RESUMEN
Melanin is a crucial pigment in melanomagenesis. Its fluorescence in human tissue is exceedingly weak but can be detected through advanced laser spectroscopy techniques. The spectral profile of melanin fluorescence distinctively varies among melanocytes, nevomelanocytes, and melanoma cells, with melanoma cells exhibiting a notably "red" fluorescence spectrum. This characteristic enables the diagnosis of melanoma both in vivo and in histological samples. Neuromelanin, a brain pigment akin to melanin, shares similar fluorescence properties. Its fluorescence can also be quantified with high spectral resolution using the same laser spectroscopic methods. Documented fluorescence spectra of neuromelanin in histological samples from the substantia nigra substantiate these findings. Our research reveals that the spectral behavior of neuromelanin fluorescence mirrors that of melanin in melanomas. This indicates that the typical red fluorescence is likely influenced by the microenvironment around (neuro)melanin, rather than by direct pigment interactions. Our ongoing studies aim to further explore this distinctive "red" fluorescence. We have observed this red fluorescence spectrum in post-mortem measurements of melanin in benign nevus. The characteristic red spectrum is also evident here (unlike the benign nevus in vivo), suggesting that hypoxia may contribute to this phenomenon. Given the central role of hypoxia in both melanoma development and treatment, as well as in fundamental Parkinson's disease mechanisms, this study discusses strategies aimed at reinforcing the hypothesis that red fluorescence from (neuro)melanin serves as an indicator of hypoxia.
Asunto(s)
Melaninas , Melanoma , Espectrometría de Fluorescencia , Humanos , Hipoxia/metabolismo , Melaninas/metabolismo , Melanocitos/metabolismo , Melanoma/metabolismo , Melanoma/patología , Espectrometría de Fluorescencia/métodosRESUMEN
BACKGROUND: Caucasians with red hair and fair skin have a remarkably increased risk of malignant melanoma compared to non-redhead Caucasians. OBJECTIVES: With the aim of a reliable melanoma diagnosis in redheads, the application of dermatofluoroscopy was analyzed in 16 patients with red hair. Most of them had been included in a clinical dermatofluoroscopy study for patients with the suspicion of melanoma. We examined whether the 25 lesions of the redheads showed the same characteristic melanin fluorescence spectra for dysplastic nevi and melanomas as those of non-redhead Caucasians or whether there was a different fluorescence pattern. This is important in view of the known significantly altered ratio of eumelanin to pheomelanin in the skin of redheads. METHODS: More than 8,000 spatially resolved fluorescence spectra of 25 pigmented lesions were measured and analysed. The spectra were excited by the stepwise absorption of two 800-nm photons (principle of dermatofluoroscopy). Furthermore, the fluorescence spectra of eumelanin and pheomelanin in hair samples were determined in the same way. RESULTS: The evaluation revealed that the melanin fluorescence spectra of dysplastic nevi and melanomas of redheads have the same spectral characteristics as those of non-redhead Caucasians. An accompanying result is that dermatofluoroscopy shows identical fluorescence spectra for eumelanin and pheomelanin. CONCLUSIONS: Dermatofluoroscopy proves to be a reliable diagnostic method also for redheads. Our results also explain our recent finding that there is a uniform fluorescence spectroscopic fingerprint for melanomas of all subtypes, which is of particular interest for hypomelanotic and apparently amelanotic melanomas containing pheomelanin.
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Fluoroscopía/métodos , Color del Cabello , Melanoma/diagnóstico por imagen , Neoplasias Cutáneas/diagnóstico por imagen , Adulto , Anciano , Detección Precoz del Cáncer , Femenino , Color del Cabello/genética , Humanos , Masculino , Melanoma/etnología , Melanoma/genética , Persona de Mediana Edad , Nevo/diagnóstico por imagen , Nevo/etnología , Nevo/genética , Neoplasias Cutáneas/etnología , Neoplasias Cutáneas/genética , Población Blanca , Adulto JovenRESUMEN
In a multifaceted investigation combining local soft X-ray and vibrational spectroscopic probes with ab initio molecular dynamics simulations, hydrogen-bonding interactions of two key principal amine compounds in aqueous solution, ammonia (NH3) and ammonium ion (NH4+), are quantitatively assessed in terms of electronic structure, solvation structure, and dynamics. From the X-ray measurements and complementary determination of the IR-active hydrogen stretching and bending modes of NH3 and NH4+ in aqueous solution, the picture emerges of a comparatively strongly hydrogen-bonded NH4+ ion via N-H donating interactions, whereas NH3 has a strongly accepting hydrogen bond with one water molecule at the nitrogen lone pair but only weakly N-H donating hydrogen bonds. In contrast to the case of hydrogen bonding among solvent water molecules, we find that energy mismatch between occupied orbitals of both the solutes NH3 and NH4+ and the surrounding water prevents strong mixing between orbitals upon hydrogen bonding and, thus, inhibits substantial charge transfer between solute and solvent. A close inspection of the calculated unoccupied molecular orbitals, in conjunction with experimentally measured N K-edge absorption spectra, reveals the different nature of the electronic structural effects of these two key principal amine compounds imposed by hydrogen bonding to water, where a pH-dependent excitation energy appears to be an intrinsic property. These results provide a benchmark for hydrogen bonding of other nitrogen-containing acids and bases.
RESUMEN
SIGNIFICANCE: Seborrheic keratoses (SKs) are harmless pigmented skin lesions (PSLs) that may be confused clinically not only with other benign conditions but also with cutaneous melanoma (CM). As SKs are one of the most common neoplasms in adults, the importance of their correct diagnosis is high. Misclassifying SK as malignant is not rare and leads to a high number of unnecessary biopsies. On the other hand, misdiagnosing CM as SK may have a large impact on prognosis or therapy. AIM: In the non-invasive technique of dermatofluoroscopy, the fluorophores in melanocytes and keratinocytes are excited in vivo with nanosecond laser pulses and the resulting spectrally resolved, melanin-dominated fluorescence signals are used to differentiate between pigmented benign lesions and CM. APPROACH: In this single-center, non-interventional study, 33 PSLs of 20 patients were scanned with dermatofluoroscopy in vivo. For all included cases, dermatofluoroscopic signals were compared to pathology classification. RESULTS: The characteristic spectral features of SK were identified, where the signals are dominated by keratin, NAD(P)H, and melanin. The fluorescence spectra of SKs differed substantially from those of CM: a characteristic spectrum of SK has been identified in 27 of 28 SKs. CONCLUSIONS: The high-accuracy differential diagnosis between CM and SK is possible with dermatofluoroscopy.
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Queratosis Seborreica , Melanoma , Neoplasias Cutáneas , Adulto , Diagnóstico Diferencial , Humanos , Queratinas , Queratosis Seborreica/diagnóstico por imagen , Melaninas , Melanoma/diagnóstico por imagen , NAD , Neoplasias Cutáneas/diagnóstico por imagenRESUMEN
2-Pyridone (PD) tautomerises to 2-hydroxypyridine (HP) in liquid solution, the equilibrium of which is solvent dependent. Dimerization of PD and HP leads to the cyclic dimers (PD)(2), (HP)(2), and (PD-HP). A combined NMR and FT-IR study [Szyc, L.; et al. J. Phys. Chem. A 2010, 114, 7749-7760] has shown that solutions of 2-pyridone in CD(2)Cl(2) constitute mainly PD-CD(2)Cl(2) solute-solvent complexes and cyclic dimers (PD)(2). Because of a lack of specific marker modes, a contribution of the cyclic dimer (HP)(2) to the NH/OH stretching absorption between 2400 and 3300 cm(-1) could not be fully ruled out. Here, we present the first ultrafast infrared (IR) pump-probe experiments on the NH/OH stretching region of a solution of 2-pyridone in CD(2)Cl(2). The temporally and spectrally resolved data reveal different rate-like relaxation processes with time constants between 150 fs and 20 ps as well as coherent low-frequency oscillations due to hydrogen bond modes. An analysis shows that the transient behavior is dominated by a single hydrogen bonded species. We compare the low-frequency wavepacket motions, observed with 99 and 150 cm(-1) frequencies, with literature values as well as our quantum chemical calculations and conclude that this single molecular species is cyclic (PD)(2).
Asunto(s)
Hidrógeno/química , Piridonas/química , Dimerización , Enlace de Hidrógeno , Modelos Biológicos , Teoría Cuántica , Soluciones , Espectrofotometría InfrarrojaRESUMEN
2-Pyridone (PD), converting to 2-hydroxypyridine (HP) through a lactam-lactim isomerization mechanism, can form three different cyclic dimers by hydrogen bond formation: (PD)(2), (PD-HP), and (HP)(2). We investigate the complexation chemistry of pyridone in dichloromethane-d(2) using a combined NMR and Fourier transform infrared (FT-IR) approach. Temperature-dependent (1)H NMR spectra indicate that at low temperatures (<200 K) pyridone in solution predominantly exists as a cyclic (PD)(2) dimer, in exchange with PD monomers. At higher temperatures a proton exchange mechanism sets in, leading to a collapse of the doublet of (15)N labeled 2-pyridone. Linear FT-IR spectra indicate the existence of several pyridone species, where, however, a straightforward interpretation is hampered by extensive spectral overlap of many vibrational transitions in both the fingerprint and the NH/OH stretching regions. Two-dimensional IR correlation spectroscopy applied on concentration-dependent and temperature-dependent data sets reveals the existence of the (PD)(2) cyclic dimer, of PD-CD(2)Cl(2) solute-solvent complexes, and of PD-PD chainlike dimers. Regarding the difference in effective time scales of the NMR and FT-IR experiments, milliseconds vs (sub)picoseconds, the cyclic dimers (PD-HP) and (HP)(2), and the chainlike conformations HP-PD, may function as intermediates in reaction pathways through which the protons exchange between PD units in cyclic (PD)(2).
Asunto(s)
Modelos Químicos , Piridonas/química , Dimerización , Enlace de Hidrógeno , Isomerismo , Lactamas/química , Espectroscopía de Resonancia Magnética , Soluciones/química , Solventes/química , Espectroscopía Infrarroja por Transformada de FourierRESUMEN
Biochemical processes occur mainly in aqueous environments, where interactions with water molecules play a key role for both the structure and function of biomolecules. Deoxyribonucleic acid (DNA), the basic carrier of genetic information, is characterized by an equilibrium double helix structure which is held together by intermolecular hydrogen bonds between base pairs and hydrated by an environment of water molecules with fluctuating hydrogen bonds. Basic vibrational motions of hydrated DNA and the fastest changes in the DNA-water interactions and hydration geometries occur in less than 1 ps. These processes can be accessed by mapping the vibrational dynamics of DNA and water in a time-resolved way by nonlinear ultrafast vibrational spectroscopy. Recent studies provide a detailed understanding of DNA vibrations and their dynamics, and give insight into nonequilibrium properties and structures of hydrated DNA.
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ADN/química , Vibración , Agua/química , Factores de TiempoRESUMEN
The need for diagnosing malignant melanoma in its earliest stages results in an increasing number of unnecessary excisions. Objective criteria beyond the visual inspection are needed to distinguish between benign and malignant melanocytic tumors in vivo. Fluorescence spectra collected during the prospective, multicenter observational study ("FLIMMA") were retrospectively analyzed by the newly developed machine learning algorithm. The formalin-fixed paraffin-embedded (FFPE) tissue samples of 214 pigmented skin lesions (PSLs) from 144 patients were examined by two independent pathologists in addition to the first diagnosis from the FLIMMA study, resulting in three histopathological results per sample. The support vector machine classifier was trained on 17,918 fluorescence spectra from 49 lesions labeled as malignant (1) and benign (0) by three histopathologists. A scoring system that scales linearly with the number of the "malignant spectra" was designed to classify the lesion as malignant melanoma (score > 28) or non-melanoma (score ≤ 28). Finally, the scoring algorithm was validated on 165 lesions to ensure model prediction power and to estimate the diagnostic accuracy of dermatofluoroscopy in melanoma detection. The scoring algorithm revealed a sensitivity of 91.7% and a specificity of 83.0% in diagnosing malignant melanoma. Using additionally the image segmentation for normalization of lesions' region of interest, a further improvement of sensitivity of 95.8% was achieved, with a corresponding specificity of 80.9%.
RESUMEN
Parkinson's disease is associated with an increased risk of melanoma (and vice versa). Several hypotheses underline this link, such as pathways affecting both melanin and neuromelanin. For the first time, the fluorescence of melanin and neuromelanin is selectively accessible using a new method of nonlinear spectroscopy, based on a stepwise two-photon excitation. Cutaneous pigmentation and postmortem neuromelanin of Parkinson patients were characterized by fluorescence spectra and compared with controls. Spectral differences could not be documented, implying that there is neither a Parkinson fingerprint in cutaneous melanin spectra nor a melanin-associated fingerprint indicating an increased melanoma risk. Our measurements suggest that Parkinson's disease occurs without a configuration change of neuromelanin. However, Parkinson patients displayed the same dermatofluorescence spectroscopic fingerprint of a local malignant transformation as controls. This is the first comparative retrospective fluorescence analysis of cutaneous melanin and postmortem neuromelanin based on nonlinear spectroscopy in patients with Parkinson's disease and controls, and this method is a very suitable diagnostic tool for melanoma screening and early detection in Parkinson patients. Our results suggest a non-pigmentary pathway as the main link between Parkinson's disease and melanoma, and they do not rule out the melanocortin-1-receptor gene as an additional bridge between both diseases.
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Melaninas/metabolismo , Melanoma/patología , Enfermedad de Parkinson/metabolismo , Adulto , Anciano , Anciano de 80 o más Años , Animales , Encéfalo/patología , Estudios de Casos y Controles , Femenino , Fluorescencia , Humanos , Masculino , Persona de Mediana Edad , Cambios Post Mortem , Factores de Riesgo , Piel/patología , Pigmentación de la Piel , Sustancia Negra/patologíaRESUMEN
N-H stretching excitations of DNA oligomers containing 23 alternating adenine-thymine base pairs are studied in femtosecond two-color pump-probe experiments. For a DNA film in a zero relative humidity atmosphere, transient vibrational spectra and their time evolution up to 10 ps demonstrate negligible spectral diffusion and allow for discerning different N-H stretching bands and the O-H stretching absorption of residual water molecules. Lifetimes on the order of 0.5 ps are found for both N-H and O-H stretching modes. The time-dependent pump-probe anisotropies of the different N-H excitations point to a pronounced coupling among them, whereas the O-H stretching anisotropy remains essentially constant.
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Adenina/química , ADN/química , Oligodesoxirribonucleótidos/química , Timina/química , Secuencia de Bases , VibraciónRESUMEN
We present a compact fluorescence imaging system developed for real-time sentinel lymph node mapping. The device uses two near-infrared wavelengths to record fluorescence and anatomical images with a single charge-coupled device camera. Experiments on lymph node and tissue phantoms confirmed that the amount of dye in superficial lymph nodes can be better estimated due to the absorption correction procedure integrated in our device. Because of the camera head's small size and low weight, all accessible regions of tissue can be reached without the need for any adjustments.
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Neoplasias de la Mama/patología , Ganglios Linfáticos/patología , Imagen Óptica/instrumentación , Imagen Óptica/métodos , Biopsia del Ganglio Linfático Centinela/instrumentación , Biopsia del Ganglio Linfático Centinela/métodos , Neoplasias Cutáneas/patología , Gráficos por Computador , Simulación por Computador , Medios de Contraste/química , Difusión , Diseño de Equipo , Femenino , Humanos , Verde de Indocianina/química , Fantasmas de Imagen , Espectroscopía Infrarroja Corta , Interfaz Usuario-Computador , Agua/químicaRESUMEN
A sample cell for performing time-resolved spectroscopy on solid samples within an atmosphere of controlled vapor composition was designed and constructed. Control over vapor composition was accomplished using a combination of passive sealing and chemical agents. Performance characteristics especially well-suited to studies using femtosecond mid-infrared spectroscopy were achieved by the use of ultrathin silicon nitride windows and a rapid and reproducible sample cell exchange mechanism.
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ADN/química , Compuestos de Silicona/química , Tensoactivos/química , Absorción , Atmósfera , Diseño de Equipo , Humedad , Óptica y Fotónica , Espectrofotometría Infrarroja/métodos , Espectroscopía Infrarroja por Transformada de Fourier/métodos , Acero Inoxidable/química , Factores de TiempoRESUMEN
NH and NH(2) stretching excitations of adenine-thymine base pairs in double-stranded DNA oligomers are studied by femtosecond two-dimensional (2D) infrared spectrosopy. The 2D spectra taken for population times of up to T = 1 ps allow for separating the NH stretching mode of thymine from the symmetric and asymmetric NH(2) stretching modes of adenine and to determine their individual line shapes. The spectra demonstrate an essentially homogeneous broadening of the NH stretching band of thymine whereas the NH(2) stretching modes display a pronounced, time-independent inhomogeneous broadening, pointing to disorder in the DNA structure. We observe a (downhill) vibrational energy transfer from the asymmetric NH(2) stretching vibration of adenine at 3350 cm(-1) to the NH stretching mode of thymine at 3200 cm(-1) on a â¼500 fs time scale whereas the inverse (uphill) transfer is negligible.
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Adenina/química , ADN/química , Timina/química , Emparejamiento Base , Enlace de Hidrógeno , Espectrofotometría Infrarroja , Factores de Tiempo , VibraciónRESUMEN
Double-stranded DNA oligomers containing 23 alternating adenine-thymine base pairs are studied at different hydration levels by femtosecond two-dimensional (2D) infrared spectrosopy. Coupled NH stretching modes of the A-T pairs and OH stretching excitations of the water shell are discerned in the 2D spectra. Limited changes of NH stretching frequencies and line shapes with increasing hydration suggest spectral dynamics governed by DNA rather than water fluctuations. In contrast, OH stretching excitations of the water shell around fully hydrated DNA undergo spectral diffusion on a ~500 fs time scale. The center line slopes of the 2D spectra of hydrated DNA demonstrate a slower decay of the frequency-time correlation function (TCF) than that in neat water, as is evident from a comparison with 2D spectra of neat H(2)O and theoretical TCFs. We attribute this behavior to reduced structural fluctuations of the water shell and a reduced rate of resonant OH stretching energy transfer.
Asunto(s)
ADN/química , Agua/química , Adenina/química , Emparejamiento Base , Secuencia de Bases , Espectrofotometría Infrarroja/métodos , Timina/químicaRESUMEN
N-H stretching vibrations of hydrogen-bonded guanosine-cytidine (G·C) base pairs in chloroform solution are studied with linear and ultrafast nonlinear infrared (IR) spectroscopy. Assignment of the IR-active bands in the linear spectrum is made possible by combining structural information on the hydrogen bonds in G·C base pairs with literature results of density functional theory calculations, and empirical relations connecting frequency shifts and intensity of the IR-active vibrations. A local mode representation of N-H stretching vibrations is adopted, consisting of ν(G)(NH(2))(f) and ν(C)(NH(2))(f) modes for free NH groups of G and C, and of ν(G)(NH(2))(b), ν(G)(NH), and ν(C)(NH(2))(b) modes associated with N-H stretching motions of hydrogen-bonded NH groups. The couplings and relaxation dynamics of the N-H stretching excitations are studied with femtosecond mid-infrared two-dimensional (2D) and pump-probe spectroscopy. The N-H stretching vibrations of the free NH groups of G and C have an average population lifetime of 2.4 ps. Besides a vibrational population lifetime shortening to subpicosecond values observed for the hydrogen-bonded N-H stretching vibrations, the 2D spectra reveal vibrational excitation transfer from the ν(G)(NH(2))(b) mode to the ν(G)(NH) and/or ν(C)(NH(2))(b) modes. The underlying intermode vibrational couplings are on the order of 10 cm(-1).
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Citidina/química , Guanosina/química , Emparejamiento Base , Hidrógeno/química , Enlace de Hidrógeno , Nitrógeno/química , Soluciones/química , Espectrofotometría Infrarroja , Factores de TiempoRESUMEN
The ionic phosphate groups in the DNA backbone play a key role for DNA hydration. We study ultrafast vibrational dynamics and local interactions of phosphate groups and water by femtosecond two-color pump-probe spectroscopy. The asymmetric (PO(2))(-) stretching vibration nu(AS)(PO(2))(-) of artificial DNA oligomers containing 23 alternating adenine-thymine base pairs displays a lifetime of 340 fs, independent of the hydration level. For DNA at zero relative humidity, excess energy from the decay of the phosphate excitation is transferred within DNA on a 20 ps time scale. For fully hydrated DNA, the water shells around the phosphates serve as a primary heat sink accepting vibrational excess energy from DNA on a femtosecond time scale. OH stretching excitation of water molecules around fully hydrated DNA induces an ultrafast nu(AS)(PO(2))(-) response which includes rearrangements of the hydration shell and a reduction of the average number of phosphate-water hydrogen bonds.