeling has been performed to validate AF data. RESULTS: Depending on the differentiation phases, cells undergo changes in morphology, AF distribution patterns, and AF emission spectral profiles. These latter reflect variations in the single endogenous fluorophore contribution to the overall emission. The coenzyme NAD(P)H accounts for up to 80% of the whole spectral area. The free form prevails on the bound one, and their changes have been investigated in terms of NAD(P)Hbound/free and redox ratios. These values vary in agreement with a slow metabolic activity and prevailing glycolytic metabolism in the undifferentiated HM1 cells, an increased metabolic activity still relying on glycolysis during the early differentiation phases, and an increased oxidative phosphorylation in EB and hematopoietic precursor cells. Lipofuscin-like lipopigments decrease following differentiation, and porphyrins contributing for less than 5%, prevail in the more actively differentiating cells. These results reflect the shift between anaerobic and aerobic respiration following differentiation, consistently with a decreased autophagy of cell organelles (i.e., mitochondria, as a strategy reported in the literature to keep the undifferentiated homeostasis state), higher mitochondrial activity with more numerous NADH binding sites and synthesis of heme as prosthetic group of proteins, that is, cytochromes. CONCLUSIONS: These data open promising perspectives for the monitoring of stem cells differentiation under living conditions without labeling with exogenous agents (inducing perturbations when used in vivo), or immunomarkers not always available for veterinary and zootechnics, by exploiting endogenous fluorophores as intrinsic biomarkers of cell morphofunctional changes.

Autofluorescence Label-Free Imaging of the Liver Reticular Structure.

Autofluorescence rising from biological substrates under proper excitation light depends on the presence of specific endogenous fluorophores and can provide information on the morpho-functional properties in which they are strictly involved. Besides the numerous endogenous fluorophores involved in metabolic functions, fibrous proteins may act as direct, label-free biomarkers of the tissue structural organization. The optical properties of collagen, in particular, are currently applied as an alternative to established histochemical procedures to investigate the connective tissue as well as its changes in diseased conditions. This is particularly true in hepatology where the histochemical procedures to label the reticular structure are not routinely applied, as they are complex and time-consuming. The morphology of the liver reticular structure and its changes are up to now poorly considered despite the increasing awareness of the regulatory role played by the remodeling of the reticular structure in pathological conditions. In this context, the autofluorescence label-free imaging has proven to be a suitable approach.

Fluorescence excitation properties of bilirubin in solution and in serum.

The bichromophore nature of bilirubin explains the presence of at least two partially overlaying bands in both absorption and fluorescence emission spectra, and accounts for interchromophore exciton transfer events responsible for the emission sensitivity to the molecular environment and excitation wavelength. These concepts were likely responsible for the previously reported good yield of the unexpected remarkable bilirubin fluorescence emission under excitation at 366 nm, at which bilirubin absorption is very low. In this connection, aim of this work is to further investigate bilirubin spectral excitation properties and their diagnostic potential, until now poorly considered. Fluorescence excitation spectra of pure bilirubin in solution with solubilizing agents observed at 520 and 570 nm showed a wide region in the 430-510 nm range, similar to the absorption profile. In addition, an excitation band centered at about 400 nm was detected. Comparable excitation features were detected in rat serum. The 430-510 nm excitation region was well separated from a main band at shorter wavelength, ascribable to other endogenous fluorophores, with a shoulder at about 400 nm which was also easily discriminated by spectral fitting analysis. The bands ascribable to bilirubin showed changes of their relative contribution to the overall spectral region after liver ischemia/reperfusion, comparable to bilirubin biochemical data. Excitation spectra proved to discriminate the fluorescence of serum bilirubin at levels much lower than emission spectra, opening promising perspectives to improve the real time fluorescence analysis of crude serum in the absence of any exogenous labelling agent, and advance the diagnostic application of optical-biopsy in experimental hepatology and biomedicine.

Human liver autofluorescence: an intrinsic tissue parameter discriminating normal and diseased conditions.

BACKGROUND AND OBJECTIVE: Autofluorescence (AF) emission is an intrinsic parameter that can provide real-time information on morpho-functional properties of biological tissue, being strictly related with their biochemical composition and structural organization. The diagnostic potentials of AF-based techniques have been investigated on normal, fibrotic, and steatotic liver tissues, in reference to histological features as evidenced by specific histochemical stainings. MATERIALS AND METHODS: AF emission under excitation at 366 nm has been examined on cryostatic tissue sections obtained from biopsies collected during surgical operation, by means of fluorescence imaging and microspectrofluorometric techniques. RESULTS: NAD(P)H, collagen, and vitamin A were found to be the endogenous fluorophores characterizing normal, fibrotic, and steatotic liver tissue AF, respectively. The differences of their photo-physical properties, in terms of emission amplitude, spectral shape, and response to irradiation, give rise to modifications of overall AF signal collected from tissues that allow the liver conditions to be distinguished. CONCLUSION: The study provides a valid premise for a development of AF-based optical biopsy techniques for a real-time discrimination of liver anatomo-pathological patterns.

Autofluorescence Spectroscopy for Monitoring Metabolism in Animal Cells and Tissues.

Excitation of biological substrates with light at a suitable wavelength can give rise to a light emission in the ultraviolet (UV)-visible, near-infrared (IR) spectral range, called autofluorescence (AF). This is a widespread phenomenon, ascribable to the general presence of biomolecules acting as endogenous fluorophores (EFs) in the organisms of the whole life kingdom. In cytochemistry and histochemistry, AF is often an unwanted signal enhancing the background and affecting in particular the detection of low signals or rare positive labeling spots of exogenous markers. Conversely, AF is increasingly considered as a powerful diagnostic tool because of its role as an intrinsic biomarker directly dependent on the nature, amount, and microenvironment of the EFs, in a strict relationship with metabolic processes and structural organization of cells and tissues. As a consequence, AF carries multiple information that can be decrypted by a proper analysis of the overall emission signal, allowing the characterization and monitoring of cell metabolism in situ, in real time and in the absence of perturbation from exogenous markers. In the animal kingdom, AF studies at the cellular level take advantage of the essential presence of NAD(P)H and flavins, primarily acting as coenzymes at multiple steps of common metabolic pathways for energy production, reductive biosynthesis and antioxidant defense. Additional EFs such as vitamin A, porphyrins, lipofuscins, proteins, and neuromediators can be detected in different kinds of cells and bulk tissues, and can be exploited as photophysical biomarkers of specific normal or altered morphofunctional properties, from the retinoid storage in the liver to aging processes, metabolic disorders or cell transformation processes. The AF phenomenon involves all living system, and literature reports numerous investigations and diagnostic applications of AF, taking advantage of continuously developing self-assembled or commercial instrumentation and measuring procedures, making almost impossible to provide their comprehensive description. Therefore a brief summary of the history of AF observations and of the development of measuring systems is provided, along with a description of the most common EFs and their metabolic significance. From our direct experience, examples of AF imaging and microspectrofluorometric procedures performed under a single excitation in the near-UV range for cell and tissue metabolism studies are then reported.

Fluorescing fatty acids in rat fatty liver models.

The autofluorescence (AF) of NAD(P)H and flavins has been at the basis of many in-situ studies of liver energy metabolism and functionality. Conversely, few data have been so far reported on fluorescing lipids. In this work we investigated the AF of liver lipid extracts from two fatty liver models, Wistar rats fed with MCD diet for 12 days (Wi-MCD), and obese (fa/fa) Zucker rats. Among the most abundant fatty acids in the lipid extracts, indicated by mass spectrometry, arachidonic acid (AA) exhibited higher quantum yield than the other fluorescing fatty acids (FLFA), and red shifted AF spectrum. This allowed to estimate the AA contribution to the overall emission of lipid extracts by curve fitting analysis. AA prevailed in obese Zucker livers, accounting for the different AF spectral profiles between the two models. AF and mass spectrometry indicated also a different balance between the fluorescing fraction and the overall amount of AA in the two models. The ability of AF to detect directly AA and FLFA was demonstrated, suggesting its supportive role as tool in wide-ranging applications, from the control of animal origin food, to experimental investigations on liver fat accumulation, lipotoxicity and disease progression, with potential translation to the clinics.

NAD(P)H and Flavin Autofluorescence Correlation with ATP in Rat Livers with Different Metabolic Steady-State Conditions.

The monitoring of NAD(P)H and flavin autofluorescence (AF) is at the basis of numerous investigations on energy metabolism. Nevertheless, the ability of these AF biomarkers to accurately represent the energy currency, ATP, is poorly explored. Here, we focused on the AF/ATP correlation in lean and fatty livers with different steady-state metabolic conditions, achieved after organ isolation, preservation and recovery, in a likely dependence on both liver intrinsic metabolic features and externally induced perturbations. Within these eventual, various conditions, a significant correlation was detected between liver NAD(P)H and flavin AF, measured via fiber-optic probe, and biochemical ATP data, strengthening AF as biomarker of energy metabolism in steady-state conditions for wide-ranging experimental and diagnostic applications.

Autofluorescence discrimination of metabolic fingerprint in nutritional and genetic fatty liver models.

Liver tissue autofluorescence (AF) has been characterized in two models with a different potential to undergo disease progression to steatohepatitis: Wistar rats, administered with a methionine, choline deficient diet (MCD), and Zucker (fa/fa) rats, homozygous for a spontaneous mutation of leptin receptor. AF spectra were recorded from liver tissue cryostatic sections by microspectrofluorometry, under 366nm excitation. Curve fitting analysis was used to estimate the contribution of different endogenous fluorophores (EFs) to the overall AF emission: i) fluorescing fatty acids, a fraction of liver lipids up to now poorly considered and complicated to detect by conventional procedures; ii) lipofuscin-like lipopigments, biomarkers of oxidizing events; iii) NAD(P)H and flavins, biomarkers of energy metabolism and tissue redox state. AF data and biochemical correlates of hepatocellular injury resulted to depend more on rat strain than on intratissue bulk lipid or ROS levels, reflecting a different metabolic ability of the two models to counteract potentially harmful agents. AF analysis can thus be proposed for extensive applications ranging from experimental hepatology to the clinics. AF based diagnostic procedures are expected to help both the prediction of the risk of fatty liver disease progression and the prescreening of marginal organs to be recruited as donors for transplantation. A support is also foreseen in the advancement and personalization of strategies to ameliorate the donor organ preservation outcome and the follow up of therapeutic interventions.

In vivo autofluorescence spectrofluorometry of central serotonin.

The autofluorescence properties of serotonin (5-HT) were investigated by light spectrofluorometry in in vitro, ex vivo and in vivo experiments. Ex vivo samples were prepared from rat brain regions containing serotonin (5-HT) i.e. cortex, striatum, hippocampus. Rats were untreated (controls) or previously submitted to chronic behavioural or pharmacological treatments known to affect endogenous 5-HT levels. Autofluorescence analysis (excitation: 366 nm) on hippocampus homogenates supplied with exogenous 5-HT revealed spectral alterations attributable to changes of endogenous 5-HT levels. In vivo, real time fluorescence studies were performed via a 50 microm diameter optic fiber probe stereotaxically implanted into selected brain areas of anaesthetised rats treated with fluoxetine or 5-OH-tryptophan. All autofluorescence data were consistent with those obtained in parallel experiments performed with ex vivo or in vivo voltammetry, confirming that auto-fluorescence spectroscopy is a suitable technique for the direct assessment of fluorescent neurotransmitters. This is a reliable evidence of the in vivo application of spectroscopy together with optic fiber probe for in vivo, in situ and real time measurement of 5-HT in discrete brain areas.

Diagnostic potential of autofluorescence for an assisted intraoperative delineation of glioblastoma resection margins.

The intrinsic autofluorescence properties of biological tissues can be affected by the occurrence of histological and biochemical alterations induced by pathological processes. In this study the potential of autofluorescence to distinguish tumor from normal tissues was investigated with the view of a real-time diagnostic application in neurosurgery to delineate glioblastoma resection margins. The autofluorescence properties of nonneoplastic and neoplastic tissues were analyzed on tissue sections and homogenates by means of a microspectrofluorometer, and directly on patients affected by glioblastoma multiforme, during surgery, with a fiber-optic probe. Scan-microspectrofluorometric analysis on tissue sections evidenced a reduction of emission intensity and a broadening of the main emission band, along with a redshift of the peak position, from peritumoral nonneoplastic to neoplastic tissues. Differences in both spectral shape and signal amplitude were found in patients when the glioblastoma lesion autofluorescence was compared with those of cortex and white matter taken as healthy tissues. Both biochemical composition and histological organization contribute to modify the autofluorescence emission of neoplastic, with respect to nonneoplastic, brain tissues. The differences found in the in vivo analysis confirm the prospects for improving the efficacy of tumor resection margin delineation in neurosurgery.

Bilirubin: an autofluorescence bile biomarker for liver functionality monitoring.

Excitation at 366-465 nm of bilirubin in aqueous solution with solubilizing agents results in emission spectra composed by two main bands. The variation of their relative contributions as shown by changes in the spectral shape are consistent with the bilirubin bichromophore nature. This latter accounts for an exciton-coupling phenomenon, intramolecular interchromophore energy transfer efficiency being affected by microenvironment. Excitation at 366 nm, despite the poor absorption of bilirubin, gives rise to appreciable emission signals from both pure compounds and bile - collected from functionally altered rat livers - favouring the spectral shape response to environment and molecular conformation changes. As compared to the merely bile flow estimation, real-time detection of fluorescence, revealing composition variations, improves near-UV optical-biopsy diagnostic potential in hepatology.