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PubMed1994

[Inhibitory effect of running exercise on age-induced muscle atrophy].

The effects of a 10-week running exercise on the histochemical and morphological properties of muscle fibers in the soleus (SOL) and tibialis anterior (TA) muscles and also in spinal motoneurons were investigated in 4 groups of female rats. Animals of the control groups were 20, 90 and 100 weeks old. Animals of the experimental group were 100 weeks old, and were exercised on a running wheel for 10 weeks. Muscle fibers examined were classified as slow-twitch oxidative (SO), fast-twitch oxidative glycolytic (FOG) or fast-twitch glycolytic (FG) according to their contractile and metabolic profiles. Spinal motoneurons innervating the SOL and TA were identified by retrograde neuronal labelling of fluorescent dye, nuclear yellow. Oxidative enzyme activities of motoneurons in the neuron pool were examined by microspectrophotometry. When compared to 20 week old rats, a decrease in the number of FOG in the SOL and of FG in the TA was observed in 90 and 100 week old rats. Findings of atrophy were observed in SO and FOG of the SOL and in FOG and FG of the TA in 90 to 100 week old rats. Practically no changes were observed with aging in the number and oxidative capacity of motoneurons in the SOL neuron pool. However, a decrease with aging was observed in the number and oxidative capacity of motoneurons in the TA neuron pool. From these results it is indicated that exercise inhibits the age-induced atrophy of SO fibers in the SOL.

Aging↗
PubMed1976

[Structural and metabolic changes occuring during the contractile activity of isolated muscle fibers; microspectrophotometric study with polarized light of the ultraviolet region].

The use of a technique of in vivo microspectrophotometry with ultraviolet polarized light on frog myocardial trabeculae or isolated fibers has indicated an increase in the fraction of oriented aromatic aminoacids and a decrease in the fraction of bounded nucleotides, during the contractile event.

Adenosine Triphosphate↗
PubMed1995

[Histochemistry of the organs--application of histochemistry to anatomy].

Histochemistry has been developed as a science to localize chemical components of cells and tissues on histological sections by using chemical, physical and biological techniques. Many textbooks are available dealing with the methodology of histochemistry, consisting of fixation of tissues, demonstration of biochemical components such as nucleic acids, proteins, sugars, lipids, enzymes, pigments, inorganic substances and amines by means of chemical reactions (Fig. 5) or application of physical (Figs. 1-3, 6-13) and biological (Fig. 4) methods such as spectrophotometry, X-ray microanalysis, radioautography and immunohistochemistry. The methodology has been well developed to form a new science which should be given a designation "general histochemistry" as similar to general histology. On the other hand, these techniques should be applied to all the organ systems, i.e. the skeletal, muscular, circulatory, digestive (Figs. 1-6), respiratory (Figs. 7, 8), urinary (Fig. 9), genital (Fig. 10), endocrine (Fig. 11), nervous (Fig. 12), and sensory organs (Fig. 13). The results of these applications to all the organs should be collected and construct a new field of histochemistry of the organs like "histology of the organs". These results now form a new field of science which should be designated as "special histochemistry". Since the present author started the career as an anatomist and histologist in 1955, the author first aimed at the study of histochemistry by developing new chemical, physical and biological techniques such as enzyme histochemistry (Fig. 5), microspectrophotometry, autoradiography (Figs. 1-3, 6-13), cryo-techniques, immunohistochemistry (Fig. 4) and lectins in the 1950s, 1960s and 1970s. However, the author and coworkers made efforts to apply these techniques to various organ systems of humans and animals since the 1970s to the present time and collected data from all the organs including the aging process from embryos, neonates, sucklings, weanlings, adults and to senescence. The data include not only 3-dimensional structures of the organs but also the 4-dimensional features taking the time dimension into account using the radioautographic techniques by labeling cells and tissues in connection with the time lapse. The original papers published from our laboratory during these 40 years were reviewed and compiled to form a new field of science, which should be designated as histochemistry of the organs or, alternatively, special histochemistry. It is hoped that this new prospect would be well developed to form a novel system in medical science.

Aging↗
PubMed1977

[DNA concentration in the nuclei of endometrial gland epithelium in the presence of dyshormonal hyperplasias of the climacteric period].

The DNA content in the nuclei of the glandular epithelium of the endometrium was studied by microspectrophotometry in 6 women of fertile age and 10 with the climacteric dyshormonal adenocystic hyperplasia. In normal uterine mucosa the DNA quantity remained unchanged at various phases of the menstrual cycle. As compared to the normal, in dyshormonal endometrial hyperplasia the DNA content was almost doubled; there was also a moderate increase of polyploidy.

Cell Nucleus↗
PubMed1997

Visual pigment assignments in regenerated retina.

Retinas of adult teleost fish can regenerate after injury. Two important issues regarding this phenomenon are the assembly of the regenerated retina and the neuronal images of the visual scene that the regenerated retina produces. Here we report experiments in which the visual pigment content of photoreceptors derived from native and regenerated sunfish retinas was determined by microspectrophotometry. In native retina, there is an apparently perfect correspondence between cone morphology and visual pigment content; all rods contain a middle-wavelength pigment, all single cones contain a different middle-wavelength pigment, and all double cone members contain a long-wavelength pigment. The visual pigments in regenerated rods and double cones were the same as in native retina; however, triple cones, a morphology never observed in native retina, contained the long-wavelength pigment. Moreover, although approximately 60% of regenerated single cones contained the expected middle-wavelength pigment, all other single cones contained the long-wavelength pigment. This mismatch between morphology of regenerated single cones and their visual pigment assignment indicated the following: (1) There is a degree of independence between the mechanisms that establish cone morphology and pigment content during regeneration, which suggests that cone photoreceptor regeneration is not a straightforward recapitulation of the normal cone photoreceptor developmental plan. (2) Although anomalous, the long-wavelength single cones may enable regenerated retina to restore the native spectral sampling of the visual scene.

Animals↗
PubMed1976

[Cytological, bioptical and cytophotometric studies on tumors of the breast (author's transl)].

209 mammary biopsies were evaluated by cytological and histological methods. 23 of them were analysed by microspectrophotometry. 9,5% of the cases could be classified as Papanicolaou III. At one hand the mammary carcinomas of this group were underestimated, on the other hand fibroadenomas and hyperplastic lesions of cystic disease of the breast were overevaluated. The DNA-histograms of these fibroadenomas and hyperplastic lesions reflect closely the tendency of proliferation of epithelium and correspond with the cytomorphological findings, a reliable differentiation of these diseases on the basis of a tumor-histogram is not possible. False negative tumor histograms are particularly found in high differentiated carcinomas. In the other groups the cytological findings were in agreement with histological examination in 92-98% of the cases.

Adenocarcinoma↗
PubMed1999

Characterization of intracellular pH gradients in human multidrug-resistant tumor cells by means of scanning microspectrofluorometry and dual-emission-ratio probes.

Multidrug-resistant cells are believed to contain a plasma-membrane-efflux pump which is hypothesized to expel anticancer drugs from the cytosol to the cell exterior. Many of these drugs are classified as weak bases whose binding to intracellular targets is pH-dependent. Slight alterations in intracellular pH gradients have been shown to affect accumulation, endocytosis and secretion of drugs. In this study, we developed a new method based on confocal spectral imaging analysis to determine intracellular pH gradients in sensitive and MDR tumor cells. Fluorescein isothiocyanate (FITC) and tetramethylrhodamine conjugated to dextran (FRD) and SNAFL-calcein-AM were used to determine pH in acidic compartments. Carboxy-SNARF1-AM was used to examine cytosolic pH. We observed that sensitive (HL60, K562, CEM and MCF7) cells exhibit lower acidity of the subcellular organelles than that corresponding to drug-resistant derivatives. Moreover, results obtained with carboxy-SNARF1-AM show that resistant cells display a more alkaline cytosolic pH. This results in a considerably larger pH gradient between the vesicular compartments and the cytosol of resistant cells than of sensitive cells. The lower pH gradient observed in sensitive cells may be related to a disruption in the organization of the trans-Golgi network (TGN). In drug-resistant cells, the organization of TGN appears compact. In addition, confocal microscopic analysis of cells labelled with FRD and SNAFL-calcein showed that sensitive cells contain a lower number of acidified vesicles. This suggest a diminished capacity of these cells to remove protonated drugs from the cytoplasm to secretory compartments followed by their secretion through the activity of the secretory and recycling pathways.

Breast Neoplasms↗
PubMed1994

Role of DNA ploidy patterns in esophageal squamous cell carcinoma. An ultraviolet microspectrophotometric study.

BACKGROUND: Controversy still exists about the influence of DNA content on the prognosis for patients with esophageal squamous cell carcinoma. METHODS: DNA ploidy was determined by microspectrophotometric (MSPM) analysis of paraffin embedded malignant tissue from 78 patients with squamous cell carcinoma of the esophagus. The DNA distribution pattern was classified as diploid, low grade aneuploid (LGA), and high grade aneuploid (HGA) pattern. The relationships among DNA distribution patterns, pathologic features, clinical findings, and prognoses were investigated. Twenty-seven of 78 patients were also selected for analyses of the DNA ploidy pattern of lymph node metastases. RESULTS: Of 78 cancers, 20 (26%) were diploid, 15 (19%) LGA, and 43 (55%) HGA. The advanced carcinomas had the higher distribution of HGA pattern (55%), whereas the diploid pattern was more frequent in early stage (70%) or superficial (65%) esophageal cancer. In patients with the HGA pattern, there was a significantly higher frequency of lymph node metastases (79%) and marked lymphatic (74.4%) and vascular (69.8%) invasion compared with those exhibiting the LGA (33.3%) or diploid (25%) pattern. Five-year survival rates for patients with diploid carcinoma (57%) was significantly better than for those with the HGA pattern (14%). The distribution of nuclear DNA content was much narrower in the metastatic lymph node than in the corresponding primary cases. CONCLUSIONS: The HGA of DNA pattern based on spectrophotometry closely correlated with the factors generally indicative of the aggressive behavior of malignant tumors. Therefore, the role of the DNA ploidy pattern as a prognostic factor was emphasized.

Aged↗
PubMed2000

Infrared spectroscopy of human cells and tissue. VIII. Strategies for analysis of infrared tissue mapping data and applications to liver tissue.

Experimental and computational methods of infrared microspectroscopy (IRI-MSP) and infrared spectral mapping (ISM) are presented. These methods are subsequently applied to the analysis of cirrhotic liver tissue. The sensitivity of infrared spectral mapping toward spectral changes caused by disease will be demonstrated. In addition, the excellent agreement between ISM data and histopathological information will be discussed.

Humans↗
PubMed2001

Effects of low-dose, noncytotoxic, intraarticular liposomal clodronate on development of erosions and proteoglycan loss in established antigen-induced arthritis in rabbits.

OBJECTIVE: To assess the clinical and histologic effects of an intraarticular application of low-dose (non-cytotoxic) liposomal clodronate in established antigen-induced monarthritis (AIA) in rabbits. METHODS: AIA was monitored by assessments of joint swelling, C-reactive protein levels, and radiographic changes in 17 NZW rabbits for 8 weeks during the course of weekly intraarticular injections of liposomal clodronate (0.145 mg/injection, low dose) or "empty" liposomes. The contralateral knee was injected with liposome buffer alone as the control. End-point analyses included macroscopic joint examination, immuno- and TUNEL staining, Safranin O staining/microspectrophotometry, and tumor necrosis factor alpha (TNFalpha) convertase enzyme (TACE) inhibition assay. RESULTS: Liposomal clodronate-treated rabbits showed a reduction and delay in joint swelling during the first 3 injections. Expression of matrix-bound (solubilized) TNFalpha, lining cell hyperplasia, and levels of RAM-11+ macrophages were low in the synovium of the liposomal clodronate treatment group, but the proportion of apoptotic lining cells was not affected. The radiologic score was low at the end of weeks 2 and 4, but at 8 weeks, no difference, compared with controls, was found in pannus formation or in the extent of joint erosion; also, joint swelling was higher than before initiation of treatment. Injections of liposomal clodronate prevented cartilage proteoglycan loss, which was significant in the superficial zone only. TACE activity was not inhibited by clodronate. CONCLUSION: Liposomal clodronate had temporary antiinflammatory and antierosive effects on established AIA in rabbits. Over the long-term, the loss of cartilage proteoglycans was halted. This observed treatment effect may be related to the inhibition of TNFalpha production and processing in the synovium.

ADAM Proteins↗
PubMed2004

Studies on acute human infections using FTIR microspectroscopy and cluster analysis.

A novel methodology for the diagnosis of acute infections using FTIR microspectroscopy (FTIR-MSP) data on blood components and cluster analysis is presented. Blood samples were collected from 11 patients suffering from various infections and 16 age-matched healthy human controls. Blood components such as white blood cells, red blood cells, and plasma were isolated using standard procedures and FTIR-MSP of these components was utilized. A cluster analysis of the FTIR spectra was performed. The spectra obtained from the three blood components of patients were different from those of controls. The FTIR spectra of white blood cells from patients suffering infections were significantly different from the controls. Cluster analyses of averaged FTIR-MSP spectra of white blood cells provided 100% classification between patients and healthy controls.

Acute Disease↗
PubMed2004

Pressure dependence of human fibrinogen correlated to the conformational alpha-helix to beta-sheet transition: an Fourier transform infrared study microspectroscopic study.

We used Fourier transform infrared (FTIR) microspectroscopy to investigate pressure-induced conformational changes in secondary structure of fibrinogen (FBG). Solid state FBG was compressed on a KBr pellet (1KBr method) or between two KBr pellets (2KBr method). The peak positions of the original and second-derivative ir spectra of compressed FBG samples prepared by the 1KBr method were similar to FBG sample without pressure. When FBG was prepared by the 2KBr method and pressure was increased up to 400 kg/cm(2), peaks at 1625 (intermolecular beta-sheet) and 1611 (beta-sheet aggregates structure and/or the side-chain absorption of the tyrosine residues) cm(-1) were enhanced. The peaks near 1661 (beta-sheet) and 1652 (alpha-helix) cm(-1) also exhibited a marked change with pressure. A linear correlation was found between the peak intensity ratio of 1611/1652 cm(-1) (r = 0.9879) or 1625/1652 cm(-1) (r = 0.9752) and applied pressure. The curve-fitted compositional changes in secondary structure of FBG also indicate that the composition of the alpha-helix structure (1657-1659 cm(-1)) was gradually reduced with the increase in compression pressure, but the composition of the beta-sheet structure (1681, 1629, and 1609 cm(-1)) gradually increased. This indicates that pressure-induced conformational changes in FBG include not only transformations from alpha-helix to beta-sheet structure, but also unfolding and denaturation of FBG and the formation of aggregates.

Fibrinogen↗
PubMed2004

Inflammatory bowel diseases as an intermediate stage between normal and cancer: a FTIR-microspectroscopy approach.

Elucidation of the evolution of inflammatory bowel disease (IBD) to cancer by clinical symptoms and histopathology of biopsies is important. Fourier transform infrared microspectroscopy (FTIR-MSP) has shown promise as a diagnostic tool for distinction of normal and cancer cells and tissues. In the present work, FTIR-MSP is used to evaluate IBD cases and to study the IR spectral characteristic with respect to cancer and normal tissues from formalin-fixed colonic biopsies from patients. Specific regions of the spectra were analyzed by statistical tools to study variations in metabolites that signified changes between the two pathological conditions: IBD and cancer. IBD tissues can be grouped with cancer or normal tissue using certain parameters such as phosphate content and RNA/DNA ratio as calculated from the spectra and show intermediate levels with regard to these metabolites. Further classification of the spectra by cluster analysis indicated which cases of Crohn's disease (3 of 10 cases) or ulcerative colitis (7 of 10 cases) were more likely to progress to cancer. The study exhibits that FTIR-MSP can detect gross biochemical changes in morphologically identical IBD and cancer tissues and suggest which cases of IBD may require further evaluation for carcinogenesis.

Cell Transformation, Neoplastic↗
PubMed1991

Chromophore motion during the bacteriorhodopsin photocycle: polarized absorption spectroscopy of bacteriorhodopsin and its M-state in bacteriorhodopsin crystals.

The three-dimensional crystallization of bacteriorhodopsin was systematically investigated and the needle-shaped crystal form analysed. In these crystals the M-intermediate forms 10 times faster and decays 15 times more slowly than in purple membranes. Polarized absorption spectra of the crystals were measured in the dark and light adapted states. A slight decrease in the angle between the transition moment and the membrane plane was detected during dark adaptation. The crystallization of a mutated bacteriorhodopsin, in which the aspartic acid at residue 96 was replaced by asparagine, provided crystals with a long lived M-intermediate. This allowed polarized absorption measurements of the M-chromophore. The change in the polarization ratio upon formation of the M-intermediate indicates an increase in the angle between the main transition dipole and the membrane plane by 2.2 degrees +/- 0.5, corresponding to a 0.5 A displacement of one end of the chromophore out of the membrane plane of the bacteriorhodopsin molecule.

Bacteriorhodopsins↗
PubMed2005

Spectral evaluation of laser-induced cell damage with photothermal microscopy.

BACKGROUND AND OBJECTIVES: Determining cell photo-damage is important for laser medicine and laser safety standards. This work evaluated the potential of photothermal (PT) technique for studying invasive laser-cell interaction, with a focus on PT evaluation of spectral dependence of laser-induced damage in visible region at single intact cell level. STUDY DESIGN/MATERIALS AND METHODS: PT is based on irradiation of a single intact cells with a tunable pump laser pulse (420-570 nm, 8 nanoseconds, 0.1-300 microJ) and monitoring of temperature-dependent variations of the refractive index with a second, collinear probe beam in pulse (imaging) mode (639 nm, 13 nanoseconds, 10 nJ), and continuous (integrated PT response) mode (633 nm, 2 mW). The local and the integrated PT responses from the individual living red blood cells, lymphocytes, and cancer cells (K562) in vitro were obtained at different pump laser fluence and wavelength and compared with data obtained by conventional viability tests (Annexin V--propidium iodide). RESULTS: The cell damage with pump pulse lead to specific change in PT response's temporal shape and PT image's structure. The photodamage thresholds varied in the range of 0.5-5 J/cm2 for red blood cells, 4.4-42 J/cm2 for lymphocytes, and 36-90 J/cm2 for blast cells in the pump wavelength range of 417-555 nm. CONCLUSION: Damage threshold at different wavelength depends on absorption spectra of cells. Spectral evaluation of laser-damage thresholds can be done in two supplements for each PT mode--PT imaging and integrated PT response. The correlation between specific change of PT parameters and cell damage permits using PT technique to rapidly estimate the invasive conditions of the laser-cell interactions.

Cell Culture Techniques↗
PubMed2001

Degradation of bisphenol A by purified laccase from Trametes villosa.

Degradation of bisphenol A (BPA), an endocrine-disrupting chemical, was studied with a purified laccase from the basidiomycete Trametes villosa. SDS--polyacrylamide gel electrophoresis of the purified laccase gave one single band with a mobility corresponding to MW 65 kDa. The absorption spectrum showed the characteristics of a blue copper protein with a maximum peak at 600 nm. HPLC analysis revealed that 2.2 micromol BPA were degraded by incubation with 1.5 units of the purified laccase in a total volume of 1.0 ml at pH 6.0 and 60 degrees C for 3 h. The enzyme reaction proceeded rapidly without requirement of mediators for the electron transfer. Isolation and identification of several reaction products are in progress, in which one product was identified as 4-isopropenylphenol by a gas chromatography--mass spectrophotometer.

Benzhydryl Compounds↗
PubMed1998

DNA content and chromatin texture of benzo[a]pyrene-transformed human breast epithelial cells as assessed by image analysis.

DNA content and chromatin supraorganization defined in terms of patterns of chromatin texture were studied by image analysis in Feulgen-stained human breast MCF-10F epithelial cells expressing different stages of tumorigenic progression after treatment with benzo[a]pyrene (BP). Nontransformed MCF-10F, nontumorigenic transformed BP1, and tumorigenic BP1-E1 cell lines were analyzed. A relationship between changes in DNA content and chromatin texture and the expression of different stages of tumorigenesis were researched in interphase nuclei. A significant decrease in Feulgen-DNA amounts and absorbing areas suggesting aneuploidy, and first detected in nontumorigenic transformed BP1 cells, became more relevant in the tumorigenic cell line BP1-E1. Simultaneously, an increase in optical densities and a change in related parameters, demonstrating increased chromatin higher order packing states, occurred in transformed BP1 cells and extended over larger chromatin areas in tumorigenic BP1-E1 cells. The results indicate loss of DNA and a change in chromatin higher order packing states accompanying the expression of different stages of the in vitro tumorigenesis process in BP-transformed human breast epithelial cells. It is suspected that the gradually acquired chromatin "condensation" supraorganization pattern is associated with activated transcription activities previously suggested for these cells.

Benzo(a)pyrene↗