The role of histopathology in forensic practice: an overview.

PubMed

Dettmeyer, R B

2014-09-01

The role of forensic histopathology in routine practice is to establish the cause of death in particular cases. This is achieved on the basis of microscopic analysis of representative cell and tissue samples taken from the major internal organs and from abnormal findings made at autopsy. A prerequisite of this is adherence to the quality standards set out for conventional histological/cytological staining and enzyme histochemical and immunohistochemical methods. The interpretation of histological findings is performed by taking into account macroscopic autopsy findings and information on previous history. Histological analysis may prompt postmortem biochemical and chemical-toxicological investigations. The results of histological analysis need to be classified by experts in the context of the available information and the need to withstand the scrutiny of other experts.

[Morphological signs of mitochondrial cytopathy in skeletal muscles and micro-vessel walls in a patient with cerebral artery dissection associated with MELAS syndrome].

PubMed

Sakharova, A V; Kalashnikova, L A; Chaĭkovskaia, R P; Mir-Kasimov, M F; Nazarova, M A; Pykhtina, T N; Dobrynina, L A; Patrusheva, N L; Patrushev, L I; Protskiĭ, S V

2012-01-01

Skin and muscles biopsy specimens of a patient harboring A3243G mutation in mitochondrial DNA, with dissection of internal carotid and vertebral arteries, associated with MELAS were studied using histochemical and electron-microscopy techniques. Ragged red fibers, regional variability of SDH histochemical reaction, two types of morphologically atypical mitochondria and their aggregation were found in muscle. There was correlation between SDH histochemical staining and number of mitochondria revealed by electron microscopy in muscle tissue. Similar mitochondrial abnormality, their distribution and cell lesions followed by extra-cellular matrix mineralization were found in the blood vessel walls. In line with generalization of cytopathy process caused by gene mutation it can be supposed that changes found in skin and muscle microvessels also exist in large cerebral vessels causing the vessel wall "weakness", predisposing them to dissection.

Some biochemical and histochemical properties of human liver serine dehydratase.

PubMed

Kashii, Tatsuhiko; Gomi, Tomoharu; Oya, Takeshi; Ishii, Yoko; Oda, Hirofumi; Maruyama, Muneharu; Kobayashi, Masashi; Masuda, Tohru; Yamazaki, Mitsuaki; Nagata, Takuya; Tsukada, Kazuhiro; Nakajima, Akinori; Tatsu, Kazuhito; Mori, Hisashi; Takusagawa, Fusao; Ogawa, Hirofumi; Pitot, Henry C

2005-03-01

In rat, serine dehydratase (SDH) is abundant in the liver and known to be a gluconeogenic enzyme, while there is little information about the biochemical property of human liver serine dehydratase because of its low content and difficulty in obtaining fresh materials. To circumvent these problems, we purified recombinant enzyme from Escherichia coli, and compared some properties between human and rat liver serine dehydratases. Edman degradation showed that the N-terminal sequence of about 75% of human serine dehydratase starts from MetSTART-Met2-Ser3- and the rest from Ser3-, whereas the N-terminus of rat enzyme begins from the second codon of MetSTART-Ala2-. The heterogeneity of the purified preparation was totally confirmed by mass spectrometry. Accordingly, this observation in part fails to follow the general rule that the first Met is not removed when the side chain of the penultimate amino acid is bulky such as Met, Arg, Lys, etc. There existed the obvious differences in the local structures between the two enzymes as revealed by limited-proteolysis experiments using trypsin and Staphylococcus aureus V8 protease. The most prominent difference was found histochemically: expression of rat liver serine dehydratase is confined to the periportal region in which many enzymes involved in gluconeogenesis and urea cycle are known to coexist, whereas human liver serine dehydratase resides predominantly in the perivenous region. These findings provide an additional support to the previous notion suggested by physiological experiments that contribution of serine dehydratase to gluconeogenesis is negligible or little in human liver.

A histochemical study of the posterior silk glands of Bombyx mori during metamorphosis from larvae to pupae using frozen sections.

PubMed

Kawamoto, K; Kawamoto, T; Shiba, H; Hosono, K

2014-02-01

The fine structures of the whole bodies and the posterior silk glands of Bombyx mori during metamorphosis from larvae to pupae in the cocoon were preserved virtually without damage when frozen sections were prepared using an adhesive plastic film. We used frozen sections for histochemical and enzyme histochemistry to characterize the metamorphosis of the posterior silk glands. Frozen sections were stained with DAPI to observe nuclear changes, examined using the TUNEL method to detect DNA fragments, and investigated using in situ hybridization to detect B. mori caspase expression. Both DNA fragments and expression of B. mori caspase increased with progressing metamorphosis. The degeneration of the posterior silk gland during metamorphosis appears to be an apoptotic event.

Can cadaverous pollution from environmental lead misguide to false positive results in the histochemical determination of gunshot residues? Study on cadaveric skin samples.

PubMed

Boracchi, Michele; Andreola, Salvatore; Collini, Federica; Gentile, Guendalina; Maciocco, Francesca; Maghin, Francesca; Zoja, Riccardo

2017-08-01

Histochemical determination of lead in gunshot residues (GSR) raises the question of possible environmental contamination by heavy metals. Authors assess the specificity of the Sodium Rhodizonate Test in the search of lead derived from environmental pollution. Sodium Rhodizonate Test and 5% HCl Sodium Rhodizonate test were applied to skin samples taken from two groups of victims, whose cause of death was not related to gunshot wounds: group A included 25 corpses found in open spaces after a long time; group B included 16 corpses exhumed after a period of 11 years. The use of these histochemical tests to skin samples from these two groups did not show metal debris histochemically similar to the lead present in GSR. These tests were confirmed to be specific in highlighting the GSR coming from gunshot, without being affected by the potential environmental contamination of the heavy metal pollutants, coming from air or soil. Copyright © 2017 Elsevier B.V. All rights reserved.

In vivo investigation on the chronic hepatotoxicity induced by sertraline.

PubMed

Almansour, Mansour I; Jarrar, Yazun B; Jarrar, Bashir M

2018-05-30

Although sertraline is widely prescribed as relatively safe antidepressant drug, hepatic toxicity was reported in some patients with sertraline treatment. The present study was conducted to investigate the morphometric, hepatotoxicity, and change in gene expression of drug metabolizing enzymes. Male healthy adult rabbits (Oryctolagus cuniculus) ranging from 1050 to 1100 g were exposed to oral daily doses of sertraline (0, 1, 2, 4, 8 mg/kg) for 9 weeks. The animals were subjected to morphometric, hepatohistological, histochemical and quantitative real-time polymerase chain reaction analyses. Sertraline chronic exposure induced morphometric changes and provoked histological and histochemical alterations including: hepatocytes hydropic degeneration, necrosis, nuclear alteration, sinusoidal dilation, bile duct hyperplasia, inflammatory cells infiltration, portal vessel congestion, Kupffer cells hyperplasia, portal fibrosis and glycogen depletion. In addition, the gene expression of drug and arachidonic acid metabolizing enzymes were reduced significantly (p value <0.05). The most affected genes were cyp4a12, ephx2, cyp2d9 and cyp1a2, demonstrating 5 folds or more down-regulation. These findings suggest that chronic sertraline treatment induced toxic histological alterations in the hepatic tissues and reduced the gene expression of drug metabolizing enzymes. Patients on chronic sertraline treatment may be on risk of hepatotoxicity with reduced capacity to metabolize drugs and fatty acids. Copyright © 2018 Elsevier B.V. All rights reserved.

[The experimental research of inner ear metabolism and electrical physiology of autoimmune sensorineural hearing loss].

PubMed

Tan, C; Cao, Y; Hu, P

1998-09-01

Inquire into the mechanism of inner ear pathological physiology in autoimmune sensorineural hearing loss (ASHL). With the auditory electric-physiological techniques and enzyme-histochemical method, the change of inner ear hearing function and enzyme activity were observed. These animals, which threshold of auditory nerve compound active potential (CAP) and cochlear microphonic potential(CM) heightening evidently, showed that the amplitude of endolymphatic potential(EP) (include-EP) bring down in various degrees, which was related to the change of the active of Na(+)-K(+)-ATPase and SDH in vascularis stria and endolymphatic sac. The abnormality of enzymes metabolism in inner ear tissues, which following autoimmune inflammation damage, is the pathological foundation of hearing dysfunction.

Hypogravity-induced atrophy of rat soleus and extensor digitorum longus muscles

NASA Technical Reports Server (NTRS)

Riley, D. A.; Ellis, S.; Slocum, G. R.; Satyanarayana, T.; Bain, J. L.; Sedlak, F. R.

1987-01-01

Prolonged exposure of humans to hypogravity causes weakening of their skeletal muscles. This problem was studied in rats exposed to hypogravity for 7 days aboard Spacelab 3. Hindlimb muscles were harvested 12-16 hours postflight for histochemical, biochemical, and ultrastructural analyses. The majority of the soleus and extensor digitorum longus fibers exhibited simple cell shrinkage. However, approximately 1% of the fibers in flight soleus muscles appeared necrotic. Flight muscle fibers showed increased glycogen, lower subsarcolemmal staining for mitochondrial enzymes, and fewer subsarcolemmal mitochondria. During atrophy, myofibrils were eroded by multiple focal losses of myofilaments; lysosomal autophagy was not evident. Tripeptidylaminopeptidase and calcium-activated protease activities of flight soleus fibers were significantly increased, implying a role in myofibril breakdown. Simple fiber atrophy appears to account for muscle weakening during spaceflight, but fiber necrosis is also a contributing factor.

Thioesters for the in vitro evaluation of agents to image brain cholinesterases.

PubMed

Macdonald, Ian R; Jollymore, Courtney T; Reid, G Andrew; Pottie, Ian R; Martin, Earl; Darvesh, Sultan

2013-06-01

Cholinesterases are associated with pathology characteristic of conditions such as Alzheimer's disease and are therefore, considered targets for neuroimaging. Ester derivatives of N-methylpiperidinol are promising potential imaging agents; however, methodology is lacking for evaluating these compounds in vitro. Here, we report the synthesis and evaluation of a series of N-methylpiperidinyl thioesters, possessing comparable properties to their corresponding esters, which can be directly evaluated for cholinesterase kinetics and histochemical distribution in human brain tissue. N-methylpiperidinyl esters and thioesters were synthesized and they demonstrated comparable cholinesterase kinetics. Furthermore, thioesters were capable, using histochemical method, to visualize cholinesterase activity in human brain tissue. N-methylpiperidinyl thioesters can be rapidly evaluated for cholinesterase kinetics and visualization of enzyme distribution in brain tissue which may facilitate development of cholinesterase imaging agents for application to conditions such as Alzheimer's disease.

[Experimental xenogenic immune pancreatitis. --Immunohistological, enzyme histochemical and ultrastructural studies (author's transl)].

PubMed

Nizze, H

1975-01-01

Repeated intraperitoneal injections of anti-mouse pancreas rabbit serum or of anti-mouse pancreas guinea pig serum produce a chronical sclerotizing pancreatitis. This study has the aim to contribute to the further elucidation of the changes which occur in the acinar cells, as well as to the etiology and pathogenesis of immune pancreatitis, by means of immunohistological, enzyme histochemical and electron microscopic studies. Anti-mouse pancreas rabbit serum was obtained by sensitization of rabbits with an admixture of AB-mouse pancreas extract (100,000 g - supernatant) and complete Freund's adjuvant [details see NIZZE, Exp. Path. (1975a)]. The presence of precipitating mouse pancreas antibodies in the rabbit serum was ascertained by the agargel diffusion test according to Duchterlony (1958). The experiments were performed with 54 adult male white mice (AB colony strain) of 22 to 30 g.b.s. (averagely 26 g). The animals were divided into 4 groups which were treated as follows: 1. 24 mice with anti-mouse pancreas rabbit serum, 2. 12 mice with rabbit normal serum, 3. 12 mice with physiological saline, 4. 6 mice remained untreated (controls) Always 4 animals of the group 1 as well as each 2 of the groups 2 and 3 were administered in total 1, 3, 5, 9, 17 or 33 intraperitoneal injections of 0.3 ml of the correspondent serum or with physiological saline within 3 hours, 1, 2, 4, 8 or 16 days. The last injection was regularly applied 3 hours before sacrification by decapitation. The time of sacrification was always at 11.00 o'clock a.m. For immunohistological and enzyme histochemical investigations 10 mum thick cryostat sections were prepared consisting of pancreatic specimens piled up to a bloc. In each case the tissue samples were taken from the experimental animals and from one control animal sacrificed at the same day. The sections were incubated in FITC-labelled anti-rabbit globulin goat serum at room temperature for 30 min in a moist chamber. For control of specificity were employed: a) initial incubation of equal sections with unlabelled anti-rabbit globulin goat serum for 30 min ("blocking test''), b) pancreatic tissue specimens of each one untreated control animal present in the cryostat sections and thus incubated like the pancreatic tissue of the experimental animals, c) native nonincubated cryostat sections from the same bloc to exclude nonspecific autofluorescence. Evaluation of the sections was done in a Zeiss-Lg-microscope with HBO-50 high pressure mercury lamp. Exciter filters were UG 1/3.5 and 1/1.5, the eyepiece was screened with a GG 9/1 filter photographs were taken on ORWO X-ray film RS 2 (VEB Filmfabrik Wolfen). The enzyme histochemical studies were performed on cryostat sections of the same tissue bloc using the following methods: lead nitrate- or calcium-Co-method after GOMORI (1952) for demonstration of acid and alkaline phosphatase, naphthylacetate method (NACHLAS and SELIGMAN 1945) for nonspecific esterase, MTT-co-method (PEARSE et al...

Effects of thyroid hormone on fast- and slow-twitch skeletal muscles in young and old rats.

PubMed Central

Larsson, L; Li, X; Teresi, A; Salviati, G

1994-01-01

1. The effects of 4 weeks of thyroid hormone treatment on contractile, enzyme-histochemical and morphometric properties and on the myosin isoform composition were compared in the slow-twitch soleus and the fast-twitch extensor digitorum longus (EDL) muscle in young (3-6 months) and old (20-24 months) male rats. 2. In the soleus of untreated controls, contraction and half-relaxation times of the isometric twitch increased by 19-32% with age. The change in contractile properties was paralleled by an age-related increase in the proportions of type I fibres and type I myosin heavy chain (MHC) and slow myosin light chain (MLC) isoforms. 3. In the EDL of controls, contraction and half-relaxation times were significantly prolonged (21-38%) in the post-tetanus twitch in the old animals. No significant age-related changes were observed in enzyme-histochemical fibre-type proportions, although the number of fibres expressing both type IIA and IIB MHCs and of fibres expressing slow MLC isoforms was increased in the old animals. 4. Serum 3,5,3',5'-tetraiodothyronine (T4) levels were lower (34%) in the old animals, but the primary byproduct of T4, 3,5,3'-triiodothyronine (T3), did not differ between young and old animals. 5. The effects of 4 weeks of thyroid hormone treatment were highly muscle specific, and were more pronounced in soleus than in EDL, irrespective of animal age. In the soleus, this treatment shortened the contraction and half-relaxation times by 35-57% and decreased the number of type I fibres by 66-77% in both young and old animals. In EDL, thyroid hormone treatment significantly shortened the contraction time by 24%, but the change was restricted to the old animals. 6. In conclusion, the ability of skeletal muscle to respond to thyroid hormone treatment was not impaired in old age and the age-related changes in speed of contraction and enzyme-histochemical properties and myosin isoform compositions were diminished after thyroid hormone treatment in both the soleus and EDL. Images Figure 3 Figure 4 PMID:7853237

Histo-chemical and biochemical analysis reveals association of er1 mediated powdery mildew resistance and redox balance in pea.

PubMed

Mohapatra, Chinmayee; Chand, Ramesh; Navathe, Sudhir; Sharma, Sandeep

2016-09-01

Powdery mildew caused by Erysiphe pisi is one of the important diseases responsible for heavy yield losses in pea crop worldwide. The most effective method of controlling the disease is the use of resistant varieties. The resistance to powdery mildew in pea is recessive and governed by a single gene er1. The objective of present study is to investigate if er1 mediated powdery mildew resistance is associated with changes in the redox status of the pea plant. 16 pea genotypes were screened for powdery mildew resistance in field condition for two years and, also, analyzed for the presence/absence of er1 gene. Histochemical analysis with DAB and NBT staining indicates accumulation of reactive oxygen species (ROS) in surrounding area of powdery mildew infection which was higher in susceptible genotypes as compared to resistant genotypes. A biochemical study revealed that the activity of superoxide dismutase (SOD) and catalase, enzymes involved in scavenging ROS, was increased in, both, resistant and susceptible genotypes after powdery mildew infection. However, both enzymes level was always higher in resistant than susceptible genotypes throughout time course of infection. Moreover, irrespective of any treatment, the total phenol (TP) and malondialdehyde (MDA) content was significantly high and low in resistant genotypes, respectively. The powdery mildew infection elevated the MDA content but decreased the total phenol in pea genotypes. Statistical analysis showed a strong positive correlation between AUDPC and MDA; however, a negative correlation was observed between AUDPC and SOD, CAT and TP. Heritability of antioxidant was also high. The study identified few novel genotypes resistant to powdery mildew infection that carried the er1 gene and provided further clue that er1 mediated defense response utilizes antioxidant machinery to confer powdery mildew resistance in pea. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

Proliferation of protease-enriched mast cells in sarcoptic skin lesions of raccoon dogs.

PubMed

Noviana, D; W Harjanti, D; Otsuka, Y; Horii, Y

2004-07-01

Skin sites, tongue, lung, liver, jejunum and rectum from two raccoon dogs with Sarcoptes scabiei infestation and five normal (control) raccoon dogs were examined in terms of the distribution, proteoglycan properties and protease activity of mast cells. Infestation with S. scabiei caused a significant increase in the number of dermal mast cells. While the number of mast cells (average +/- standard deviation) in specimens of skin from the dorsum, dorsal neck, dorsal hind foot and dorsal fore foot was 40.0 +/- 19.8/mm2 in control animals, it was 236.1 +/- 58.9/mm2 in the skin of mange-infested animals. Histochemical analysis revealed the glycosaminoglycan, heparin, within the mast cells of all organs examined in both control and affected animals. Enzyme-histochemical detection of serine proteases demonstrated an increase in mast-cell-specific protease activity (i.e., chymase and tryptase) in the skin of infested animals. The percentage of mast cells demonstrating chymase activity was 53.0 +/- 27.4% in control animals and 73.8 +/- 19.4% in mite-infested animals. The corresponding results for tryptase activity were 53.5 +/- 25.2% and 89.4 +/- 9.8%. Increases in mast cell chymase or tryptase activity, or both, were also observed within other organs of the infected animals, but the total number of mast cells found at such sites (with the exception of liver and ventrolateral pinna) did not differ from those of control animals. Copyright 2004 Elsevier Ltd.

Contraction-free, fume-fixed longitudinal sections of fresh frozen muscle

NASA Technical Reports Server (NTRS)

Riley, Danny A.; Slocum, Glenn R.

1988-01-01

Contraction damage occurring when longitudinal frozen sections of fresh unfixed muscles are thawed on microscope slides has limited histological examination of this tissue mainly to cross sections. Longitudinally oriented sections are advantageous for investigating properties that vary along the length of the muscle fibers. A fume fixation technique has been developed for preventing contraction of thick longitudinal frozen sections. The technique is compatible with histochemical staining of enzymes.

CONSIDERATIONS OF THE HISTOLOGIC AND HISTOCHEMICAL INJURY INDUCED BY TOTAL ROENTGEN IRRADIATION IN THE FEMALE GENITAL ORGANS OF PUBERTAL AND PREPUBERTAL MICE

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cecio, A.; Cerasuolo, G.

1961-07-15

Changes in uterus and ovaries were studied in mice exposed to 700 r whole-body irradiation. Endometrial glands showed an increase in mitotic figures after irradiation and somewhat decreased numbers of cells although histochemical tests for polysaccharides and ribonucleic acid (RNA) and deoxyribonucleic acid (DNA) were intensified. The results were similar in adult and immature mice. In ovary, irradiation caused degeneration of follicles with separation of granulosa cells, which was followed by hyperplasia during the reparative phase. Here also polysaccharides were increased in amounts, especially in the zona pellucida of the oocytes. Epithelium of Fallopiant tube also showed increased histochemical reactionsmore » for polysaccharides, RNA, and DNA. Most of these changes were evident within 24 hr after irradiation; they increased in intensity up to the 2nd day, then tended to decrease. (H.H.D.)« less

Histochemical investigation of the activity of oxidoreductases in the skin lesions of lepromatous leprosy patients*

PubMed Central

Žuravleva, G. F.

1972-01-01

This paper reports an investigation of the activity of three basic groups of oxidoreductases in lepromatous leprosy: specific dehydrogenases, flavoprotein enzymes, and cytochrome oxidase. The activity of the enzymes was studied before treatment, at various stages of treatment during exacerbations, and in the stage of regression. The data obtained are of importance for evaluating metabolic process in the cells of the specific infiltrates and the dermal connective tissue in leprosy, for determining the nature and intensity of the inflammatory process, and for control purposes in cases of regression. ImagesFig. 4Fig. 5Fig. 6Fig. 1Fig. 2Fig. 3 PMID:4342274

Liver‐Directed Human Amniotic Epithelial Cell Transplantation Improves Systemic Disease Phenotype in Hurler Syndrome Mouse Model

PubMed Central

Rodriguez, Natalie S.; Yanuaria, Lisa; Parducho, Kevin Murphy R.; Garcia, Irving M.; Varghese, Bino A.; Grubbs, Brendan H.

2017-01-01

Abstract Mucopolysaccharidosis type 1 (MPS1) is an inherited lysosomal storage disorder caused by a deficiency in the glycosaminoglycan (GAG)‐degrading enzyme α‐l‐iduronidase (IDUA). In affected patients, the systemic accumulation of GAGs results in skeletal dysplasia, neurological degeneration, multiple organ dysfunction, and early death. Current therapies, including enzyme replacement and bone marrow transplant, improve life expectancy but the benefits to skeletal and neurological phenotypes are limited. In this study, we tested the therapeutic efficacy of liver‐directed transplantation of a placental stem cell, which possesses multilineage differentiation potential, low immunogenicity, and high lysosomal enzyme activity. Unfractionated human amniotic epithelial cells (hAECs) were transplanted directly into the liver of immunodeficient Idua knockout mouse neonates. The hAECs engraftment was immunohistochemically confirmed with anti‐human mitochondria staining. Enzyme activity assays indicated that hAECs transplantation restored IDUA function in the liver and significantly decreased urinary GAG excretion. Histochemical and micro‐computed tomography analyses revealed reduced GAG deposition in the phalanges joints and composition/morphology improvement of cranial and facial bones. Neurological assessment in the hAEC treated mice showed significant improvement of sensorimotor coordination in the hAEC treated mice compared to untreated mice. Results confirm that partial liver cell replacement with placental stem cells can provide long‐term (>20 weeks) and systemic restoration of enzyme function, and lead to significant phenotypic improvement in the MPS1 mouse model. This preclinical data indicate that liver‐directed placental stem cell transplantation may improve skeletal and neurological phenotypes of MPS1 patients. Stem Cells Translational Medicine 2017;6:1583–1594 PMID:28585336

Liver-Directed Human Amniotic Epithelial Cell Transplantation Improves Systemic Disease Phenotype in Hurler Syndrome Mouse Model.

PubMed

Rodriguez, Natalie S; Yanuaria, Lisa; Parducho, Kevin Murphy R; Garcia, Irving M; Varghese, Bino A; Grubbs, Brendan H; Miki, Toshio

2017-07-01

Mucopolysaccharidosis type 1 (MPS1) is an inherited lysosomal storage disorder caused by a deficiency in the glycosaminoglycan (GAG)-degrading enzyme α-l-iduronidase (IDUA). In affected patients, the systemic accumulation of GAGs results in skeletal dysplasia, neurological degeneration, multiple organ dysfunction, and early death. Current therapies, including enzyme replacement and bone marrow transplant, improve life expectancy but the benefits to skeletal and neurological phenotypes are limited. In this study, we tested the therapeutic efficacy of liver-directed transplantation of a placental stem cell, which possesses multilineage differentiation potential, low immunogenicity, and high lysosomal enzyme activity. Unfractionated human amniotic epithelial cells (hAECs) were transplanted directly into the liver of immunodeficient Idua knockout mouse neonates. The hAECs engraftment was immunohistochemically confirmed with anti-human mitochondria staining. Enzyme activity assays indicated that hAECs transplantation restored IDUA function in the liver and significantly decreased urinary GAG excretion. Histochemical and micro-computed tomography analyses revealed reduced GAG deposition in the phalanges joints and composition/morphology improvement of cranial and facial bones. Neurological assessment in the hAEC treated mice showed significant improvement of sensorimotor coordination in the hAEC treated mice compared to untreated mice. Results confirm that partial liver cell replacement with placental stem cells can provide long-term (>20 weeks) and systemic restoration of enzyme function, and lead to significant phenotypic improvement in the MPS1 mouse model. This preclinical data indicate that liver-directed placental stem cell transplantation may improve skeletal and neurological phenotypes of MPS1 patients. Stem Cells Translational Medicine 2017;6:1583-1594. © 2017 The Authors Stem Cells Translational Medicine published by Wiley Periodicals, Inc. on behalf of AlphaMed Press.

Application of an enzyme-labeled antigen method for visualizing plasma cells producing antibodies against Strep A, a carbohydrate antigen of Streptococcus pyogenes, in recurrent tonsillitis.

PubMed

Onouchi, Takanori; Mizutani, Yasuyoshi; Shiogama, Kazuya; Inada, Ken-ichi; Okada, Tatsuyoshi; Naito, Kensei; Tsutsumi, Yutaka

2015-01-01

Streptococcus pyogenes is the main causative pathogen of recurrent tonsillitis. Histologically, lesions of recurrent tonsillitis contain numerous plasma cells. Strep A is an antigenic carbohydrate molecule on the cell wall of S. pyogenes. As expected, plasma cells in subjects with recurrent tonsillitis secrete antibodies against Strep A. The enzyme-labeled antigen method is a novel histochemical technique that visualizes specific antibody-producing cells in tissue sections by employing a biotin-labeled antigen as a probe. The purpose of the present study was to visualize plasma cells producing antibodies reactive with Strep A in recurrent tonsillitis. Firstly, the lymph nodes of rats immunized with boiled S. pyogenes were paraformaldehyde-fixed and specific plasma cells localized in frozen sections with biotinylated Strep A. Secondly, an enzyme-labeled antigen method was used on human tonsil surgically removed from 12 patients with recurrent tonsillitis. S. pyogenes genomes were PCR-detected in all 12 specimens. The emm genotypes belonged to emm12 in nine specimens and emm1 in three. Plasma cells producing anti-Strep A antibodies were demonstrated in prefixed frozen sections of rat lymph nodes, 8/12 human specimens from patients with recurrent tonsillitis but not in two control tonsils. In human tonsils, Strep A-reactive plasma cells were observed within the reticular squamous mucosa and just below the mucosa, and the specific antibodies belonged to either IgA or IgG classes. Our technique is effective in visualizing immunocytes producing specific antibodies against the bacterial carbohydrate antigen, and is thus a novel histochemical tool for analyzing immune reactions in infectious disorders. © 2014 The Authors. Microbiology and Immunology Published by The Societies and Wiley Publishing Asia Pty Ltd.

Ataxia with loss of Purkinje cells in a mouse model for Refsum disease

PubMed Central

Ferdinandusse, Sacha; Zomer, Anna W. M.; Komen, Jasper C.; van den Brink, Christina E.; Thanos, Melissa; Hamers, Frank P. T.; Wanders, Ronald J. A.; van der Saag, Paul T.; Poll-The, Bwee Tien; Brites, Pedro

2008-01-01

Refsum disease is caused by a deficiency of phytanoyl-CoA hydroxylase (PHYH), the first enzyme of the peroxisomal α-oxidation system, resulting in the accumulation of the branched-chain fatty acid phytanic acid. The main clinical symptoms are polyneuropathy, cerebellar ataxia, and retinitis pigmentosa. To study the pathogenesis of Refsum disease, we generated and characterized a Phyh knockout mouse. We studied the pathological effects of phytanic acid accumulation in Phyh−/− mice fed a diet supplemented with phytol, the precursor of phytanic acid. Phytanic acid accumulation caused a reduction in body weight, hepatic steatosis, and testicular atrophy with loss of spermatogonia. Phenotype assessment using the SHIRPA protocol and subsequent automated gait analysis using the CatWalk system revealed unsteady gait with strongly reduced paw print area for both fore- and hindpaws and reduced base of support for the hindpaws. Histochemical analyses in the CNS showed astrocytosis and up-regulation of calcium-binding proteins. In addition, a loss of Purkinje cells in the cerebellum was observed. No demyelination was present in the CNS. Motor nerve conduction velocity measurements revealed a peripheral neuropathy. Our results show that, in the mouse, high phytanic acid levels cause a peripheral neuropathy and ataxia with loss of Purkinje cells. These findings provide important insights in the pathophysiology of Refsum disease. PMID:19004801

Effect of retinol on the hyperthermal response of normal tissue in vivo

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rogers, M.A.; Marigold, J.C.L.; Hume, S.P.

The effect of prior administration of retinol, a membrane labilizer, on the in vivo hyperthermal response of lysosomes was investigated in the mouse spleen using a quantitative histochemical assay for the lysosomal enzyme acid phosphatase. A dose of retinol which had no effect when given alone enhanced the thermal response of the lysosome, causing an increase in lysosomal membrane permeability. In contrast, the same dose of retinol had no effect on the gross hyperthermal response of mouse intestine; a tissue which is relatively susceptible to hyperthermia. Thermal damage to intestine was assayed directly by crypt loss 1 day after treatmentmore » or assessed as thermal enhancement of x-ray damage by counting crypt microcolonies 4 days after a combined heat and x-ray treatment. Thus, although the hyperthermal response of the lysosome could be enhanced by the administration of retinol, thermal damage at a gross tissue level appeared to be unaffected, suggesting that lysosomal membrane injury is unlikely to be a primary event in hyperthermal cell killing.« less

[Cytochemical localization and properties of selected nucleolytic enzymes].

PubMed

Sierakowska, Halina

2015-01-01

In the article there are shortly outlined studies on cytochemical localization of selected nucleolytic enzymes carried out between 1957-1986 by David Shugar and his coworkers. The histochemical localization of several nucleolytic enzymes in animal and plant tissues was determined by synthesis of specific substrates, alpha-naphthyl esters of 5'- and 3'-nucleotides and their derivatives. In rat tissues phosphodiesterase I was localized in the plasma membrane whereas phosphodiesterase II in the lizosomes, reflecting their physiological roles. The localization of pancreatic type ribonuclease in animal tissues was determined, indicating its role in extracellular digestion. Plant nucleotide pyrophosphatase was localized in several tissues, purified to near homogeneity from potato tubers and its properties and substrate specificity were determined. Application of this enzyme for removal of m7GMP from the "cap" of eukaryotic mRNA allowed to elucidate the role of "cap" in mRNA binding to ribosomes in the process of translation. Furthermore, cyclic nucleotide phosphodiesterase was isolated from potato tubers and its physicochemical properties, oligomeric structure and substrate specificity were elucidated.

Loss of histochemical identity in mast cells lacking carboxypeptidase A.

PubMed

Feyerabend, Thorsten B; Hausser, Heinz; Tietz, Annette; Blum, Carmen; Hellman, Lars; Straus, Anita H; Takahashi, Hélio K; Morgan, Ellen S; Dvorak, Ann M; Fehling, Hans Jörg; Rodewald, Hans-Reimer

2005-07-01

Mast cell carboxypeptidase A (Mc-cpa) is a highly conserved secretory granule protease. The onset of expression in mast cell progenitors and lineage specificity suggest an important role for Mc-cpa in mast cells. To address the function of Mc-cpa, we generated Mc-cpa-null mice. Mc-cpa-/- mast cells lacked carboxypeptidase activity, revealing that Mc-cpa is a nonredundant enzyme. While Mc-cpa-/- peritoneal mast cells were ultrastructurally normal and synthesized normal amounts of heparin, they displayed striking histochemical and biochemical hallmarks of immature mast cells. Wild-type peritoneal mast cells had a mature phenotype characterized by differential histochemical staining with proteoglycan-reactive dyes (cells do not stain with alcian blue but stain with safranin and with berberine) and a high side scatter to forward scatter ratio by flow cytometry and were detergent resistant. In contrast, Mc-cpa-/- peritoneal mast cells, like immature bone marrow-derived cultured mast cells, stained with alcian blue normally or weakly and either did not stain with safranin and berberine or stained weakly, had a low side scatter to forward scatter ratio, and were detergent sensitive. This phenotype was partially ameliorated with age. Thus, histochemistry and flow cytometry, commonly used to measure mast cell maturation, deviated from morphology in Mc-cpa-/- mice. The Mc-cpa-/- mast cell phenotype was not associated with defects in degranulation in vitro or passive cutaneous anaphylaxis in vivo. Collectively, Mc-cpa plays a crucial role for the generation of phenotypically mature mast cells.

Loss of Histochemical Identity in Mast Cells Lacking Carboxypeptidase A

PubMed Central

Feyerabend, Thorsten B.; Hausser, Heinz; Tietz, Annette; Blum, Carmen; Hellman, Lars; Straus, Anita H.; Takahashi, Hélio K.; Morgan, Ellen S.; Dvorak, Ann M.; Fehling, Hans Jörg; Rodewald, Hans-Reimer

2005-01-01

Mast cell carboxypeptidase A (Mc-cpa) is a highly conserved secretory granule protease. The onset of expression in mast cell progenitors and lineage specificity suggest an important role for Mc-cpa in mast cells. To address the function of Mc-cpa, we generated Mc-cpa-null mice. Mc-cpa−/− mast cells lacked carboxypeptidase activity, revealing that Mc-cpa is a nonredundant enzyme. While Mc-cpa−/− peritoneal mast cells were ultrastructurally normal and synthesized normal amounts of heparin, they displayed striking histochemical and biochemical hallmarks of immature mast cells. Wild-type peritoneal mast cells had a mature phenotype characterized by differential histochemical staining with proteoglycan-reactive dyes (cells do not stain with alcian blue but stain with safranin and with berberine) and a high side scatter to forward scatter ratio by flow cytometry and were detergent resistant. In contrast, Mc-cpa−/− peritoneal mast cells, like immature bone marrow-derived cultured mast cells, stained with alcian blue normally or weakly and either did not stain with safranin and berberine or stained weakly, had a low side scatter to forward scatter ratio, and were detergent sensitive. This phenotype was partially ameliorated with age. Thus, histochemistry and flow cytometry, commonly used to measure mast cell maturation, deviated from morphology in Mc-cpa−/− mice. The Mc-cpa−/− mast cell phenotype was not associated with defects in degranulation in vitro or passive cutaneous anaphylaxis in vivo. Collectively, Mc-cpa plays a crucial role for the generation of phenotypically mature mast cells. PMID:15988029

Further Studies on the Lyo and Desmo Components of Several Hydrolytic Enzymes and Their Histochemical Significance

PubMed Central

Hannibal, Mark J.; Nachlas, Marvin M.

1959-01-01

This report describes additional studies of the lyo and desmo components of esterase, alkaline phosphatase, acid phosphatase, leucine aminopeptidase, and β-glucuronidase. The techniques used have already been reported (7). Enzyme diffusion occurs to different degrees in different fixatives, and varies somewhat with each enzyme. Loss of enzymatic activity during fixation occurs as a result of both inactivation due to the chemical reaction of the fixative with the enzymic protein, and diffusion of the lyo component into the fixative. The amount of diffusion into formalin can be reduced by the addition of salts, sucrose, or methocel. The pH of the aqueous medium significantly influences the removal of the lyo fraction from the tissue section. A striking similarity can be noted in the proportions of each fraction of enzyme present in the kidney of the rat, dog, and man. The procedure of fixation and paraffin embedding of tissue blocks does not wholly prevent the diffusion of the lyo component from the tissue sections when they are subsequently immersed in the aqueous incubation medium. PMID:13654449

Observations of mechanisms of attachment in the green alga Ulva mutabilis Føyn. An ultrastructural and light microscopical study of zygotes and rhizoids.

PubMed

Bråten, T

1975-01-01

The development of the rhizoid cells of the green alga Ulva mutabilis was investigated at the ultrastructural level paying special attention to the mechanism of attachment of the plant. Cytochemical data concerning the initial settling of the early zygote are also given. On the basis of histochemical staining and enzyme treatment it is concluded that the adhesive material secreted by the rhizoid cells is chemically different from that secreted by the zygote during the initial settling of the alga.

Manipulation of host plant cells and tissues by gall-inducing insects and adaptive strategies used by different feeding guilds.

PubMed

Oliveira, D C; Isaias, R M S; Fernandes, G W; Ferreira, B G; Carneiro, R G S; Fuzaro, L

2016-01-01

Biologists who study insect-induced plant galls are faced with the overwhelming diversity of plant forms and insect species. A challenge is to find common themes amidst this diversity. We discuss common themes that have emerged from our cytological and histochemical studies of diverse neotropical insect-induced galls. Gall initiation begins with recognition of reactive plant tissues by gall inducers, with subsequent feeding and/or oviposition triggering a cascade of events. Besides, to induce the gall structure insects have to synchronize their life cycle with plant host phenology. We predict that reactive oxygen species (ROS) play a role in gall induction, development and histochemical gradient formation. Controlled levels of ROS mediate the accumulation of (poly)phenols, and phytohormones (such as auxin) at gall sites, which contributes to the new cell developmental pathways and biochemical alterations that lead to gall formation. The classical idea of an insect-induced gall is a chamber lined with a nutritive tissue that is occupied by an insect that directly harvests nutrients from nutritive cells via its mouthparts, which function mechanically and/or as a delivery system for salivary secretions. By studying diverse gall-inducing insects we have discovered that insects with needle-like sucking mouthparts may also induce a nutritive tissue, whose nutrients are indirectly harvested as the gall-inducing insects feeds on adjacent vascular tissues. Activity of carbohydrate-related enzymes across diverse galls corroborates this hypothesis. Our research points to the importance of cytological and histochemical studies for elucidating mechanisms of induced susceptibility and induced resistance. Copyright © 2015 Elsevier Ltd. All rights reserved.

Effect of depletion of glycosaminoglycans and non-collagenous proteins on interstitial hydraulic permeability in rabbit synovium

PubMed Central

Scott, D; Coleman, P J; Abiona, A; Ashhurst, D E; Mason, R M; Levick, J R

1998-01-01

The hydraulic resistance of synovial interstitium helps to retain a lubricating fluid within the joint cavity. The contributions of sulphated glycosaminoglycans to resistance were assessed by selective depletion by chondroitinase ABC, keratanase and heparinases I, II and III in vivo. Also, since glycosaminoglycans do not account fully for the resistance, the contribution of non-collagenous, structural proteins in interstitium was assessed by treatment with chymopapain, a collagen-sparing protease. Ringer solution containing enzyme was injected into the synovial cavity of the knee in anaesthetized rabbits. After ≥ 30 min the intra-articular pressure was raised and the relation between pressure (Pj) and trans-synovial outflow (Q̇s) determined. The slope dQ̇s/dPj at low pressures, i.e. below yield pressure, represents the hydraulic conductance of the lining, i.e. 1/resistance. The contralateral joint received Ringer solution without enzyme as a control. Action of enzymes on the tissue was confirmed by histochemical and immunohistochemical studies. Treatment with chondroitinase ABC (5 joints) increased the hydraulic conductance of the lining by 2.3 times (control, 1.34 ± 0.22 μl min−1 cmH2O−1; post-enzyme, 3.11 ± 0.45 μl min−1 cmH2O−1). This was significantly less than the effects of leech, Streptomyces and testicular hyaluronidases, which caused an average 4.7 times increase (P < 0.001, ANOVA). Analogous findings were made above yield pressure. Treatment with keratanase (3 joints) or heparinases I, II and III (3 joints) caused no significant increase in trans-synovial flows or conductance, even though the concentration of heparan sulphate in synovium is higher than that of chondroitin sulphates or hyaluronan. Treatment with chymopapain (7 joints) caused the greatest increases in trans-synovial flow, which exceeded control flow by an order of magnitude in one case. After 0.1 U chymopapain the average conductance was 6.6 times the control conductance below yield pressure. Immunohistochemical studies confirmed that chymopapain treatment removed the synovial proteoglycans. It is concluded that, despite their similar resistivities in vitro, the different glycosaminoglycans do not contribute equally, weight for weight, to interstitial resistance in vivo. Hyaluronan is the dominant glycosaminoglycan governing synovial interstitial resistance. In addition, non-collagenous structural proteins contribute significantly to interstitial resistance. PMID:9706037

An investigation into the enzyme histochemistry of adenocarcinomas of human large intestine and of the transitional epithelium immediately adjacent to them

PubMed Central

Marsden, J. R.; Dawson, I. M. P.

1974-01-01

Histochemical enzymatic studies were performed on 30 freshly resected large bowel carcinomas, 30 samples of normal colonic epithelium, and six samples of the histologically normal epithelium (so-called transitional epithelium) immediately adjacent to a carcinoma. Five enzymes were studied: nicotine adenine dinucleotide tetrazolium reductase (NADH-TR), glucose-6-phosphate dehydrogenase, succinate dehydrogenase, monoamine oxidase, and acid phosphatase. Quantitative and qualitative differences in enzyme activity were observed between normal, transitional, and carcinomatous mucosa as follows: monoamine oxidase activity was moderate in normal mucosa, high in transitional mucosa, and low in carcinoma. Succinate dehydrogenase activity was high in transitional mucosa and low or moderate in normal and carcinomatous mucosa. Glucose-6-phosphate dehydrogenase activity showed a gradation from low in normal mucosa to high in carcinoma while acid phosphatase showed the reverse of this pattern. The tetrazolium reductase activity was low or moderate in normal and transitional mucosa and high in carcinoma. These differences in enzyme activity and their possible clinical and metabolic significance are discussed. ImagesFig 2Fig 3 PMID:4154840

[Novel CHST6 compound heterozygous mutations cause macular corneal dystrophy in a Chinese family].

PubMed

Qi, Yan-hua; Dang, Xiu-hong; Su, Hong; Zhou, Nan; Liang, Ting; Wang, Zheng; Huang, Shang-zhi

2010-02-01

The aim of this study was to identify mutations of CHST6 gene in a Chinese family with macular corneal dystrophy (MCD) and to investigate the histopathological changes of MCD. Corneal button of the proband was obtained from penetrating keratoplasty for the treatment of severe corneal dystrophy. The sections and ultrathin sections of this specimen were examined under light microscope and transmission electron microscope (TEM). Genomic DNA was extracted from leukocytes in peripheral blood from the family members. The coding region of CHST6 was amplified by polymerase chain reaction (PCR). The PCR products were analyzed by direct sequencing and restriction enzyme digestion. Histochemical study revealed positive results of colloidal iron stain. TEM revealed enlargement of smooth endoplasmic reticulum and the presence of intracytoplasmic vacuoles. Two mutations, Q298X Y358H, were identified in exon 3 of CHST6. Three patients were compound heterozygotes of these two mutations. The C892T transversion occurred at codon 298 turned the codon of glutamine to a stop codon; the T1072C transversion occurred at codon 358 caused a missense mutation, tyrosine to histidine. All six unaffected family members were heterozygotes. These two mutations were not detected in any of the 100 control subjects. The novel compound heterozygous mutation results in loss of CHST6 function and causes the occurrence of MCD. This is the first report of this gene mutation.

The hatching gland cells of trout embryos: characterisation of N- and O-linked oligosaccharides

PubMed Central

DE GASPAR, IGNACIO; BLANQUEZ, MARIA JOSE; FRAILE, BENITO; PANIAGUA, RICARDO; ARENAS, MARIA ISABEL

1999-01-01

A histochemical, light and electron microscopy study of the hatching gland cells (HGCs) in incubated 50-d-old trout embryos is reported. The distribution of carbohydrate residues in the glycoconjugates of these cells was studied by means of a battery of 13 different lectins conjugated with horseradish peroxidase (PNA, ConA, LCA, WGA, SBA, UEA-I, HPA, DBA) or digoxigenin (DSA, MAA, AAA, SNA, GNA). Identification of N- and O-linked oligosaccharides in HGCs was performed by application of both chemical and enzymatic treatments. Present results suggest that HGCs are seromucous cells which store both high choriolytic enzyme (HCE) and low choriolytic enzyme (LCE), and that their cytoplasmic granules, endoplasmic reticulum and Golgi complex contain additional sialic acid-rich glycoproteins. The negative charge of these glycoproteins might be responsible for the rapid expansion of mucin to form a highly hydrated gel, which would facilite the action of these enzymes in programmed cell death and might play a major role during the morphogenic events. PMID:10227672

α-Viniferin-Induced Structural and Functional Alterations in Raillietina echinobothrida, a Poultry Tapeworm.

PubMed

Roy, Bishnupada; Giri, Bikash R

2015-04-01

α-Viniferin, an active component of the plant Carex baccans L., is known for its anticancer, antidiabetic, and anti-inflammatory properties. In Northeast India, different tribes traditionally consume C. baccans to control intestinal helminth infections. Therefore, the present study was carried out to assess the extent of tegumental alteration caused by α-viniferin in Raillietina echinobothrida, a widely prevalent poultry helminth in northeast India. Helminths were exposed in vitro to various doses of α-viniferin (50, 100, and 200 µM/mL of physiological buffered saline) and their motility and mortality were recorded. Stereoscan observations on the parasite exposed to the active compound showed extensive distortion and destruction of the surface fine topography of the tegument compared with controls. The compound also caused extensive damage to the tegument by disintegration of microtriches, disorganization of muscle bundles, and loss of cellular organelles combined with distortion and disruption of the plasma membrane, nuclear membrane, nucleolus, mitochondrial membrane, and cristae. Histochemical and biochemical studies carried out parasites exposed to α-viniferin revealed a decline in the activity of vital tegumental enzymes like acid phosphatase, alkaline phosphatase, and adenosine triphosphatase. Extensive structural and functional alterations observed in the treated parasites are indicative of efficient cestocidal activity of the compound.

Histochemistry and cytochemistry of glucose-6-phosphate dehydrogenase.

PubMed

Van Noorden, C J

1984-01-01

Histochemistry and cytochemistry of glucose-6-phosphate dehydrogenase has found many applications in biomedical research. However, up to several years ago, the methods used often appeared to be unreliable because many artefacts occurred during processing and staining of tissue sections or cells. The development of histochemical methods preventing loss or redistribution of the enzyme by using either polyvinyl alcohol as a stabilizer or a semipermeable membrane interposed between tissue section and incubation medium, has lead to progress in the topochemical localization of glucose-6-phosphate dehydrogenase. Optimization of incubation conditions has further increased the precision of histochemical methods. Precise cytochemical methods have been developed either by the use of a polyacrylamide carrier in which individual cells have been incorporated before staining or by including polyvinyl alcohol in the incubation medium. In the present text, these methods for the histochemical and cytochemical localization of glucose-6-phosphate dehydrogenase for light microscopical and electron microscopical purposes are extensively discussed along with immunocytochemical techniques. Moreover, the validity of the staining methods is considered both for the localization of glucose-6-phosphate dehydrogenase activity in cells and tissues and for cytophotometric analysis. Finally, many applications of the methods are reviewed in the fields of functional heterogeneity of tissues, early diagnosis of carcinoma, effects of xenobiotics on cellular metabolism, diagnosis of inherited glucose-6-phosphate dehydrogenase deficiency, analysis of steroid-production in reproductive organs, and quality control of oocytes of mammals. It is concluded that the use of histochemistry and cytochemistry of glucose-6-phosphate dehydrogenase is of highly significant value in the study of diseased tissues. In many cases, the first pathological change is an increase in glucose-6-phosphate dehydrogenase activity and detection of these early changes in a few cells by histochemical means only, enables prediction of other subsequent abnormal metabolic events. Analysis of glucose-6-phosphate dehydrogenase deficiency in erythrocytes has been improved as well by the development of cytochemical tools. Heterozygous deficiency can now be detected in a reliable way. Cell biological studies of development or maturation of various tissues or cells have profited from the use of histochemistry and cytochemistry of glucose-6-phosphate dehydrogenase activity.(ABSTRACT TRUNCATED AT 400 WORDS)

Lipid Peroxidation Is an Early Symptom Triggered by Aluminum, But Not the Primary Cause of Elongation Inhibition in Pea Roots1

PubMed Central

Yamamoto, Yoko; Kobayashi, Yukiko; Matsumoto, Hideaki

2001-01-01

Pea (Pisum sativum) roots were treated with aluminum in a calcium solution, and lipid peroxidation was investigated histochemically and biochemically, as well as other events caused by aluminum exposure. Histochemical stainings were observed to distribute similarly on the entire surface of the root apex for three events (aluminum accumulation, lipid peroxidation, and callose production), but the loss of plasma membrane integrity (detected by Evans blue uptake) was localized exclusively at the periphery of the cracks on the surface of root apex. The enhancement of four events (aluminum accumulation, lipid peroxidation, callose production, and root elongation inhibition) displayed similar aluminum dose dependencies and occurred by 4 h. The loss of membrane integrity, however, was enhanced at lower aluminum concentrations and after longer aluminum exposure (8 h). The addition of butylated hydroxyanisole (a lipophilic antioxidant) during aluminum treatment completely prevented lipid peroxidation and callose production by 40%, but did not prevent or slow the other events. Thus lipid peroxidation is a relatively early symptom induced by the accumulation of aluminum and appears to cause, in part, callose production, but not the root elongation inhibition; by comparison, the loss of plasma membrane integrity is a relatively late symptom caused by cracks in the root due to the inhibition of root elongation. PMID:11154329

Outdoor post-mortem bite injuries by Tapinoma nigerrimum (Hymenoptera, Formicidae) on a human corpse: Case report.

PubMed

Bonacci, Teresa; Vercillo, Vannio

2015-07-01

Ants are among the insects that colonize exposed human and animal corpses during the early stage of decomposition. In Calabria, Italy (as well as in other countries), Formicidae have been observed preying on immature stages of Diptera and other insects, as well as causing irregular scalloped areas of superficial skin loss on human corpses and animal carcasses. We present a case of injuries on a human corpse caused by ant feeding. The macroscopic appearance is described and the results of a histochemical investigation of the skin lesions caused by worker ants are reported for the first time. The investigation was carried out on the fresh corpse of a 53-year-old man discovered in a rural area of Cosenza province (Calabria, southern Italy). Numerous irregular areas of superficial skin loss caused by the ant Tapinoma nigerrimum (Nylander 1856) (Hymenoptera, Formicidae) were observed on the body surface, inflicted very early in the post-mortem period. Because the classification of lesions is of crucial importance for forensic investigations, the macroscopic appearance and distribution pattern of the lesions on the corpse are illustrated. The histochemical investigation of the damaged skin explains, for the first time, the mechanism of production of the lesions. Copyright © 2015 Elsevier Ltd and Faculty of Forensic and Legal Medicine. All rights reserved.

The effect of retinol on the hyperthermal response of normal tissue in vivo

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rogers, M.A.; Marigold, J.C.; Hume, S.P.

The effect of prior administration of retinol, a membrane labilizer, on the in vivo hyperthermal response of lysosomes was investigated in the mouse spleen using a quantitative histochemical assay for the lysosomal enzyme acid phosphatase. A dose of retinol which had no effect when given alone enhanced the thermal response of the lysosome, causing an increase in lysosomal membrane permeability. In contrast, the same dose of retinol had no effect on the gross hyperthermal response of mouse intestine; a tissue which is relatively susceptible to hyperthermia. Thermal damage to intestine was assayed directly by crypt loss 1 day after treatmentmore » or assessed as thermal enhancement of X-ray damage by counting crypt microcolonies 4 days after a combined heat and X-ray treatment. Thus, although the hyperthermal response of the lysosome could be enhanced by the administration of retinol, thermal damage at a gross tissue level appeared to be unaffected, suggesting that lysosomal membrane injury is unlikely to be a primary event in hyperthermal cell killing.« less

Effects of folic acid deficiency in pregnant Wistar rats on the activities of D5-3 beta hydroxysteroid dehydrogenase and glucose-6 phosphate dehydrogenase in the ovaries of their litters.

PubMed

Uche-Nwachi, E O; Caxton-Martins, A E

1997-06-01

Histochemical studies of the activities of glucose-6-phosphate dehydrogenase (G-6-PD) and D5-3 beta-hydroxysteroid dehydrogenase (D5-3 beta-HSD) in the ovaries of 40 day old litters of Wistar rats whose mothers were folic acid deficient from the 13th day of gestation showed very weak or no enzyme activity. Biochemical estimations of these enzymes showed that the specific activity of 3 beta-HSD in the experimental animal was 20% that of control while that of G-6-PD in the experimental animals was 14% that of control. This implies that folic acid deficiency instituted at a critical period in gestation in Wistar rats adversely affects steroidogenesis in the ovaries of their litters.

Metabolomic, enzymatic, and histochemical analyzes of cassava roots during postharvest physiological deterioration.

PubMed

Uarrota, Virgílio Gavicho; Maraschin, Marcelo

2015-11-05

Under postharvest physiological deterioration cassava root tubers alter the expression of biosynthetic pathways of certain primary and secondary metabolites, as well as the activity of some scavenging enzymes. Therefore, in this study we hypothesized that cassava cultivars differ as to their physiological responses to deterioration and their biochemical profiles can be an indicative of the tolerance or susceptibility to deterioration. The results corroborate the working hypothesis, revealing that high Levels of phenolic acids, scopoletin, carotenoids, proteins, and augmented activities of guaiacol peroxidase and hydrogen peroxide in non-stored cassava roots can be used as potential biomarkers of cassava deterioration. Cassava physiological deterioration depends on cultivar and many compounds are up and downregulated during storage time. Secondary metabolites, enzymes, scopoletin, scavenging reactive oxygen species, and acidic polysaccharides are activated as responses to the physiological stress induced in root tubers.

A Simple Picaxe Microcontroller Pulse Source for Juxtacellular Neuronal Labelling.

PubMed

Verberne, Anthony J M

2016-10-19

Juxtacellular neuronal labelling is a method which allows neurophysiologists to fill physiologically-identified neurons with small positively-charged marker molecules. Labelled neurons are identified by histochemical processing of brain sections along with immunohistochemical identification of neuropeptides, neurotransmitters, neurotransmitter transporters or biosynthetic enzymes. A microcontroller-based pulser circuit and associated BASIC software script is described for incorporation into the design of a commercially-available intracellular electrometer for use in juxtacellular neuronal labelling. Printed circuit board construction has been used for reliability and reproducibility. The current design obviates the need for a separate digital pulse source and simplifies the juxtacellular neuronal labelling procedure.

Characterizing intestinal strictures with acoustic resolution photoacoustic microscopy

NASA Astrophysics Data System (ADS)

Lei, Hao; Xu, Guan; Liu, Shengchun; Johnson, Laura A.; Moons, David S.; Higgins, Peter D. R.; Rice, Michael D.; Ni, Jun; Wang, Xueding

2016-03-01

Crohn's disease (CD) is an autoimmune disease, which may cause obstructing intestinal strictures due to inflammation, fibrosis (deposition of collagen), or a combination of both. Identifying the different stages of the disease progression is still challenging. In this work, we indicated the feasibility of non-invasively characterizing intestinal strictures using photoacoustic imaging (PAI), utilizing the uniquely optical absorption of hemoglobin and collagen. Surgically removed human intestinal stricture specimens were investigated with a prototype PAI system. 2D PA images with acoustic resolution at wavelength 532, 1210 and 1310 nm were formulated, and furthermore, the PA histochemical components images which show the microscopic distributions of histochemical components were solved. Imaging experiments on surgically removed human intestinal specimens has demonstrated the solved PA images were significantly different associated with the presence of fibrosis, which could be applied to characterize the intestinal strictures for given specimens.

Partial transformation from fast to slow muscle fibers induced by deafferentation of capsaicin-sensitive muscle afferents.

PubMed

Brunetti, O; Barazzoni, A M; Della Torre, G; Clavenzani, P; Pettorossi, V E; Bortolami, R

1997-11-01

Mechanical and histochemical characteristics of the lateral gastrocnemius (LG) muscle of the rat were examined 21 days after capsaicin injection into the LG muscle. The capsaicin caused a decrease in generation rate of twitch and tetanic tension and an increase in fatigue resistance of LG muscle. The histochemical muscle fiber profile evaluated by myosin adenosine triphosphatase and reduced nicotinamide adenine dinucleotide tetrazolium reductase methods showed an increase of type I and IIC fibers and a decrease of the type IIB in whole muscle, and a decrease of the IIA, IIX fibers in the red part accompanied by their increase in the white part. Therefore the capsaicin treatment, which selectively eliminated fibers belonging to the III and IV groups of muscle afferents, induced muscle fiber transformation from fast contracting fatiguing fibers to slowly contracting nonfatiguing ones.

Histochemical study of alkali-burned rabbit anterior eye segment in which severe lesions were prevented by aprotinin treatment.

PubMed

Cejková, J; Lojda, Z; Salonen, E M; Vaheri, A

1989-01-01

Activities of different enzymes (acid glycosidases, phosphatases, Na+ - K+ -dependent ATPase, proteases, dehydrogenases) and acid glycosaminoglycans were studied by histochemical methods in sections of rabbit anterior eye segments after experimental alkali burn and treatment with aprotinin, an inhibitor of plasmin and other serine proteinases. Solutions of sodium hydroxide (0.25-1.0 M) were applied on corneas using 12-mm-diameter plastic tube for 15-60 s. After wiping with cotton and rinsing with tap water aprotinin solutions were applied in saline (in experimental animals) and saline (in control animals) dropwise in 12-h intervals for a month. Within the first two weeks aprotinin was used at a concentration of 5000 IU/ml. During the subsequent two weeks the aprotinin concentration was reduced to 2500 IU/ml. Striking differences in enzyme activities and in the healing between treated and untreated eyes were found. Without aprotinin, ulcers developed in most corneas within 3 weeks and plasmin was regularly demonstrated in tears and in the aqueous. When aprotinin treatment was started within 24 h after the burn, the number of enzymatically active inflammatory cells was significantly lower, not only in the cornea itself but also in the whole anterior eye segment. With aprotinin treatment no ulcerations and no plasmin in tears and the aqueous were observed and the corneas healed within a month. The healing process started from the zone of enzymatically activated corneal cells in the unburned zone at the corneal periphery. In the regenerating epithelium and endothelium high activities of Na+ -K+ -dependent ATPase, gamma-glutamyltransferase, lactate and succinate dehydrogenases appeared very soon.(ABSTRACT TRUNCATED AT 250 WORDS)

CHOLINESTERASE IN DENERVATED END PLATES AND MUSCLE FIBRES

PubMed Central

Brzin, Miro; Majcen-Tkačev, Živa

1963-01-01

Parallel studies were made of cholinesterase activities and localizations in denervated rat and rabbit gastrocnemius muscle. Koelle's histochemical reaction was used for demonstrating the localization of cholinesterases. Enzyme activities in whole sliced muscle were measured by electrometric titration. The Cartesian ampulla-diver technique was used for cholinesterase activity determinations in end plate regions or in small pieces of the muscle fibre itself. No changes in the activity of cholinesterases (ChE) were found in the whole denervated muscle which would account for its chemical supersensitivity. The ChE distribution pattern was changed so that the end plate region became less active in the denervated muscle than in the normal one. The decrease in ChE activity in the end plates seems to be largely compensated for by an increase of this enzyme elsewhere in the muscle. A possible connection between the spatial spread of cholinesterase activity and the enlargement of the acetylcholine-sensitive surface is discussed. PMID:14086761

Study of the role of epididymal alpha-glucosidase in the fertility of male rats by the administration of the enzyme inhibitor castanospermine.

PubMed

Yeung, C H; Cooper, T G

1994-11-01

The activity of epididymal alpha-glucosidase in adult rats was rapidly suppressed to histochemically undetectable levels within 2 days by the continuous release of the enzyme inhibitor castanospermine via a peritoneal osmotic pump at a rate of 100-200 nmol h-1. It was established that mating activities overnight depleted 72% of the spermatozoa in the distal cauda, which was replenished in 2 days, and that fertility began to decline 3 weeks after efferent duct ligation. Male rats of proven mating proficiency and fertility were treated with castanospermine, or buffered saline as control, for up to 30 days and enzyme inhibition was confirmed at the end of treatment by histochemistry. Fertility was normal at the first mating test on day 7, significantly decreased at the second mating on day 9, but recovered in a stepwise manner at subsequent matings on days 12 and 14. Delaying the third mating until day 25 did not sustain the transient subfertility. However, prolonging sperm storage in the distal cauda epididymides and preventing replenishment with freshly matured spermatozoa, by efferent duct ligation for 14 days performed on day 15 during castanospermine administration, caused a decrease in fertility and a change in the kinematics of epididymal spermatozoa of the castanospermine-treated group. In control rats, binding of epididymal spermatozoa to Vicia faba, a lectin specific for glucose and glucosamine, and mannose and mannosamine residues, decreased from the proximal caput to the distal corpus coincident with the increase in alpha-glucosidase activity on the epithelial brush border. Lectin binding then increased in the cauda where enzyme activity was absent. However, castanospermine treatment did not significantly alter this binding profile. The findings suggest that epididymal alpha-glucosidase does not play a crucial role in the development of sperm fertilizing capacity, but may be involved in the preparation of spermatozoa for storage.

Characterisation of Nitric Oxide Synthase in Three Cnidarian-Dinoflagellate Symbioses

PubMed Central

Safavi-Hemami, Helena; Young, Neil D.; Doyle, Jason; Llewellyn, Lyndon; Klueter, Anke

2010-01-01

Background Nitric oxide synthase (NOS) is an enzyme catalysing the conversion of L-arginine to L-citrulline and nitric oxide (NO), the latter being an essential messenger molecule for a range of biological processes. Whilst its role in higher vertebrates is well understood little is known about the role of this enzyme in early metazoan groups. For instance, NOS-mediated signalling has been associated with Cnidaria-algal symbioses, however controversy remains about the contribution of enzyme activities by the individual partners of these mutualistic relationships. Methodology/Principal Findings Using a modified citrulline assay we successfully measured NOS activity in three cnidarian-algal symbioses: the sea anemone Aiptasia pallida, the hard coral Acropora millepora, and the soft coral Lobophytum pauciflorum, so demonstrating a wide distribution of this enzyme in the phylum Cnidaria. Further biochemical (citrulline assay) and histochemical (NADPH-diaphorase) investigations of NOS in the host tissue of L. pauciflorum revealed the cytosolic and calcium dependent nature of this enzyme and its in situ localisation within the coral's gastrodermal tissue, the innermost layer of the body wall bearing the symbiotic algae. Interestingly, enzyme activity could not be detected in symbionts freshly isolated from the cnidarians, or in cultured algal symbionts. Conclusions/Significance These results suggest that NOS-mediated NO release may be host-derived, a finding that has the potential to further refine our understanding of signalling events in cnidarian-algal symbioses. PMID:20442851

Characterisation of nitric oxide synthase in three cnidarian-dinoflagellate symbioses.

PubMed

Safavi-Hemami, Helena; Young, Neil D; Doyle, Jason; Llewellyn, Lyndon; Klueter, Anke

2010-04-28

Nitric oxide synthase (NOS) is an enzyme catalysing the conversion of L-arginine to L-citrulline and nitric oxide (NO), the latter being an essential messenger molecule for a range of biological processes. Whilst its role in higher vertebrates is well understood little is known about the role of this enzyme in early metazoan groups. For instance, NOS-mediated signalling has been associated with Cnidaria-algal symbioses, however controversy remains about the contribution of enzyme activities by the individual partners of these mutualistic relationships. Using a modified citrulline assay we successfully measured NOS activity in three cnidarian-algal symbioses: the sea anemone Aiptasia pallida, the hard coral Acropora millepora, and the soft coral Lobophytum pauciflorum, so demonstrating a wide distribution of this enzyme in the phylum Cnidaria. Further biochemical (citrulline assay) and histochemical (NADPH-diaphorase) investigations of NOS in the host tissue of L. pauciflorum revealed the cytosolic and calcium dependent nature of this enzyme and its in situ localisation within the coral's gastrodermal tissue, the innermost layer of the body wall bearing the symbiotic algae. Interestingly, enzyme activity could not be detected in symbionts freshly isolated from the cnidarians, or in cultured algal symbionts. These results suggest that NOS-mediated NO release may be host-derived, a finding that has the potential to further refine our understanding of signalling events in cnidarian-algal symbioses.

The clinical, histochemical, and molecular spectrum of PEO1 (Twinkle)-linked adPEO

PubMed Central

Fratter, C.; Gorman, G.S.; Stewart, J.D.; Buddles, M.; Smith, C.; Evans, J.; Seller, A.; Poulton, J.; Roberts, M.; Hanna, M.G.; Rahman, S.; Omer, S.E.; Klopstock, T.; Schoser, B.; Kornblum, C.; Czermin, B.; Lecky, B.; Blakely, E.L.; Craig, K.; Chinnery, P.F.; Turnbull, D.M.; Horvath, R.; Taylor, R.W.

2010-01-01

Background: Mutations in the Twinkle (PEO1) gene are a recognized cause of autosomal dominant progressive external ophthalmoplegia (adPEO), resulting in the accumulation of multiple mitochondrial DNA (mtDNA) deletions and cytochrome c oxidase (COX)-deficient fibers in skeletal muscle secondary to a disorder of mtDNA maintenance. Patients typically present with isolated extraocular muscle involvement, with little apparent evidence of the clinical heterogeneity documented in other mtDNA maintenance disorders, in particular POLG-related disease. Methods: We reviewed the clinical, histochemical, and molecular genetics analysis of 33 unreported patients from 26 families together with all previous cases described in the literature to define the clinical phenotype associated with PEO1 mutations. Results: Ptosis and ophthalmoparesis were almost universal clinical features among this cohort, with 52% (17/33) reporting fatigue and 33% (11/33) having mild proximal myopathy. Features consistent with CNS involvement were rarely described; however, in 24% (8/33) of the patients, cardiac abnormalities were reported. Mitochondrial histochemical changes observed in muscle showed remarkable variability, as did the secondary mtDNA deletions, which in some patients were only detected by PCR-based assays and not Southern blotting. Moreover, we report 7 novel PEO1 variants. Conclusions: Our data suggest a shared clinical phenotype with variable mild multiorgan involvement, and that the contribution of PEO1 mutations as a cause of adPEO may well be underestimated. Direct sequencing of the PEO1 gene should be considered in adPEO patients prior to muscle biopsy. GLOSSARY adPEO = autosomal dominant progressive external ophthalmoplegia; COX = cytochrome c oxidase; IOSCA = infantile-onset spinocerebellar ataxia; mtDNA = mitochondrial DNA; PEO = progressive external ophthalmoplegia; SANDO = sensory ataxic neuropathy, dysarthria, and ophthalmoparesis; SDH = succinate dehydrogenase. PMID:20479361

Histogenesis of splenic lesions in Hodgkin's disease.

PubMed

Yam, L T; Li, C Y

1976-12-01

Histochemical markers were used to identify the various cellular and structural components of the human spleen, and to investigate the histogenesis of the splenic lesions of Hodgkin's disease. The early lesions appear in areas near the central artery (periarterial lymphatic sheath) in the white pulp. The white pulp becomes hypertrophic. The lesions enlarge, extend into the red pulp, and compress the sinuses and the cords of Billroth. The derivations of various "histiocytes" contained with the lesions are differentiated by using cytochemical stains for lysosomal enzymes and for granulocytes. The epithelioid cells in the granulomas are rich in those lysosomal enzymes typically seen in phagocytic histiocytes, suggesting that they are indeed true histiocytes. The malignant "histiocytes," including the mononuclear Hodgkin cells, the binucleated Sternberg-Reed cells, and the multinucleated giant cells, do not contain significant amounts of lysosomal enzymes and more closely resemble stimulated lymphocytes. The splenic lesions in Hodkin's disease may be the result of a lymphocytic and histiocytic cellular response to an unknown agent, which reaches the spleen through the central artery in the white pulp.

A Simple Picaxe Microcontroller Pulse Source for Juxtacellular Neuronal Labelling †

PubMed Central

Verberne, Anthony J. M.

2016-01-01

Juxtacellular neuronal labelling is a method which allows neurophysiologists to fill physiologically-identified neurons with small positively-charged marker molecules. Labelled neurons are identified by histochemical processing of brain sections along with immunohistochemical identification of neuropeptides, neurotransmitters, neurotransmitter transporters or biosynthetic enzymes. A microcontroller-based pulser circuit and associated BASIC software script is described for incorporation into the design of a commercially-available intracellular electrometer for use in juxtacellular neuronal labelling. Printed circuit board construction has been used for reliability and reproducibility. The current design obviates the need for a separate digital pulse source and simplifies the juxtacellular neuronal labelling procedure. PMID:28952589

[Histochemical characteristics of the secretory cells of gastric glands compared].

PubMed

Shubich, M G; Mogil'naia, G M; Dudetskiĭ, V I; Bogatyr', L Ia

1978-02-01

The work is dedicated to complex histological studies of the secreting cells in the gastric fundal glands, in their comparative aspect. In the representatives of Amphibia, Reptilians and birds, histochemical differentiation of oxyntopeptic cells was demonstrated to be independent on the peculiarities of the animal nutrition. In mammals, histochemical characteristic of the carbohydrate component in the glandular secreting cells depends on the type of nutrition.

Muscle morphology and mitochondrial investigations of a family with autosomal dominant cerebellar ataxia and retinal degeneration mapped to chromosome 3p12-p21.1.

PubMed

Forsgren, L; Libelius, R; Holmberg, M; von Döbeln, U; Wibom, R; Heijbel, J; Sandgren, O; Holmgren, G

1996-12-01

The autosomal dominant cerebellar ataxias (ADCA) are a group of neurodegenerative disorders with ataxia and dysarthria as early and dominant signs. In ADCA type II, retinal degeneration causes severe visual impairment. ADCA type II has recently been mapped to chromosome 3p by three independent groups. In the family with ADCA type II studied here, the disease has been mapped to chromosome 3p12-p21.1. Histochemical examination of muscle biopsies in 5 cases showed slight neurogenic atrophy and irregular lobulated appearance or focal decreases of enzyme activity when staining for NADH dehydrogenase, succinic dehydrogenase and cytochrome oxidase. Ragged-red fibres were scarce. Electron microscopic examination showed uneven distribution of mitochondria with large fibre areas devoid of mitochondria and/or large subsarcolemmal accumulations of small rounded mitochondria, and frequent autophagic vacuoles. These vacuoles contained remnants of multiple small rounded organelles, possibly mitochondria, and had a remarkably consistent ultrastructural appearance. Biochemical investigation of mitochondrial function showed reduced activity of complex IV and slightly reduced activity of complex I in the respiratory chain in a severely affected child while no abnormalities were found in his affected uncle.

Histopathological and histochemical changes in the liver of Bagrus bayad caused by environmental pollution.

PubMed

Gaber, Hanan S; Ibrahim, Seham A; El-Kasheif, Midhat A

2015-09-01

This study focused on two areas (Abu-Zaabal and El-Tal El-Kabir) of the Ismailia canal. The samples represented the effect of the factories' effluent discharge onto the canal. Physical characteristics (water temperature and pH) and chemical characteristics (total hardness, dissolved oxygen, biological oxygen demand and chemical oxygen demand) were measured to identify the water quality in the two areas of the Ismailia canal. These measurements showed slight variations between the two stations. Additionally, the histological and histochemical analyses in the liver of Bagrus bayad fish samples were done. Histological studies of liver showed more severe degeneration in the fish samples of El-Tal El-Kabir station. Histochemical studies on protein and glycogen contents in the liver of B. bayad showed a progressive decrease in their staining intensity to bromophenol blue and periodic acid Schiff at two stations. This study, therefore, concludes that Ismailia canal water (Abu-Zaabal and El-Tall El-Kabir) stations create metabolic crisis and impairment in fish liver. The depletion in the levels of protein and glycogen points leads to the exhaustion of cell energy that is of high demand for fish during stressful conditions. Therefore, it is imperative for the authorities concerned to ensure that treated effluent discharge comply with acceptable standards to save our environment from destruction. © The Author(s) 2013.

High resolution and image processing of otoconia matrix

NASA Technical Reports Server (NTRS)

Fermin, C. D.

1993-01-01

This study was designed to investigate patterns of fibrils organization in histochemically stained otoconia. Transmission electron microscope and video imaging were used. These data indicate that otoconia of the chick (Gallus domesticus) inner ear may have central cores in vivo. The data also show that the ultrastructural organization of fibrils fixed with aldehydes and histochemical stains follows trajectories that conform to the hexagonal shape of otoconia. These changes in direction may contribute to the formation of a central core. The existence of central cores is important for the in vivo buoyancy of otoconia. Packing of fibrils is tighter after phosphotungstic acid (PTA) stained otoconia than with other histochemical stains, which usually produce looser packing of fibrils and seemingly larger central core. TEM of tilted and untilted material showed that turning of fibrils occurs at the points where the face angles of otoconia form and where central cores exist. Video image processing of the images allowed reconstructing a template which, if assumed to repeat and change trajectories, would fit the pattern of fibrils seen in fixed otoconia. Since it is highly unlikely that aldehyde primary fixation or PTA stain caused such drastic change in the direction of fibrils, the template derived from these results may closely approximate patterns of otoconia fibrils packing in vivo. However, if the above is correct, the perfect crystallographic diffraction pattern of unfixed otoconia do not correspond to patterns of fixed fibrils.

Beta-glucuronidase and hexosaminidase are marker enzymes for different compartments of the endo-lysosomal system in mussel digestive cells.

PubMed

Izagirre, U; Angulo, E; Wade, S C; ap Gwynn, I; Marigómez, I

2009-02-01

In environmental toxicology, the most commonly used techniques used to visualise lysosomes in order to determine their responses to pollutants (LSC test: lysosomal structural changes test; LMS test: lysosomal membrane stability test) are based on the histochemical application of lysosomal marker enzymes. In mussel digestive cells, the marker enzymes used are beta-glucuronidase (beta-Gus) and hexosaminidase (Hex). The present work has been aimed at determining the distribution of these lysosomal marker enzymes in the various compartments of the endo-lysosomal system (ELS) of mussel digestive cells and at exploring whether intercellular transfer of lysosomal enzymes occurs between digestive and basophilic cells. Immunogold cytochemistry has allowed us to conclude that beta-Gus is present in every compartment of the digestive cell ELS, whereas Hex is not so widely distributed. Moreover, Hex is intimately linked to the lysosomal membrane, whereas beta-Gus appears to be not necessarily membrane-bound. Therefore, two populations of heterolysosomes with different enzyme load and membrane stability have been distinguished in the digestive cell. In addition, heterolysosomes of different electron density have been commonly observed merging together by contact; we suggest that some might act as storage granules for lysosomal enzymes. On the other hand, beta-Gus seems to be released to the digestive alveolar lumen in secretory lysosomes produced by basophilic cells and endocytosed by digestive cells. Regarding the implications of the present study on the interpretation of lysosomal biomarkers, we conclude that beta-Gus, but not Hex, histochemistry provides an appropriate marker for the LSC test and that, although both lysosomal marker enzymes can be employed in the LMS test, different values would be obtained depending on the marker enzyme employed.

Histochemical and biochemical studies of carbonic anhydrase activity in the opercular epithelium of the euryhaline teleost, Fundulus heteroclitus.

PubMed

Lacy, E R

1983-01-01

Carbonic anhydrase (CAH) activity was biochemically measured and histochemically localized (at both the light and electron microscope levels) in isolated opercular membranes from teleost fish, Fundulus heteroclitus, adapted to freshwater (FW), seawater (SW), and double-strength seawater (2 x SW). The normal morphology of this membrane showed that its epithelial portion consisted of five cell types: (1) chloride cells, which have been previously implicated as responsible for the active chloride transport across the epithelium; (2) mucous cells; (3) pavement cells, which formed the major portion of the free epithelial surface; (4) supportive cells, which had an abundance of intermediate (10 nm)-type filaments suggesting a structural role for these cells; and (5) vesicular cells, which were characterized by various types of membrane-bound vesicles, including lysosomes, and numerous free ribosomes. Vesicular cells may be stem cells and/or endocrine cells. Hansson's histochemical method for CAH revealed cobalt sulfide reaction product confined to the following structures in fish from each environment: (1) chloride cells: throughout the cytoplasm and some nuclear staining; (2) mucous cells: throughout the cytoplasm, some nuclear staining, and some in mucous granules; (3) vesicular cells: confined to lysosomes, some of the vesicles, and nucleoli; (4) a small portion of the intracellular space between adjacent vesicular cells and supportive cells; and (5) supportive cells: in nucleoli and occasionally in larger membrane-bound lysosomelike structures. Acetazolamide (10(-5) M) and potassium cyanate (KCNO) (10(-1) M) in Hansson's incubation medium completely inhibited the formation of reaction product. Biochemical determination of CAH activity on vascularly perfused, isolated opercular membranes showed no statistically significant difference in enzyme activity between environmental groups. The following units of activity/mg opercular membrane protein were measured: FW: 0.63 +/- 0.02; SW: 0.43 +/- 0.08; 2 x SW: 0.64 +/- 0.09.

Separation of periportal and perivenous rat hepatocytes by fluorescence-activated cell sorting: confirmation with colloidal gold as an exogenous marker.

PubMed

Braakman, I; Keij, J; Hardonk, M J; Meijer, D K; Groothuis, G M

1991-01-01

Periportal and perivenous hepatocytes are known to display various functional differences. In this study we present a new method to separate periportal and perivenous cells: after selectively loading zone 1 or zone 3 with the fluorescent label acridine orange in an antegrade or retrograde perfusion, respectively, we separated the isolated hepatocytes on a fluorescence-activated cell sorter. The common way to check on proper separation is to estimate activities of enzymes known to exhibit a heterogeneous acinar distribution. Using enzyme histochemistry, however, we found that already on short collagenase perfusion, some enzymes displayed a more shallow gradient than in vivo, making enzyme activities less suitable as zonal markers. We therefore used colloidal gold granules (17 nm) injected intravenously (2.5 mg) into the rat 2 to 3 hr before cell isolation. The gold is taken up predominantly by perivenous hepatocytes, probably because of the efficient removal of gold granules in zone 1 by competing Kupffer cells. We compared acridine orange fluorescence, presence of gold particles and activities of six marker enzymes, three biochemically and three histochemically determined. Acridine orange and gold both pointed to a high enrichment of the fractions, whereas most enzyme activities were more randomly distributed among the cells as a result of the isolation procedure. Our separation procedure yielded fractions highly enriched in either viable periportal or perivenous cells, both from one liver. The use of colloidal gold as a marker to monitor separation is a valuable alternative to the more risky estimation of enzyme activities.

Histochemical Detection of Acetogenins and Storage Molecules in the Endosperm of Annona Macroprophyllata Donn Sm. Seeds

PubMed Central

Hernández, G. Laguna; Franco, A.E. Brechú; De-la-Cruz-Chacón, I.; González-Esquinca, A.R.

2015-01-01

Acetogenins (ACGs) are bioactive compounds with cytotoxic properties in different cell lines. They are antitumoural, antiparasitic, antimalarial, insecticidal, antimicrobial, anti-fungal and antibacterial. These secondary metabolites function in plant defence and are found in specific organelles and specific cells, thereby preventing toxicity to the plant itself and permitting site-specific defence. The aim of this work was to histochemically determine the in situ localisation of ACGs in the endosperm of Annona macroprophyllata seeds using Kedde’s reagent. Additionally, the co-localisation of ACGs with other storage molecules was analysed. The seeds were analysed after 6 and 10 days of imbibition, when 1 or 2 cm of the radicle had emerged and metabolism was fully established. The seeds were then transversally cut in half at the midline and processed using different histological and histochemical techniques. Positive reactions with Kedde’s reagent were only observed in fresh, unfixed sections that were preserved in water, and staining was found only in the large cells (the idioblasts) at the periphery of the endosperm. The ACGs’ positive reaction with Sudan III corroborated their lipid nature. Paraffin sections stained with Naphthol Blue Black showed reactions in the endosperm parenchyma cells and stained the proteoplasts blue, indicating that they might correspond to storage sites for albumin-like proteins. Lugol’s iodine, which is similar in chemical composition to Wagner’s reagent, caused a golden brown reaction product in the cytoplasm of the idioblasts, which may indicate the presence of alkaloids. Based on these results, we propose that Kedde’s reagent is an appropriate histochemical stain for detecting ACGs in situ in idioblasts and that idioblasts store ACGs and probably alkaloids. ACGs that are located in idioblasts found in restricted, peripheral areas of the endosperm could serve as a barrier that protects the seeds against insects and pathogen attack. PMID:26428881

Antinephrolithiatic and antioxidative efficacy of Dolichos biflorus seeds in a lithiasic rat model.

PubMed

Saha, Sarmistha; Verma, Ramtej J

2015-01-01

Dolichos biflorus sensu auct non L. (Fabaceae) is widely used for the treatment of kidney stones, leucorrhoea, urinary disorders, and menstrual troubles, and is known for its antioxidant activity. To evaluate the preventive effect of hydro-alcoholic extract of Dolichos biflorus seeds (DBE) in ethylene glycol induced nephrolithiasis. In vitro antioxidative capacity of DBE was estimated in terms of reducing power, superoxide radical, 2,2- diphenyl-1-picrylhydrazyl radical, and nitric oxide scavenging activity. A validated HPLC method was used for standardization using quercetin as a marker. Adult female Wistar rats were administered with DBE (150 and 300 mg/kg body weight/day) along with ethylene glycol (0.75%, v/v) for 28 d. The various biochemical parameters were measured in urine, serum, and kidney followed by histochemistry. Ethylene glycol caused a significant increase in calcium, oxalate, phosphate, and total protein in urine as well as in kidney whereas decrease in calcium, sodium, and magnesium in serum was observed (p < 0.001). Ethylene glycol also caused a significant increase in lipid peroxidation and concurrent decrease in activities of antioxidant enzymes in kidney (p < 0.001). However, the seed extract of D. biflorus caused significant restoration of all these parameters (p < 0.001). Histopathological and histochemical studies also showed the reduced calcifications in kidney of seed extract treated rats. These results indicated that seeds of D. biflorus have significant prophylactic effect in preventing the nephrolithiasis, which might be due to the antioxidant activity of the active compounds of the plant.

Pathogenesis of mitral valve disease in mucopolysaccharidosis VII dogs.

PubMed

Bigg, Paul W; Baldo, Guilherme; Sleeper, Meg M; O'Donnell, Patricia A; Bai, Hanqing; Rokkam, Venkata R P; Liu, Yuli; Wu, Susan; Giugliani, Roberto; Casal, Margret L; Haskins, Mark E; Ponder, Katherine P

2013-11-01

Mucopolysaccharidosis VII (MPS VII) is due to the deficient activity of β-glucuronidase (GUSB) and results in the accumulation of glycosaminoglycans (GAGs) in lysosomes and multisystemic disease with cardiovascular manifestations. The goal here was to determine the pathogenesis of mitral valve (MV) disease in MPS VII dogs. Untreated MPS VII dogs had a marked reduction in the histochemical signal for structurally-intact collagen in the MV at 6 months of age, when mitral regurgitation had developed. Electron microscopy demonstrated that collagen fibrils were of normal diameter, but failed to align into large parallel arrays. mRNA analysis demonstrated a modest reduction in the expression of genes that encode collagen or collagen-associated proteins such as the proteoglycan decorin which helps collagen fibrils assemble, and a marked increase for genes that encode proteases such as cathepsins. Indeed, enzyme activity for cathepsin B (CtsB) was 19-fold normal. MPS VII dogs that received neonatal intravenous injection of a gamma retroviral vector had an improved signal for structurally-intact collagen, and reduced CtsB activity relative to that seen in untreated MPS VII dogs. We conclude that MR in untreated MPS VII dogs was likely due to abnormalities in MV collagen structure. This could be due to upregulation of enzymes that degrade collagen or collagen-associated proteins, to the accumulation of GAGs that compete with proteoglycans such as decorin for binding to collagen, or to other causes. Further delineation of the etiology of abnormal collagen structure may lead to treatments that improve biomechanical properties of the MV and other tissues. © 2013.

Pathogenesis of Mitral Valve Disease in Mucopolysaccharidosis VII Dogs

PubMed Central

Bigg, Paul W.; Baldo, Guilherme; Sleeper, Meg M.; O'Donnell, Patricia A.; Bai, Hanqing; Rokkam, Venkata R.P.; Liu, Yuli; Wu, Susan; Giugliani, Roberto; Casal, Margret L.; Haskins, Mark E.; Ponder, Katherine P.

2013-01-01

Mucopolysaccharidosis VII (MPS VII) is due to deficient activity of β-glucuronidase (GUSB) and results in the accumulation of glycosaminoglycans (GAGs) in lysosomes and multisystemic disease with cardiavascular manifestations. The goal here was to determine the pathogenesis of mitral valve (MV) disease in MPS VII dogs. Untreated MPS VII dogs had a marked reduction in the histochemical signal for structurally-intact collagen in the MV at 6 months of age, when mitral regurgitation had developed. Electron microscopy demonstrated that collagen fibrils were of normal diameter, but failed to align into large parallel arrays. mRNA analysis demonstrated a modest reduction in the expression of genes that encode collagen or collagen-associated proteins such as the proteoglycan decorin which helps collagen fibrils assemble, and a marked increase for genes that encode proteases such as cathepsins. Indeed, enzyme activity for cathepsin B (CtsB) was 19-fold normal. MPS VII dogs that received neonatal intravenous injection of a gamma retroviral vector had an improved signal for structurally-intact collagen, and reduced CtsB activity relative to that seen in untreated MPS VII dogs. We conclude that MR in untreated MPS VII dogs was likely due to abnormalities in MV collagen structure. This could be due to upregulation of enzymes that degrade collagen or collagen-associated proteins, to the accumulation of GAGs that compete with proteoglycans such as decorin for binding to collagen, or to other causes. Further delineation of the etiology of abnormal collagen structure may lead to treatments that improve biomechanical properties of the MV and other tissues. PMID:23856419

Hydrophilic acrylic intraocular lens optic and haptics opacification in a diabetic patient: bilateral case report and clinicopathologic correlation.

PubMed

Pandey, Suresh K; Werner, Liliana; Apple, David J; Kaskaloglu, Mahmut

2002-11-01

To report clinicopathologic and ultrastructural features of two opacified single-piece hydrophilic acrylic intraocular lenses (IOLs) explanted from a diabetic patient. Interventional case report with clinicopathologic correlation. A 64-year-old white female underwent phacoemulsification and implantation of a single-piece hydrophilic acrylic lens (SC60B-OUV; Medical Developmental Research, Inc., Clear Water, FL) in October 1998 in the left eye and in July 1999 in the right eye. The best-corrected visual acuity after surgery was 20/60 in the left eye and 20/50 in the right eye. The patient had a marked decrease in visual acuity in June 2000 as a result of a milky, white opalescence of both lenses. Intraocular lens explantation and exchange was performed in both eyes and the explanted IOLs were submitted to our center for detailed pathologic, histochemical, and ultrastructural evaluation. They were stained with alizarin red and the von Kossa method for calcium, and also underwent scanning electron microscopy and energy dispersive radiograph spectroscopy to ascertain the nature of the deposits leading to opacification. Documentation of calcium deposits confirmed by histochemical stains and surface analyses. Opacification of the IOL was found to be the cause of decreased visual acuity. The opacification involved both the IOL optic and the haptics in the left eye and was confined to the IOL optic in the right eye. Histochemical and ultrastructural analyses revealed that the opacity was caused by deposition of calcium and phosphate within the lens optic and haptics. There are two features that distinguish this case from those reported earlier. This is the first clinicopathologic report of lens opacification that has involved completely the lens optic and the haptics. Second, these two explanted IOLs document the first bilateral case. This process of intraoptic and haptic opacification represents dystrophic calcification of unknown cause. Diabetic patients appear to be more severely and more often affected by lens opacification. Long-term follow-up of diabetic patients implanted with this IOL design should be maintained by surgeons and manufacturers.

Centronuclear myopathy in a Border collie dog.

PubMed

Eminaga, S; Cherubini, G B; Shelton, G D

2012-10-01

A two-year old, male entire Border collie was presented with a one-year history of exercise-induced collapsing on the pelvic limbs. Physical examination revealed generalised muscle atrophy. Neurological examination supported a generalised neuromuscular disorder. Electromyography revealed spontaneous electrical activity in almost all muscles. Unfixed and formaldehyde-fixed biopsy samples were collected from the triceps brachii, longissimus and vastus lateralis muscles. Histopathological, histochemical and ultrastructural examinations of biopsy specimens were consistent with either centronuclear or myotubular myopathy. The dog clinically improved with supportive treatment with L-carnitine, co-enzyme Q10 and vitamin B compound. To the authors' knowledge, this is the first report of centronuclear/myotubular myopathy in a Border collie. © 2012 British Small Animal Veterinary Association.

Structural-metabolic organization of field 4 of the cat brain in normal conditions and after unilateral enucleation of the eye.

PubMed

Zykin, P A

2005-01-01

Comparative data on the structural-metabolic organization of field 4 of the cat brain in normal conditions and after unilateral enucleation of the eye are presented. Cytochrome oxidase was detected histochemically. Data were processed by a computerized method using an original video capture system. Data were obtained demonstrating the uneven distribution of enzyme along sublayer IlIb of field 4 in animals with unilateral enucleation. A hypothesis based on published data is suggested whereby the alternation of high- and low-reactive areas is evidence for the ordering of the retinal representations of the right and left eyes in the sensorimotor cortex.

Fatal human eosinophilic meningo-encephalitis caused by CNS co-infection with Halicephalobus gingivalis and West Nile virus.

PubMed

Anwar, M A; Gokozan, H N; Ball, M K; Otero, J; McGwire, B S

2015-10-01

The saprophytic nematode Halicephalobus is a rare cause of fatal human meningo-encephalitis, and West Nile virus is neurotropic flavivirus implicated in a variety of clinical neurologic syndromes. Here we report a case of rapidly progressive CNS encephalopathy and death. Serologic, immuno-histochemical, histopathologic and nucleic acid studies demonstrate the presence of active Halicephalobus and West Nile virus in the CNS tissue. This is the first reported case of co-infection with these neurotropic pathogens. Copyright © 2015. Published by Elsevier Ireland Ltd.

Carbon Dioxide Metabolism in Leaf Epidermal Tissue 1

PubMed Central

Willmer, C. M.; Pallas, J. E.; Black, C. C.

1973-01-01

A number of plant species were surveyed to obtain pure leaf epidermal tissue in quantity. Commelina communis L. and Tulipa gesnariana L. (tulip) were chosen for further work. Chlorophyll a/b ratios of epidermal tissues were 2.41 and 2.45 for C. communis and tulip, respectively. Phosphoenolpyruvate carboxylase, ribulose-1,5-diphosphate carboxylase, malic enzyme, and NAD+ and NADP+ malate dehydrogenases were assayed with epidermal tissue and leaf tissue minus epidermal tissue. In both species, there was less ribulose 1,5-diphosphate than phosphoenolpyruvate carboxylase activity in epidermal tissue whether expressed on a protein or chlorophyll basis whereas the reverse was true for leaf tissue minus epidermal tissue. In both species, malic enzyme activities were higher in epidermal tissue than in the remaining leaf tissue when expressed on a protein or chlorophyll basis. In both species, NAD+ and NADP+ malate dehydrogenase activities were higher in the epidermal tissue when expressed on a chlorophyll basis; however, on a protein basis, the converse was true. Microautoradiography of C. communis epidermis and histochemical tests for keto acids suggested that CO2 fixation occurred predominantly in the guard cells. The significance and possible location of the enzymes are discussed in relation to guard cell metabolism. Images PMID:16658581

Fluorometric method of quantitative cell mutagenesis

DOEpatents

Dolbeare, Frank A.

1982-01-01

A method for assaying a cell culture for mutagenesis is described. A cell culture is stained first with a histochemical stain, and then a fluorescent stain. Normal cells in the culture are stained by both the histochemical and fluorescent stains, while abnormal cells are stained only by the fluorescent stain. The two stains are chosen so that the histochemical stain absorbs the wavelengths that the fluorescent stain emits. After the counterstained culture is subjected to exciting light, the fluorescence from the abnormal cells is detected.

Fluorometric method of quantitative cell mutagenesis

DOEpatents

Dolbeare, F.A.

1980-12-12

A method for assaying a cell culture for mutagenesis is described. A cell culture is stained first with a histochemical stain, and then a fluorescent stain. Normal cells in the culture are stained by both the histochemical and fluorescent stains, while abnormal cells are stained only by the fluorescent stain. The two stains are chosen so that the histochemical stain absorbs the wavelengths that the fluorescent stain emits. After the counterstained culture is subjected to exciting light, the fluorescence from the abnormal cells is detected.

The action of aminoguanidine on the liver of trained diabetic rats

PubMed Central

2013-01-01

Background This study evaluated the effect of aminoguanidine on liver of diabetic rats subject to physical exercises using histological and histochemical techniques. Methods The rats used in this study were divided into five groups: sedentary control, sedentary diabetic, trained diabetic, sedentary diabetic and treated with aminoguanidine, trained diabetic and treated with aminoguanidine. Results The results showed no effect of aminoguanidine on the liver tissue, although there was improvement with exercise training showing cytological, morpho-histological and histochemical alterations in liver cells of animals from groups trained diabetic and/or treated diabetic compared to those individuals in the sedentary control and sedentary diabetic. These changes included: hepatocytes hypertrophy, presence and distribution of polysaccharides in the hepatocytes cytoplasm and, especially, congestion of the liver blood vessels. Conclusion Our results suggest that aminoguanidine is not hepatotoxic, when used at dosage of 1 g/L for the treatment of diabetes complications, and confirmed that the practice of moderate physical exercise assuaged the damage caused by diabetes without the use of insulin. PMID:23837632

Japanese quail acute exposure to methamidophos: experimental design, lethal, sub-lethal effects and cholinesterase biochemical and histochemical expression.

PubMed

Foudoulakis, Manousos; Balaskas, Christos; Csato, Attila; Szentes, Csaba; Arapis, Gerassimos

2013-04-15

We exposed the Japanese quail (Coturnix coturnix japonica) to the organophosphate methamidophos using acute oral test. Mortality and sub-lethal effects were recorded in accordance to internationally accepted protocols. In addition cholinesterases were biochemically estimated in tissues of the quail: brain, liver and plasma. Furthermore, brain, liver and duodenum cryostat sections were processed for cholinesterase histochemistry using various substrates and inhibitors. Mortalities occurred mainly in the first 1-2h following application. Sub-lethal effects, such as ataxia, ruffled feathers, tremor, salivation and reduced or no reaction to external stimuli were observed. Biochemical analysis in the brain, liver and plasma indicates a strong cholinesterase dependent inhibition with respect to mortality and sub-lethal effects of the quail. The histochemical staining also indicated a strong cholinesterase inhibition in the organs examined and the analysis of the stained sections allowed for an estimation and interpretation of the intoxication effects of methamidophos, in combination with tissue morphology visible by Haematoxylin and Eosin staining. We conclude that the use of biochemistry and histochemistry for the biomarker cholinesterase, may constitute a significantly novel approach for understanding the results obtained by the acute oral test employed in order to assess the effects of methamidophos and other chemicals known to inhibit this very important nervous system enzyme. Copyright © 2012 Elsevier B.V. All rights reserved.

Functional characterization of cinnamyl alcohol dehydrogenase and caffeic acid O-methyltransferase in Brachypodium distachyon

PubMed Central

2013-01-01

Background Lignin is a significant barrier in the conversion of plant biomass to bioethanol. Cinnamyl alcohol dehydrogenase (CAD) and caffeic acid O-methyltransferase (COMT) catalyze key steps in the pathway of lignin monomer biosynthesis. Brown midrib mutants in Zea mays and Sorghum bicolor with impaired CAD or COMT activity have attracted considerable agronomic interest for their altered lignin composition and improved digestibility. Here, we identified and functionally characterized candidate genes encoding CAD and COMT enzymes in the grass model species Brachypodium distachyon with the aim of improving crops for efficient biofuel production. Results We developed transgenic plants overexpressing artificial microRNA designed to silence BdCAD1 or BdCOMT4. Both transgenes caused altered flowering time and increased stem count and weight. Downregulation of BdCAD1 caused a leaf brown midrib phenotype, the first time this phenotype has been observed in a C3 plant. While acetyl bromide soluble lignin measurements were equivalent in BdCAD1 downregulated and control plants, histochemical staining and thioacidolysis indicated a decrease in lignin syringyl units and reduced syringyl/guaiacyl ratio in the transgenic plants. BdCOMT4 downregulated plants exhibited a reduction in total lignin content and decreased Maule staining of syringyl units in stem. Ethanol yield by microbial fermentation was enhanced in amiR-cad1-8 plants. Conclusion These results have elucidated two key genes in the lignin biosynthetic pathway in B. distachyon that, when perturbed, may result in greater stem biomass yield and bioconversion efficiency. PMID:23902793

Histochemical localisation of carbonic anhydrase in the inner ear of developing cichlid fish, Oreochromis mossambicus

NASA Astrophysics Data System (ADS)

Beier, M.; Hilbig, R.; Anken, R.

2008-12-01

Inner ear otolith growth in terms of mineralisation mainly depends on the enzyme carbonic anhydrase (CAH). CAH is located in specialised, mitochondria-rich macular cells (ionocytes), which are involved in the endolymphatic ion exchange, and the enzyme is responsible for the provision of the pH-value necessary for otolithic calcium carbonate deposition. In the present study, for the first time the localisation of histochemically demonstrated CAH was analysed during the early larval development of a teleost, the cichlid fish Oreochromis mossambicus. CAH-reactivity was observed already in stage 7 animals (onset of otocyst development; staging follows Anken et al. [Anken, R., Kappel, T., Slenzka, K., Rahmann, H. The early morphogenetic development of the cichlid fish, Oreochromis mossambicus (Perciformes, Teleostei). Zool. Anz. 231, 1-10, 1993]). Neuroblasts (from which sensory and supporting cells are derived) proved to be CAH-positive. Already at stage 12 (hatch), CAH-positive regions could be attributed to ionocyte containing regions both in the so-called meshwork and patches area of the macula (i.e., clearly before ionocytes can be identified on ultrastructural level or by employing immunocytochemistry). In contrast to the circumstances observed in mammalian species, sensory hair cells stained negative for CAH in the cichlid. With the onset of stage 16 (finray primordia in dorsal fin, yolk-sac being increasingly absorbed), CAH-reactivity was observed in the vestibular nerve. This indicates the onset of myelinisation and thus commencement of operation. The localisation of CAH in the inner ear of fish (especially the differences in comparison to mammals) is discussed on the basis of its role in otolith calcification. Since the vestibular system is a detector of acceleration and thus gravity, also aspects regarding effects of altered gravity on CAH and hence on the mineralisation of otoliths in an adaptive process are addressed.

Underbody Blast Models of TBI Caused by Hyper-Acceleration and Secondary Head Impact

DTIC Science & Technology

2014-02-01

Cleveland RO, Tanzi RE, Stanton PK, McKee AC. (2012) Chronic traumatic encephalopathy in blast-exposed military veterans and a blast neurotrauma mouse model...exposure, remove brains, and process for electron microscopic analysis of cyto- and axonal ultrastructure and for histochemical evidence of acute ...of Trauma and Acute Care Surgery (see Appendix). These observations include increased axonopathy (silver staining) in the cerebellum and astrocyte

Immunolocalization of steroidogenic enzymes in the vaginal mucous of Galea spixii during the estrous cycle.

PubMed

Dos Santos, Amilton Cesar; Conley, Alan James; de Oliveira, Moacir Franco; Oliveira, Gleidson Benevides; Viana, Diego Carvalho; Assis Neto, Antônio Chaves de

2017-04-24

The synthesis of sex steroids is controlled by several enzymes such as17α-hydroxylase cytochrome P450 (P450c17) catalyzing androgen synthesis and aromatase cytochrome P450 (P450arom) catalyzing estrogen synthesis, both of which must complex with the redox partner NADPH-cytochrome P450 oxidoreductase (CPR) for activity. Previous studies have identified expression of steroidogenic enzymes in vaginal tissue, suggesting local sex steroid synthesis. The current studies investigate P450c17, P450aromatase and CPR expression in vaginal mucosa of Galea spixii (Spix cavy) by immuno-histochemical and western immunoblot analyses. Stages of estrous cyclicity were monitored by vaginal exfoliative cytology. After euthanasia, vaginal tissues were retrieved, fixed and frozen at diestrus, proestrus, estrus and metestrus. The ovaries and testis were used as positive control tissues for immunohistochemistry. Data from cytological study allowed identification of different estrous cycle phases. Immunohistochemical analysis showed different sites of expression of steroidogenic enzymes along with tissue response throughout different phases of the estrous cycle. However, further studies are needed to characterize the derived hormones synthesized by, and the enzymes activities associated with, vaginal tissues. Current results not only support the expression of enzymes involved in sex steroid synthesis in the wall of the vagina, they also indicate that expression changes with the stage of the cycle, both the levels and types of cells exhibiting expression. Thus, changes in proliferation of vaginal epithelial cells and the differentiation of the mucosa may be influenced by local steroid synthesis as well as circulating androgens and estrogens.

Comparison between histochemical and immunohistochemical methods for diagnosis of sporotrichosis.

PubMed Central

Marques, M E; Coelho, K I; Sotto, M N; Bacchi, C E

1992-01-01

AIMS: To compare the efficacy of histochemical and immunohistochemical methods in detecting forms of Sporothrix schenckii in tissue. METHODS: Thirty five cutaneous biopsy specimens from 27 patients with sporotrichosis were stained by histochemical haematoxylin and eosin, periodic acid Schiff, and Gomori's methenamine silver methods and an immunohistochemical (avidin-biotin complex immunoperoxidase) (ABC) technique associated with a newly produced rabbit polyclonal antibody anti-Sporothrix schenckii. RESULTS: A total of 29 (83%) cases were positive by the ABC method used in association with anti-Sporothrix schenckii rabbit polyclonal antibodies. Histochemical methods, using silver staining, periodic acid Schiff, and conventional haematoxylin and eosin detected 37%, 23%, and 23% of forms of S schenckii, respectively. The ABC technique was significantly more reliable than periodic acid Schiff and silver staining techniques. CONCLUSIONS: It is concluded that immunostaining is an easy and rapid method which can efficiently increase the accuracy of the diagnosis of sporotrichosis in human tissue. Images PMID:1479036

Histochemical stains as promising means for the laser histochemical surgery of a number of pathologies

NASA Astrophysics Data System (ADS)

Piruzyan, L. A.; Mikhailovskiy, Ye. M.; Piruzyan, A. L.

1999-12-01

The directions of laboratory and clinical studies oriented to experimental confirmation of the priority concept of `laser histochemical surgery' are presented. The goal of the studies is reproduction on experimental model of a number of pathologies (in vivo and in vitro) of the `sensitization to laser radiation by staining' effect. Testing of the histochemical stains as sensitizers to laser irradiation of their `address substrates', i.e. vitally stained intracellular structures which participate in the pathologic processes evolution is under planning. The processes include: (a) metabolic disorders in the brain cells, i.e. disseminated sclerosis; (b) generalized metabolic disorders- -mucopolysaccharidosis and collagenosises (periarteritis nodosa, rheumatism, rheumatoid arthritis, sclerodermia); (3) metabolic disorders in individual organs--vessel atherosclerosis, hypercholesterolemia, myocardial infarction, cardiosclerosis, caries and parodontosis. The conditions of the studies are detailed in the recommendations along the positions: (1) disease name; (2) disease characteristics: (a) pathomorphologic, (b) biochemical; (3) stains revealing the disease signs and recommended for testing; (4) `address substrates' of the stains that are targets for laser radiation; (5) lasers recommended for the testing after the cells staining in vivo in the corresponding pathology; (6) experimental models of the pathologies suggested for the testing; (7) criteria of the stain efficiency as target sensitizer to the laser light (criteria of the `laser sensitization by staining' efficiency). Possible perspectives for the experimental clinical medicine are indicated of common histochemical stains and lasers use and of practice introduction of the `laser histochemical surgery' in the case the described concept is confirmed in experiments and clinically.

Induction of resistance to diabetes in non-obese diabetic mice by targeting CD44 with a specific monoclonal antibody

PubMed Central

Weiss, Lola; Slavin, Shimon; Reich, Shoshana; Cohen, Patrizia; Shuster, Svetlana; Stern, Robert; Kaganovsky, Ella; Okon, Elimelech; Rubinstein, Ariel M.; Naor, David

2000-01-01

Inflammatory destruction of insulin-producing β cells in the pancreatic islets is the hallmark of insulin-dependent diabetes mellitus, a spontaneous autoimmune disease of non-obese diabetic mice resembling human juvenile (type I) diabetes. Histochemical analysis of diabetic pancreata revealed that mononuclear cells infiltrating the islets and causing autoimmune insulitis, as well as local islet cells, express the CD44 receptor; hyaluronic acid, the principal ligand of CD44, is detected in the islet periphery and islet endothelium. Injection of anti-CD44 mAb 1 hr before cell transfer of diabetogenic splenocytes and subsequently on alternate days for 4 weeks induced considerable resistance to diabetes in recipient mice, reflected by reduced insulitis. Contact sensitivity to oxazolone was not influenced by this treatment. A similar antidiabetic effect was observed even when the anti-CD44 mAb administration was initiated at the time of disease onset: i.e., 4–7 weeks after cell transfer. Administration of the enzyme hyaluronidase also induced appreciable resistance to insulin-dependent diabetes mellitus, suggesting that the CD44–hyaluronic acid interaction is involved in the development of the disease. These findings demonstrate that CD44-positive inflammatory cells may be a potential therapeutic target in insulin-dependent diabetes. PMID:10618410

Differential expression of the polysialyl capsule during blood-to-brain transit of neuropathogenic Escherichia coli K1.

PubMed

Zelmer, Andrea; Bowen, Mark; Jokilammi, Anne; Finne, Jukka; Luzio, J Paul; Taylor, Peter W

2008-08-01

Escherichia coli K1 isolates synthesize a polysialic acid (polySia) capsule, are components of the adult gastrointestinal microbiota and may cause lethal bacteraemia and meningitis if acquired maternally by newborn infants. We used a neonatal rat pup K1 infection model to establish that prompt administration of a selective capsule depolymerase reverses the bacteraemic state and prevents death of almost all pups. In untreated animals, bacteria colonize the gastrointestinal tract and gain entry to the blood compartment, where they express the non-O-acetylated form of polySia. The bacteria invade the major organs of the host; histological and histochemical analysis of brain sections revealed that at least some bacteria enter the central nervous system through the blood-cerebrospinal fluid barrier at the choroid plexus prior to colonization of the meninges. Once in this location, they cease expression of polySia. The unexpected abrogation of polySia, a factor associated with the pathogenesis of meningitis and essential for transit through the blood, suggests that the neuropathogen dispenses with its protective capsule once it has colonized protected niches. Thus, systemic infections due to encapsulated pathogens may be resolved by capsule depolymerization only if the enzyme modifies the bacteria whilst they are in the blood compartment.

[Localization of 17 beta-hydroxysteroid dehydrogenase in the gonads of bivalve mollusks--the sea pecten (Patinopecten yessoensis Jay) and Gray's mussel (Crenomytilus grayanus Dunker)].

PubMed

Varaksina, G S; Varaksin, A A

1988-11-01

Presence of 17 beta-hydroxysteroid dehydrogenase (17 beta-HSD) in testes and ovaries of the common mussels--Patinopecten yessoensis (Jay) and Crenomytilus grayanus (Dunker) has been demonstrated histochemically. The enzyme is revealed in some granular amebocytes and germ cells. In growing oocytes its activity is higher that in oocytes completed their growth. 17 beta-HSD is revealed electron microscopically near agranular endoplasmic reticulum, or on the external surface of its membranes and in globules, possessing, evidently, lipid nature. The data obtained demonstrate that synthesis and metabolism of steroid hormones are possible both in additional gonadal elements and in germ cells of the animals investigated.

Histochemical and ultrastructural studies on the calcareous corpuscles and eggs of Taenia taeniaeformis and Dipylidium caninum.

PubMed

Khalifa, Refaat M A; Mazen, Nawal A M; Marawan, Aziza M A; Thabit, Hasnaa T M

2011-08-01

Calcareous corpuscles were noticed by several previous workers to be present in larval and adult cestodes without knowing their function. However, nothing was mentioned in the available literature about distribution of these corpuscles and their density, structure and composition in different parts of the body of different cestodes. Hence, in the present work, a comparative study of their distribution, density, histochemical and ultrastructural characters in different parts of the body was performed in Taenia taeniaeformis and Dipylidium caninum. Due to the presence of the eggs in their gravid segments, their histochemical and ultrastructural characteristics were also studied. It was found that the size, location and density of the calcareous bodies were different in different body parts of the same and the other cestode. Histochemically, the main component of these corpuscles was calcium; while other constituents as polysaccharides, lipids, protrins and mucopolysaccharides were found in their outer rim. Ultrastructurally, they were quite similar in the two studied cestodes and different stages of their development were exhibited. Histochemically, the eggs of both cestodes were similar in their contents. However, some ultrastructural differences have been demonstrated particularly in relation to the size and shape of the rods in the embryophore and the structures in between the embryophore and onchosphere.

Agaricus blazei Murill extract abrogates CCl4-induced liver injury in rats.

PubMed

Wu, Ming-Fang; Hsu, Yu-Ming; Tang, Ming-Chu; Chen, Hsueh-Chin; Chung, Jing-Gung; Lu, Hsu-Feng; Lin, Jing-Pin; Tang, Nou-Ying; Yeh, Chun; Yeh, Ming-Yang

2011-01-01

Agaricus blazei Murill (ABM) is enriched with polysaccharides, lipids, vitamins, fibers and minerals. Many studies have shown that ABM possesses immune-enhancing and anti-tumor effects. However, little is known about its protective effects on liver function. We employed carbon tetrachloride (CCl(4)) to induce hepatic fibrosis in a rat model to examine the protective effects of ABM on the liver in this study. The experiments included non-treatment control, CCl(4)-only control, and treatment with 200 mg and 2,000 mg of ABM extracts (per kilogram rat weight). All groups other than the non-treatment control were treated with intraperitoneal injections of CCl(4) twice a week. Experimental and control rats were tube-fed with experimental ABM extracts or double-distilled water, respectively, on the remaining four days each week. The whole experimental protocol lasted 8 weeks; blood and liver samples were collected for biochemical and tissue histochemical analysis. Plasma alanine aminotransferase and aspartate aminotransferase, and the activities of the anti-oxidative enzymes glutathione peroxidase, superoxide dismutase and catalase in the liver were measured. We found that high-dose ABM treatment reduced hepatic necrosis and fibrosis caused by CCl(4) in comparison with the CCl(4) control group. ALT and AST activities in the sera collected from ABM-treated rats were lower than those in the CCl(4) control rats. These results suggested that ABM extract was capable of either enhancing liver recovering from CCl(4) damage or attenuating CCl(4) toxicity. Results of anti-oxidative enzyme activity analysis showed no apparent differences among ABM-treated groups and CCl(4) control groups, indicating that removal of free radicals does not explain the protective/recovery effects observed in this study.

Evolution of the localisation and composition of phenolics in grape skin between veraison and maturity in relation to water availability and some climatic conditions.

PubMed

Cadot, Yves; Chevalier, Michel; Barbeau, Gerard

2011-08-30

Several studies have investigated the composition of phenolics in grape skin during grape maturation under various conditions of light exposure, water stress, nitrogen supply and mineral nutrition, but their localisation during berry development is not well known. In this study the composition and localisation of proanthocyanidins were monitored for three years on four plots known to induce a distinctive behaviour of the vine (Cabernet Franc). The composition of phenolics was determined by spectrophotometry; also, in one year, proanthocyanidins were determined by high-performance liquid chromatography. Further information was obtained histochemically by means of toluidine blue O staining and image analysis. The results indicated that clear differences in phenolic quantification existed between the biochemical and histochemical approaches; the proportion of cells without phenolics was not linked with the quantity determined by the analytical methods used. The histochemical method showed the evolution of the localisation and typology of cells with and without phenolics during ripening. The number of cells without any phenolic compounds appeared to be very dependent on the mesoclimatic conditions and only slightly dependent on the site water status. Clear differences in phenolic quantification existed between the biochemical and histochemical approaches; the proportion of cells with phenolics was not linked with the quantity determined by biochemistry. The histochemical method showed an evolution of the localisation and typology of cells with and without phenolics in which mesoclimatic conditions were the most influential factor. Finally, the study showed some advantages of the histochemical approach: it gives information about the anatomy of the tissue as well as the nature and distribution of some of the large macromolecules and allows reconstruction of the three-dimensional plant structure. Copyright © 2011 Society of Chemical Industry.

Distribution of glycogen phosphorylase and cytochrome oxidase in the central nervous system of the turtle Trachemys dorbigni.

PubMed

Partata, W A; Krepsky, A M; Xavier, L L; Marques, M; Achaval, M

1999-10-01

Glycogen phosphorylase (GP) and cytochrome oxidase (CO) activities were mapped histochemically in the brain of the turtle Trachemys dorbigni. In the telencephalon, both activities occurred in the olfactory bulb, in all cortical areas, in the dorsal ventricular ridge, striatum, primordium hippocampi and olfactory tubercle. In the diencephalon, they were identified in some areas of the hypothalamus, and in rotundus and geniculate nuclei. Both reactions were detected in the oculomotor, trochlear, mesencephalic trigeminal nuclei, the nucleus of the posterior commissure, torus semicircularis, substantia nigra and ruber and isthmic nuclei of the mesencephalon. In all layers of the optic tectum GP activity was found, but CO only labelled the stratum griseum centrale. In the medulla oblonga both enzymes appear in the reticular, raphe and vestibular nuclei, locus coeruleus and nuclei of cranial nerves. In the cerebellum, the granular and molecular layers, and the deep cerebellar nuclei were positive for both enzymes. The Purkinje cells were only reactive for CO. In the spinal cord, motor and commissural neurones exhibited a positive reaction for the two enzymes. However, CO also occurred in the marginal nucleus and in the lateral funiculus. These results may be useful as a basis for subsequent studies on turtle brain metabolism.

Polyamine metabolism influences antioxidant defense mechanism in foxtail millet (Setaria italica L.) cultivars with different salinity tolerance.

PubMed

Sudhakar, Chinta; Veeranagamallaiah, Gounipalli; Nareshkumar, Ambekar; Sudhakarbabu, Owku; Sivakumar, M; Pandurangaiah, Merum; Kiranmai, K; Lokesh, U

2015-01-01

Polyamines can regulate the expression of antioxidant enzymes and impart plants tolerance to abiotic stresses. A comparative analysis of polyamines, their biosynthetic enzymes at kinetic and at transcriptional level, and their role in regulating the induction of antioxidant defense enzymes under salt stress condition in two foxtail millet (Setaria italica L.) cultivars, namely Prasad, a salt-tolerant, and Lepakshi, a salt-sensitive cultivar was conducted. Salt stress resulted in elevation of free polyamines due to increase in the activity of spermidine synthase and S-adenosyl methionine decarboxylase enzymes in cultivar Prasad compared to cultivar Lepakshi under different levels of NaCl stress. These enzyme activities were further confirmed at the transcript level via qRT-PCR analysis. The cultivar Prasad showed a greater decrease in diamine oxidase and polyamine oxidase activity, which results in the accumulation of polyamine pools over cultivar Lepakshi. Generation of free radicals, such as O 2 (·-) and H2O2, was also analyzed quantitatively. A significant increase in O 2 (·-) and H2O2 in the cultivar Lepakshi compared with cultivar Prasad was recorded in overall pool sizes. Further, histochemical staining showed lesser accumulation of O 2 (·-) and of H2O2 in the leaves of cultivar Prasad than cultivar Lepakshi. Our results also suggest the ability of polyamine oxidation in regulating the induction of antioxidative defense enzymes, which involve in the elimination of toxic levels of O 2 (·-) and H2O2, such as Mn-superoxide dismutase, catalase and ascorbate peroxidase. The contribution of polyamines in modulating antioxidative defense mechanism in NaCl stress tolerance is discussed.

Immunolocalization of choline acetyltransferase of common type in the central brain mass of Octopus vulgaris

PubMed Central

Casini, A.; Vaccaro, R.; D'Este, L.; Sakaue, Y.; Bellier, J.P.; Kimura, H.; Renda, T.G.

2012-01-01

Acetylcholine, the first neurotransmitter to be identified in the vertebrate frog, is widely distributed among the animal kingdom. The presence of a large amount of acetylcholine in the nervous system of cephalopods is well known from several biochemical and physiological studies. However, little is known about the precise distribution of cholinergic structures due to a lack of a suitable histochemical technique for detecting acetylcholine. The most reliable method to visualize the cholinergic neurons is the immunohistochemical localization of the enzyme choline acetyltransferase, the synthetic enzyme of acetylcholine. Following our previous study on the distribution patterns of cholinergic neurons in the Octopus vulgaris visual system, using a novel antibody that recognizes choline acetyltransferase of the common type (cChAT), now we extend our investigation on the octopus central brain mass. When applied on sections of octopus central ganglia, immunoreactivity for cChAT was detected in cell bodies of all central brain mass lobes with the notable exception of the subfrontal and subvertical lobes. Positive varicosed nerves fibers where observed in the neuropil of all central brain mass lobes. PMID:23027350

Immunolocalization of choline acetyltransferase of common type in the central brain mass of Octopus vulgaris.

PubMed

Casini, A; Vaccaro, R; D'Este, L; Sakaue, Y; Bellier, J P; Kimura, H; Renda, T G

2012-07-19

Acetylcholine, the first neurotransmitter to be identified in the vertebrate frog, is widely distributed among the animal kingdom. The presence of a large amount of acetylcholine in the nervous system of cephalopods is well known from several biochemical and physiological studies. However, little is known about the precise distribution of cholinergic structures due to a lack of a suitable histochemical technique for detecting acetylcholine. The most reliable method to visualize the cholinergic neurons is the immunohistochemical localization of the enzyme choline acetyltransferase, the synthetic enzyme of acetylcholine. Following our previous study on the distribution patterns of cholinergic neurons in the Octopus vulgaris visual system, using a novel antibody that recognizes choline acetyltransferase of the common type (cChAT), now we extend our investigation on the octopus central brain mass. When applied on sections of octopus central ganglia, immunoreactivity for cChAT was detected in cell bodies of all central brain mass lobes with the notable exception of the subfrontal and subvertical lobes. Positive varicosed nerves fibers where observed in the neuropil of all central brain mass lobes.

ISOLATION AND PROPERTIES OF LIVER CELL NUCLEOLI

PubMed Central

Monty, K. J.; Litt, M.; Kay, E. R. M.; Dounce, A. L.

1956-01-01

1. The significance of the term nucleolus has been discussed. 2. A detailed method for the isolation of nucleoli from already isolated rat or cat liver nuclei has been presented. 3. The presence of DNA in isolated liver cell nucleoli has been indicated by histochemical methods. 4. The percentages of DNA and RNA in the isolated nucleoli have been determined by chemical analysis. 5. The specific activities of aldolase, arginase, and catalase have been determined for two subnuclear fractions and for the isolated nucleoli of rat and cat liver, and the relative amounts of these enzymes in the same subnuclear fractions and nucleoli of rat liver have been measured. 6. The significance of the above findings has been discussed and consideration has been given to what types of isolated nuclei might best serve as starting material for the isolation of nucleoli. 7. A new hypothesis has been presented that nucleoli of the liver cell type may function primarily in furnishing (directly or indirectly) templates for the synthesis of the particular enzymes that must govern the chemistry of mitosis. PMID:13319377

Lectin histochemistry and alkaline phosphatase activity in the pia mater vessels of spontaneously hypertensive rats (SHR).

PubMed

Szumańska, G; Gadamski, R

1992-01-01

Some lectins were used to study the localization of sugar residues on the endothelial cell surface in the pia mater blood vessels of control (WKY) and hypertensive rats (SHR). The lectins tested recognized the following residues: beta-D-galactosyl (Ricinus communis agglutinin 120, RCA-1), alpha-L-fucosyl (Ulex europaeus agglutinin, UEA-1), N-acetylglucosaminyl and sialyl (Wheat germ agglutinin, WGA), N-glycolyl-neuraminic acid (Limax flavus agglutinin, LFA), and N-acetyl-D-galactosaminyl (Helix pomatia agglutinin, HPA). Several differences were revealed in the presence of sugar receptors on the surface of endothelial cells between the control and the hypertensive rats. Our studies showed also differences in the localization of the tested glycoconjugates between pial capillaries, small, medium-size and large pial arteries. The histochemical evaluation of alkaline phosphatase revealed an increased activity of the enzyme in the pial vessels of SHRs as compared with control rats with a similar localization of the enzyme activity. Some differences in the distribution of lectin binding sites and alkaline phosphatase activity could be associated with the different functions of particular segments of the pial vascular network.

Influence of long-term hyper-gravity on the reactivity of succinic acid dehydrogenase and NADPH-diaphorase in the central nervous system of fish: a histochemical study

NASA Astrophysics Data System (ADS)

Anken, R. H.; Rahmann, H.

In the course of a densitometric evaluation, the histochemically demonstrated reactivity of succinic acid dehydrogenase (SDH) and of NADPH-diaphorase (NADPHD) was determined in different brain nuclei of two teleost fish (cichlid fish Oreochromis mossambicus, swordtail fish Xiphophorus helleri), which had been kept under 3g hyper-gravity for 8 days. SDH was chosen since it is a rate limiting enzyme of the Krebs cycle and therefore it is regarded as a marker for metabolic and neuronal activity. NADPHD reactivity reflects the activity of nitric oxide synthase. Nitric oxide (NO) is a gaseous intercellular messenger that has been suggested to play a major role in several different in vivo models of neuronal plasticity including learning. Within particular vestibulum-connected brain centers, significant effects of hyper-gravity were obtained, e.g., in the magnocellular nucleus, a primary vestibular relay ganglion of the brain stem octavolateralis area, in the superior rectus subdivision of the oculomotoric nucleus and within cerebellar eurydendroid cells, which in teleosts possibly resemble the deep cerebellar nucleus of higher vertebrates. Non-vestibulum related nuclei did not respond to hypergravity in a significant way. The effect of hyper-gravity found was much less distinct in adult animals as compared to the circumstances seen in larval fish (Anken et al., Adv. Space Res. 17, 1996), possibly due to a development correlated loss of neuronal plasticity.

Parkia pendula lectin as histochemistry marker for meningothelial tumour.

PubMed

Beltrão, E I C; Medeiros, P L; Rodrigues, O G; Figueredo-Silva, J; Valença, M M; Coelho, L C B B; Carvalho, L B

2003-01-01

Lectins have been intensively used in histochemical techniques for cell surface characterization. These proteins are involved in several biological processes and their use as histochemical markers have been evaluated since they can indicate differences in cell surfaces. Parkia pendula lectin (PpeL) was evaluated as histochemical marker for meningothelial meningioma biopsies. Tissue slices were incubated with PpeL conjugated to horseradish peroxidase (PpeL-HRP) and Concanavalin A-HRP (ConA-HPR) and the binding visualized with diaminobenzidine and hydrogen peroxide. The lectin-tissue binding was inhibited with D-glucose. PpeL showed to be a useful tool for the characterization of meningothelial tumour and clinico-pathological diagnosis.

Electron microprobe analysis and histochemical examination of the calcium distribution in human bone trabeculae: a methodological study using biopsy specimens from post-traumatic osteopenia

DOE Office of Scientific and Technical Information (OSTI.GOV)

Obrant, K.J.; Odselius, R.

1984-01-01

Energy dispersive X-ray microanalysis (EDX) (or electron microprobe analysis) of the relative intensity for calcium in different bone trabeculae from the tibia epiphysis, and in different parts of one and the same trabecula, was performed on 3 patients who had earlier had a fracture of the ipsilateral tibia-diaphysis. The variation in intensity was compared with the histochemical patterns obtained with both the Goldner and the von Kossa staining techniques for detecting calcium in tissues. Previously reported calcium distribution features, found to be typical for posttraumatic osteopenia, such as striated mineralization patterns in individual trabeculae and large differences in mineralization levelmore » between different trabeculae, could be verified both by means of the two histochemical procedures and from the electron microprobe analysis. A pronounced difference was observed, however, between the two histochemical staining techniques as regards their sensitivity to detect calcium. To judge from the values obtained from the EDX measurements, the sensitivity of the Goldner technique should be more than ten times higher than that of von Kossa. The EDX measurements gave more detailed information than either of the two histochemical techniques: great variations in the intensity of the calcium peak were found in trabeculae stained as unmineralized as well as mineralized.« less

Address substrates as promising targets for laser histochemical surgery as a nontraditional line in medicine

NASA Astrophysics Data System (ADS)

Piruzyan, L. A.; Mikhailovskiy, Ye. M.; Piruzyan, A. L.

1999-12-01

The priority concept of the laser histochemical surgery as a potentially novel line in medicine is presented. The histochemical stains, selectively coloring some targets (address substrates), that are cells or their biochemical ingredients, sensitize them to the laser irradiation. Such sensitization to laser irradiation by staining turns the colored targets into targets for the laser beam. The action of the irradiation onto its specific targets beats out of the cell its ingredients which participate in a pathology process. In particular, the beating of a stained ferment out of the general stage of biochemical processes characteristic for the pathology interrupts their currence. The laser beam, when beating out its stained targets without any damage of the unstained tissues, acts like a scalpel that cuts off affected tissues not brushing healthy ones. A scheme for testing stains as sensitizers of the `address substrates' to the laser irradiation is presented. As the criterion of the stain sensitization the fact was chosen of absence or weakness of pathomorphologic and biochemical signs of the disease in an experimental model of the pathology irradiated with laser after a stain use, while the pathology signs are present in a control sample. The basis is done for study of the histochemical stains as potential means for the laser histochemical surgery of disseminated sclerosis, mucopolysaccharidosis, hypercholesterolemia, myocardial infarction, cardiosclerosis, caries and parodontosis.

Apoplastic polyesters in Arabidopsis surface tissues--a typical suberin and a particular cutin.

PubMed

Franke, Rochus; Briesen, Isabel; Wojciechowski, Tobias; Faust, Andrea; Yephremov, Alexander; Nawrath, Christiane; Schreiber, Lukas

2005-11-01

Cutinized and suberized cell walls form physiological important plant-environment interfaces as they act as barriers limiting water and nutrient loss and protect from radiation and invasion by pathogens. Due to the lack of protocols for the isolation and analysis of cutin and suberin in Arabidopsis, the model plant for molecular biology, mutants and transgenic plants with a defined altered cutin or suberin composition are unavailable, causing that structure and function of these apoplastic barriers are still poorly understood. Transmission electron microscopy (TEM) revealed that Arabidopsis leaf cuticle thickness ranges from only 22 nm in leaf blades to 45 nm on petioles, causing the difficulty in cuticular membrane isolation. We report the use of polysaccharide hydrolases to isolate Arabidopsis cuticular membranes, suitable for depolymerization and subsequent compositional analysis. Although cutin characteristic omega-hydroxy acids (7%) and mid-chain hydroxylated fatty acids (8%) were detected, the discovery of alpha,omega-diacids (40%) and 2-hydroxy acids (14%) as major depolymerization products reveals a so far novel monomer composition in Arabidopsis cutin, but with chemical analogy to root suberin. Histochemical and TEM analysis revealed that suberin depositions were localized to the cell walls in the endodermis of primary roots and the periderm of mature roots of Arabidopsis. Enzyme digested and solvent extracted root cell walls when subjected to suberin depolymerization conditions released omega-hydroxy acids (43%) and alpha,omega-diacids (24%) as major components together with carboxylic acids (9%), alcohols (6%) and 2-hydroxyacids (0.1%). This similarity to suberin of other species indicates that Arabidopsis roots can serve as a model for suberized tissue in general.

THE LOCALIZATION OF MYOFIBRILLAR ATPASE ACTIVITY IN THE FLIGHT MUSCLES OF THE BLOWFLY, CALLIPHORA ERYTHROCEPHALA

PubMed Central

Tice, Lois W.; Smith, David S.

1965-01-01

The distribution of ATPase activity in the asynchronous flight muscles of Calliphora erythrocephala (Diptera) was studied at a fine structural level, using preparations of teased fibers, both unfixed and after brief fixation in hydroxyadipaldehyde, incubated in a medium for the histochemical demonstration of myosin or actomyosin ATPase. In relaxed fibrils, activity was found confined to the A bands and was absent from the H zones as well as from the Z and I band regions. At high magnification, deposits of final product, lead phosphate, appeared primarily related to the thick filaments, or to short lateral extensions from them. Evidence was gathered which indicated that this enzyme activity was that of a triphosphatase which did not act on dinucleoside or non-nucleoside substrates. PMID:4221034

Slow to fast alterations in skeletal muscle fibers caused by clenbuterol, a beta(2)-receptor agonist

NASA Technical Reports Server (NTRS)

Zeman, Richard J.; Ludemann, Robert; Easton, Thomas G.; Etlinger, Joseph D.

1988-01-01

The effects of a beta(2)-receptor agonist, clenbuterol, and a beta(2) antagonist, butoxamine, on the skeletal muscle fibers of rats were investigated. It was found that chronic treatment of rats with clenbuterol caused hypertrophy of histochemically identified fast-twitch, but not slow-twitch, fibers within the soleus, while in the extensor digitorum longus the mean areas of both fiber types were increased; in both muscles, the ratio of the number of fast-twitch to slow-twitch fibers was increased. In contrast, a treatment with butoxamine caused a reduction of the fast-twitch fiber size in both muscles, and the ratio of the fast-twitch to slow-twitch fibers was decreased.

Topochemistry of trace metals in nasal mucosa. Potentialities of some histochemical methods and energy dispersive X-ray microanalysis.

PubMed

Torjussen, W; Haug, F M; Olsen, A; Andersen, I

1978-01-01

Histochemical methods and energy dispersive X-ray micro-analysis (EDX-analysis) were evaluated in model experiments and on tissue sections for their usefulness in detecting traces of metals in biological tissue. The goal for this study was to establish a method for localization of nickel deposits in the nasal mucosa, where it has been found in concentrations between 1 and 40 microgram/g in nickel exposed individuals. The histochemical methods tested were staining with dimethylglyoxime, rubeanic acid and dithizone, the Turnbull and Prussian blue methods and TIMM'S sulphide silver procedure. In model experiments nickel-, cobalt-, copper-, zinc- and ironsalts were applied to thin-layer chromatography sheets (TLC-sheets) and stained by the histochemical methods. Spots containing 500 and 50 ng of these metals represented the smallest amounts that could consistently be detected in these experiments, except for the sulphide silver method which seemed a little more sensitive. With the latter method, moreover, zinc was detected in 40 micrometer thick cryostat sections of gelatine made up with 1 microgram/g of the metal. For nickel the corresponding figure was 10 to 50 microgram/g. On specimens of nasal mucosa from nickel-exposed workers, a faint colour was obtained in 40 micron thick cryostat sections from specimens that had been immersed in dithizone, but the colour was too weak for histological analysis. None of the other coloured chelating agents caused noticeable staining when applied to blocks or to cryostat sections. TIMM'S sulphide silver method caused strong staining of the basal layers of the surface epithelium and of fibroblast-like cells in the underlying connective tissue. This staining pattern is described in more detail in a separate report. Rat liver tissue was analyzed by atomic absorption before and after araldite embedding. Blocks of gelatine made up with nickel, copper, zinc and iron were embedded in epoxy resin and analyzed by atomic absorption. Large changes in the metal concentrations, usually an increase, were found after embedding. Ultrathin sections from this material were used to test the sensitivity of the EDX-equipment. Referring to the concentrations determined by atomic absorption in the embedded material, iron was detected at 1215 microgram/g and 362 microgram/g (gelatine standards) but not at 167 microgram/g (rat liver). Similar values could not be determined for nickel, copper or zinc, because of background radiation resulting from the presence of these metals in the instrument. We did not succeed in establishing a procedure for detecting nickel deposits in nasal mucosa with any of the methods which were tested. The most sensitive but least specific of the tested methods for visualizing heavy metals in the nasal mucosa, was TIMM'S sulphide silver procedure. The preparation of tissue for this method is discussed.

Histochemical investigation of Cochlospermum regium (Schrank) Pilg. leaves and chemical composition of its essential oil.

PubMed

Inácio, Marielle Cascaes; Paz, Tiago Antunes; Bertoni, Bianca Waléria; Vieira, Maria Aparecida Ribeiro; Marques, Márcia Ortiz Mayo; Pereira, Ana Maria Soares

2014-01-01

Essential oil from Cochlospermum regium (Schrank) Pilg. leaves (CR-EO) has been extracted by hydrodistillation; we analysed the CR-EO by gas chromatography coupled with mass spectrometry. We also conducted histochemical analysis on cross-sections of the central vein of young and adult leaves. A total of 32 compounds were qualitatively and quantitatively analysed, which represented 94.87% of the total CR-EO oil content. The CR-EO basically consisted of sesquiterpenes (96.87%); its main component was β-copaen-4-α-ol (18.73%), followed by viridiflorol (12.67%). The histochemical analyses identified the main classes of compounds present in both young and adult leaves.

[Orthotopic cartilage transplantation. Morphologic, angiography and histochemical studies of the vitality of free septum transplants].

PubMed

Gubisch, W; Donath, K

1999-11-01

Orthotopic cartilage transplantation is a technique frequently used in modern septal surgery. The prerequisite for a stable long-term result is viability of the transplanted cartilage. Therefore, we studied the healing process histologically, angiographically, and histochemically. We found a characteristic picture. Due to chondronal structure of the cartilage, the healing process varied in time and location. Reintegration took place by chondroneogenesis, commencing at the inner perichondrium. Reintegration depended directly on the distance of the cartilage cells to the surrounding vessels. Histochemically, we found an intact respiratory chain in the mitochondria and thus, we were able to demonstrate the preservation of viability in orthotopic transplanted cartilage.

Ontogenetical and histochemical studies on the translator apparatus in Calotropis procera R. Br. I. The retinaculum.

PubMed

Vijayaraghavan, M R; Cheema, K

1977-01-01

Translator apparatus consists of 2 parts--the corpusculum, which is attached to the margin of the stigmatic head between the anthers, and a pair of arms by which the corpusculum is attached to the pollinia of the adjacent anther halves. Retinaculum spans the lateral blade of the corpusculum to the acellular beak of the pollinium. Ontogenetically, it is the secretory product of the stigma along the secondary stigmatic groove. Histochemically, it is composed of lipid, lignin, cutin and a little amount of proteins. Retinaculum, like corpusculum, is stigmatic in origin, but differs histochemically from the latter being composed mainly of lipo-protein complex and lacking phenolic compounds.

An Investigation of the Memory Response of the Local Immune System to Shigella Antigens

DTIC Science & Technology

1989-03-03

CWM. Tugan NA: The histochemical demonstration of enterology 76:1035, 1979 protease by gelatin-silver film substance. J Histochem Cvtochem 23. Kotler DP...L. R. Armstrong , and J. E. Brown The Department of Pathology, University of Michigan Medical School, Ann Arbor, Michigan 48109, USA INTRODUCTION The

SOME HISTOCHEMICAL RESPONSES OF GUINEA PIG TISSUES TO COLD,

DTIC Science & Technology

Guinea pigs weighing approximately 300 gm were kept in a cold room, held at 6C, for two weeks. Various organs were then studied histochemically...Liver glycogen is rapidly used up in cold-exposed guinea pigs . The fate of liver lipids is unknown. Lipids in the cortex of the adrenals appear to

Age-related ultrastructural and monoamine oxidase changes in the rat optic nerve.

PubMed

Taurone, S; Ripandelli, G; Minni, A; Lattanzi, R; Miglietta, S; Pepe, N; Fumagalli, L; Micera, A; Pastore, F S; Artico, M

2016-01-01

The aim of this paper is to study the morphology and the distribution of the monoamine oxidase enzymatic system in the optic nerve of 4 month-old Wistar (young) and 28 month-old Wistar (old) rats. The optic nerve was harvested from 20 young and old rats. The segment of optic nerve was divided longitudinally into two pieces, each 0.1 mm in length. The first piece was used for transmission electron microscopy. The second piece was stained with histochemical reaction for monoamine oxidase. The agerelated changes in the optic nerve of rats include micro-anatomical details, ultrastructure and monoamine oxidase histochemical staining. A strong decrease of the thin nerve fibers and a swelling of the thick ones can be observed in optic nerve fibers of old rats. Increased monoamine oxidase histochemical staining of the optic nerve of aged rats is well demonstrated. The increase of meningeal shealth and the decrease of thin nerve fibers of the optic nerve in old rats are well documented. Morphological, ultrastructural and histochemical changes observed in optic nerve fibers of the old rats show a close relation with aging.

Is there still room for novelty, in histochemical papers?

PubMed

Pellicciari, Carlo

2016-12-16

Histochemistry continues to be widely applied in biomedical research, being nowadays mostly addressed to detect and locate single molecules or molecular complexes inside cells and tissues, and to relate structural organization and function at the high resolution of the more advanced microscopical techniques. In the attempt to see whether histochemical novelties may be found in the recent literature, the articles published in the European Journal of Histochemistry in the period 2014-2016 have been reviewed. In the majority of the published papers, standardized methods have been preferred by scientists to make their results reliably comparable with the data in the literature, but  many papers (approximately one fourth of the published articles) described novel histochemical methods and procedures.  It is worth noting that there is a growing interest for minimally-invasive in vivo techniques (magnetic resonance imaging, autofluorescence spectroscopy) which may parallel conventional  histochemical analyses to obtain information not only on the morphological features of living organs and tissues, but also on their functional, biophysical and molecular characteristics. Thanks to this unceasing methodological refinement, histochemistry will continue to provide innovative applications in the biomedical field.

Histochemical and microspectrophotometric analyses of early wound responses of resistant and susceptible Populus tremuloides inoculated with Entoleuca mammata (=Hypoxylon mammatum)

Treesearch

B. Bucciarelli; Michael E. Ostry; R. G. Fulcher; N. A. Anderson; C. P. Vance

1999-01-01

Stem tissue of resistant and susceptible genotypes of Poyulus tremuloides Michx, wounded or woundinoculated with Entoleuca mammata (Wahlenberg: Fr.) J.D. Rogers & Y.M. Ju was prepared for histochemical and microspectrophotometric analysis. Samples were collected over a 96-h period. Parenchyma cell walls associated with the...

Specific fibre composition and metabolism of the rectus abdominis muscle of bovine Charolais cattle

PubMed Central

2010-01-01

Background An important variability of contractile and metabolic properties between muscles has been highlighted. In the literature, the majority of studies on beef sensorial quality concerns M. longissimus thoracis. M. rectus abdominis (RA) is easy to sample without huge carcass depreciation and may appear as an alternative to M. longissimus thoracis for fast and routine physicochemical analysis. It was considered interesting to assess the muscle fibres of M. rectus abdominis in comparison with M. longissimus thoracis (LT) and M. triceps brachii (TB) on the basis of metabolic and contractile properties, area and myosin heavy chain isoforms (MyHC) proportions. Immuno-histochemical, histochemical, histological and enzymological techniques were used. This research concerned two populations of Charolais cattle: RA was compared to TB in a population of 19 steers while RA was compared to LT in a population of 153 heifers. Results RA muscle had higher mean fibre areas (3350 μm2 vs 2142 to 2639 μm2) than the two other muscles. In RA muscle, the slow-oxidative fibres were the largest (3957 μm2) and the fast-glycolytic the smallest (2868 μm2). The reverse was observed in TB muscle (1725 and 2436 μm2 respectively). In RA muscle, the distinction between fast-oxidative-glycolytic and fast-glycolytic fibres appeared difficult or impossible to establish, unlike in the other muscles. Consequently the classification based on ATPase and SDH activities seemed inappropriate, since the FOG fibres presented rather low SDH activity in this muscle in comparison to the other muscles of the carcass. RA muscle had a higher proportion of I fibres than TB and LT muscles, balanced by a lower proportion either of IIX fibres (in comparison to TB muscle) or of IIA fibres (in comparison to LT muscle). However, both oxidative and glycolytic enzyme activities were lower in RA than in TB muscle, although the LDH/ICDH ratio was higher in RA muscle (522 vs 340). Oxidative enzyme activities were higher in RA than in LT muscle, whereas glycolytic enzyme activity was lower. In RA muscle, contractile and metabolic properties appeared to be less well-correlated than in the two other muscles. Conclusions RA muscle has some particularities in comparison to the LT and TB muscles, especially concerning the unusual large cross-section surface of SO fibres and the very low oxidative activity of intermediate IIA fibres. PMID:20205735

Potential role of cyanidin 3-glucoside (C3G) in diabetic cardiomyopathy in diabetic rats: An in vivo approach.

PubMed

Li, Weizhen; Chen, Songwen; Zhou, Genqing; Li, Hongli; Zhong, Lan; Liu, Shaowen

2018-03-01

The present study aimed to evaluate the importance of cyanidin 3-glucoside (C3G) of diabetic cardiomyopathy in diabetic rats. The rats were induced with diabetic using streptozotocin and total triglyceride (TG) and total cholesterol (TC) were determined. The range of myocardial enzymes such as aspartate aminotransferase (AST), creatine kinase (CK) and lactate dehydrogenase (LD) were also estimated, further, the Immuno histochemical analysis and western blot investigation were determined for the actual activity of C3G. Results indicated that the marker enzymes such as CK, LD and AST were significantly ( P  < 0.05) increased in STZ administered rats (DM group), while the levels of these elevated marker enzymes of cardiac injury significantly ( P  < 0.05) declined in the DM + C3G group, as compared to the diabetic group of rats. Additionally, a decrease in the level of TNF-alpha and interleukin-6, was noticed in the C3G treated group as compared to diabetic group. Finally, blotting analysis clearly confirmed that theC3G treatment resulted to higher level response of Bcl-2 and lower level response of caspase-3 and BAX. In conclusion, C3G a natural antioxidant may prevent cardiovascular complications by ameliorating oxidative damage, inflammation, metabolic dysfunctions and apoptosis pathways in type 2 diabetes.

[Cylindroma of the breast. A histochemical and histogenetic study (author's transl)].

PubMed

van Bogaert, L J; Maldague, P; Pham-Maldague, H; Staquet, J P

1975-10-20

The histochemical study of mucopolysaccharide components of the ground substance of the cylindroma lends force to their epithelial origine. The myoepithelial cell, which is ectodermal in origin, plays an essential role in their secretion. Silver impregnation of normal and dysplastic breast illustrates the secretory function of myoepithelial cells and their possible implication in the histogenesis of cylindroma.

Improvement of skin condition by oral administration of collagen hydrolysates in chronologically aged mice.

PubMed

Wang, Zhenbin; Wang, Qing; Wang, Lin; Xu, Weidong; He, Yuanqing; Li, Yunliang; He, Song; Ma, Haile

2017-07-01

Collagen hydrolysates (CHs) have been demonstrated to have positive effects on skin photoaging by topical application or oral ingestion. However, there has been little research on their influence on skin chronological aging. In this study, 9-month-old female ICR mice were given normal AIN-93M diets containing CHs (2.5, 5 and 10% w/w) from Nile tilapia scale. After 6 months, the collagen content and antioxidant enzyme (superoxide dismutase and glutathione peroxidase) activities increased significantly (P < 0.05), while the survival rate, viscera indices and contents of moisture, fat and non-collagenous protein in skin did not change (P > 0.05). The color, luster and quantity of hair were obviously ameliorated. Moreover, the structure of epidermis and dermis, the density and distribution of collagen fibers and the ratio of type I to type III collagen were improved in a dose-dependent manner as shown by histochemical staining. Oral ingestion of CHs increased the collagen content and antioxidant enzyme activities and improved the appearance and structure of skin. These results suggest the potential of CHs as an anti-skin-aging ingredient in nutraceuticals or functional foods. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

Lignin metabolism involves Botrytis cinerea BcGs1- induced defense response in tomato.

PubMed

Yang, Chenyu; Liang, Yingbo; Qiu, Dewen; Zeng, Hongmei; Yuan, Jingjing; Yang, Xiufen

2018-06-04

BcGs1, a cell wall-degrading enzyme (CWDE), was originally derived from Botrytis cinerea. Our previous study revealed that BcGs1 could trigger defense responses and protect plants against various pathogens. We researched the defense response mechanism underlying this BcGs1 elicitation in tomato. We revealed that the two domains were required for BcGs1's full necrosis activity. According to analysis and quantitative real-time PCR of the up-regulated proteins and genes filtered by iTRAQ-based quantitative proteome approach, oxidative metabolism and phenylpropanoid metabolism were speculated to be involved in BcGs1-triggered defense response in tomato. Furthermore, experimental evidence showed that BcGs1 triggered reactive oxygen species (ROS) burst and increased the level of phenylalanine-ammonia lyase (PAL) and peroxidase (POD) enzyme activity, as well as lignin accumulation. Moreover, histochemical analysis revealed that infiltration of BcGs1 in tomato leaves exhibited cell wall thickening compared with untreated plants. The results suggested that BcGs1 activated the basal defense response included lignin metabolism contributed to BcGs1-induced resistance to Botrytis. cinerea infection in tomato.

Histochemical characteristics of a tonic smooth muscle.

PubMed

Gilloteaux, J; Stalmans-Falys, M

1979-09-01

It is suggested that ABRM, smooth muscle of Mytilus edulis L. and Mytilus galloprovincialis Lmk. (Mollusca Pelecypoda), is composed of one histochemical fibre type. The fibres are characterized by a low myofibrillar ATPase activity. Succinic and nicotinamide adenine dinucleotide oxidoreductase activities are distributed in a reverse pattern than that of the ATPase activity. Glycogen phosphorylase is richly represented in ABRM fibres and this detection is in opposition with the negative detection of alkaline phosphatase activity. These preliminary histochemical observations are similar to those found in some vertebrate smooth muscles. Mitochondrial glycerol-3-phosphate, 6-phosphogluconate, lactate and octopine dehydrogenases are not detected in muscle fibres whereas glio-interstitial tissues show weak but distinct reactivity. These last results especially characterize Mytilus catch fibres and are briefly discussed in relationship with previous physiological, biochemical and morphological observations.

[Mucous retention cysts of the minor salivary glands. A specific type of mucocele].

PubMed

Kakarantza-Angelopoulou, E; Triantaphyllou, A

1989-08-01

The mucous retention cyst of the minor salivary glands represent a specific type of oral mucocele which is lined by epithelium. It is caused probably from partial or complete obstruction of a duct. It affects older patients (over 40 years of age) most commonly women and it is located in different sites than the ordinary mucocele. In this paper we studied the histologic and histochemical features of four cases. The lining epithelium varied from cuboidal to columnar or flattened. Among the cells of the lining epithelium oncocytes were observed.

Copper treatment of the digestive gland of the slug, Arion ater L. 1. Bioassay conduction and histochemical analysis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Marigomez, J.A.; Angulo, E.; Moya, J.

1986-04-01

Effects of experimental treatment with copper on the digestive gland of the terrestrial slug, Arion ater, have been studied from a histochemical perspective. The purposes of histotoxicological studies must be: (a) to explain metal-metal interaction; and (b) to describe regulating mechanisms with regard to the use of molluscs in environmental analysis of metals.

Quantification of histochemical stains using whole slide imaging: development of a method and demonstration of its usefulness in laboratory quality control.

PubMed

Gray, Allan; Wright, Alex; Jackson, Pete; Hale, Mike; Treanor, Darren

2015-03-01

Histochemical staining of tissue is a fundamental technique in tissue diagnosis and research, but it suffers from significant variability. Efforts to address this include laboratory quality controls and quality assurance schemes, but these rely on subjective interpretation of stain quality, are laborious and have low reproducibility. We aimed (1) to develop a method for histochemical stain quantification using whole slide imaging and image analysis and (2) to demonstrate its usefulness in measuring staining variation. A method to quantify the individual stain components of histochemical stains on virtual slides was developed. It was evaluated for repeatability and reproducibility, then applied to control sections of an appendix to quantify H&E staining (H/E intensities and H:E ratio) between automated staining machines and to measure differences between six regional diagnostic laboratories. The method was validated with <0.5% variation in H:E ratio measurement when using the same scanner for a batch of slides (ie, it was repeatable) but was not highly reproducible between scanners or over time, where variation of 7% was found. Application of the method showed H:E ratios between three staining machines varied from 0.69 to 0.93, H:E ratio variation over time was observed. Interlaboratory comparison demonstrated differences in H:E ratio between regional laboratories from 0.57 to 0.89. A simple method using whole slide imaging can be used to quantify and compare histochemical staining. This method could be deployed in routine quality assurance and quality control. Work is needed on whole slide imaging devices to improve reproducibility. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.

Brenner tumor of the ovary: a correlative histologic, histochemical, immunohistochemical, and ultrastructural investigation.

PubMed

Santini, D; Gelli, M C; Mazzoleni, G; Ricci, M; Severi, B; Pasquinelli, G; Pelusi, G; Martinelli, G

1989-08-01

The histologic, histochemical, immunohistochemical, and ultrastructural features of Brenner tumor (BT) were studied. BT was compared with transitional bladder cells, and close similarities between the two tissues were identified. Abundant glycogen in all cellular layers, an alcianophilic/sialomucinic surface mucous coat, and argyrophilic cells characterized both BT and bladder epithelium. Immunohistochemically, chromogranin and neuron-specific enolase reactivity was observed in all cases examined. An additional relevant finding was the presence of serotonin-storing cells in both BT and urothelium. Moreover, carcinoembryonic antigen, epithelial membrane antigen, and keratin reaction were found in BT and urothelium, indicating an additional antigenic similarity. Additionally, malignant Brenner tumor was ultrastructurally found to share many common features with the bladder tissue. The distinct histochemical, ultrastructural, and antigenic pattern of BT, primarily of the transitional type, is emphasized.

Aluminium-induced excessive ROS causes cellular damage and metabolic shifts in black gram Vigna mungo (L.) Hepper.

PubMed

Chowra, Umakanta; Yanase, Emiko; Koyama, Hiroyuki; Panda, Sanjib Kumar

2017-01-01

Aluminium-induced oxidative damage caused by excessive ROS production was evaluated in black gram pulse crop. Black gram plants were treated with different aluminium (Al 3+ ) concentrations (10, 50 and 100 μM with pH 4.7) and further the effects of Al 3+ were characterised by means of root growth inhibition, histochemical assay, ROS content analysis, protein carbonylation quantification and 1 H-NMR analysis. The results showed that aluminium induces excessive ROS production which leads to cellular damage, root injury, stunt root growth and other metabolic shifts. In black gram, Al 3+ induces cellular damage at the earliest stage of stress which was characterised from histochemical analysis. From this study, it was observed that prolonged stress can activate certain aluminium detoxification defence mechanism. Probably excessive ROS triggers such defence mechanism in black gram. Al 3+ can induce excessive ROS initially in the root region then transported to other parts of the plant. As much as the Al 3+ concentration increases, the rate of cellular injury and ROS production also increases. But after 72 h of stress, plants showed a lowered ROS level and cellular damage which indicates the upregulation of defensive mechanisms. Metabolic shift analysis also showed that the black gram plant under stress has less metabolic content after 24 h of treatment, but gradually, it was increased after 72 h of treatment. It was assumed that ROS played the most important role as a signalling molecule for aluminium stress in black gram.

[NO and H2S brain systems of the Japanese shore crab Hemigrapsus sanguineus under conditions of anoxia].

PubMed

Kotsiuba, E P

2012-01-01

The topography and dynamics of the activity of the enzymes of the synthesis of nitric oxide (NO) and hydrogen sulfide (H2S) in the brain of the Japanese shore crab Hemigrapsus sanguineus after 1, 6, and 12 h ofanoxia was studied histochemically and immunocytochemically. Changes in the activity and number of NO- and CBS-immune-positive cells that take place due to anoxia and the intensity of which depends on the duration of the influence were revealed. The fact that the balance between the nitric oxide and hydrogen sulfide systems in the brain of the crabs H. sanguineus is preserved indicates the joint participation of those systems in the central regulation of adaptive mechanisms under the influence of anoxia and, apparently, plays an important role in the adaptation of these hydrobionts to oxygen deficit.

Pseudocyanotic pigmentation of the skin induced by amiodarone: a light and electron microscopic study.

PubMed Central

Delage, C.; Lagacé, R.; Huard, J.

1975-01-01

An unusual bluish discolouration of the nose was noticed in a woman 9 months after she had begun treatment with a coronary vasodilator, amiodarone hydrochloride. Cutaneous biopsies of the nose were obtained 6 and 9 months later for light and electron microscopic studies. In the dermis were histiocytes containing cytoplasmic yellow-brown granules with histochemical properties of melanin and lipofuscin. Ultrastructurally the granules appeared as lysosomal membrane-bound dense bodies similar to lipofuscin. Similar granules were observed at diascopy in both corneas. The pathogenesis is obscure. A storage disease involving the drug or its metabolites cannot be ruled out. Another possibility is that amiodarone accelerates the normal cellular autophagocytosis, resulting in increased production of lipofuscin, which then accumulates in lysosomes because of a deficiency in lipolytic enzymes. Images FIG. 1 FIG. 2 FIG. 3 FIG. 4 FIG. 5 FIG. 6 FIG. 7 FIG. 8 PMID:47784

Morphological and biochemical changes in soleus and extensor digitorum longus muscles of rats orbited in Spacelab 3

NASA Technical Reports Server (NTRS)

Riley, D. A.; Slocum, T.; Bain, J. L. W.; Sedlak, F. R.; Elis, S.; Satyanarayana, T.

1985-01-01

Muscle atrophy in rats exposed to hypogravity for seven days aboard Spacelab 3 is examined. Hindlimb muscles were harvested 12-16 days postflight, and prepared for enzyme studies and electron microscopy. Simple cell shrinkage was found, with a mean fiber area decrease of 35.8 percent for soleus and 24.9 percent for extensor digitorum longus (EDL) flight muscle fibers, as compared with control muscle fibers. EDL and soleus muscles showed increases in alkaline myofibrillar ATPase, alpha glycerophosphate dehydrogenase, and glycogen, and a decrease in NADH dehydrogenase staining. The 26 percent increase in calcium activated protease suggests that the focal degradation of myofibrils is the key process of myofibril breakdown. The presence in the flight soleus muscles of one percent necrotic fibers is unexplained. The observed shift towards histochemical fast-muscle type properties is consistent with previous findings.

Morphological and transcript changes in the biosynthesis of lignin in oil palm (Elaeis guineensis) during Ganoderma boninense infections in vitro.

PubMed

Goh, Kar Mun; Dickinson, Matthew; Supramaniam, Christina V

2018-03-01

Lignification of the plant cell wall could serve as the first line of defense against pathogen attack, but the molecular mechanisms of virulence and disease between oil palm and Ganoderma boninense are poorly understood. This study presents the biochemical, histochemical, enzymology and gene expression evidences of enhanced lignin biosynthesis in young oil palm as a response to G. boninense (GBLS strain). Comparative studies with control (T1), wounded (T2) and infected (T3) oil palm plantlets showed significant accumulation of total lignin content and monolignol derivatives (syringaldehyde and vanillin). These derivatives were deposited on the epidermal cell wall of infected plants. Moreover, substantial differences were detected in the activities of enzyme and relative expressions of genes encoding phenylalanine ammonia lyase (EC 4.3.1.24), cinnamate 4-hydroxylase (EC 1.14.13.11), caffeic acid O-methyltransferase (EC 2.1.1.68) and cinnamyl alcohol dehydrogenase (CAD, EC 1.1.1.195). These enzymes are key intermediates dedicated to the biosynthesis of lignin monomers, the guaicyl (G), syringyl (S) and ρ-hydroxyphenyl (H) subunits. Results confirmed an early, biphasic and transient positive induction of all gene intermediates, except for CAD enzyme activities. These differences were visualized by anatomical and metabolic changes in the profile of lignin in the oil palm plantlets such as low G lignin, indicating a potential mechanism for enhanced susceptibility toward G. boninense infection. © 2017 Scandinavian Plant Physiology Society.

Detection of endogenous alkaline phosphatase activity in intact cells by flow cytometry using the fluorogenic ELF-97 phosphatase substrate

NASA Technical Reports Server (NTRS)

Telford, W. G.; Cox, W. G.; Stiner, D.; Singer, V. L.; Doty, S. B.

1999-01-01

BACKGROUND: The alkaline phosphatase (AP) substrate 2-(5'-chloro-2'-phosphoryloxyphenyl)-6-chloro-4-(3H)-quinazolinone (ELF((R))-97 for enzyme-labeled fluorescence) has been found useful for the histochemical detection of endogenous AP activity and AP-tagged proteins and oligonucleotide probes. In this study, we evaluated its effectiveness at detecting endogenous AP activity by flow cytometry. METHODS: The ELF-97 phosphatase substrate was used to detect endogenous AP activity in UMR-106 rat osteosarcoma cells and primary cultures of chick chondrocytes. Cells were labeled with the ELF-97 reagent and analyzed by flow cytometry using an argon ultraviolet (UV) laser. For comparison purposes, cells were also assayed for AP using a Fast Red Violet LB azo dye assay previously described for use in detecting AP activity by flow cytometry. RESULTS: The ELF-97 phosphatase substrate effectively detected endogenous AP activity in UMR-106 cells, with over 95% of the resulting fluorescent signal resulting from AP-specific activity (as determined by levamisole inhibition of AP activity). In contrast, less than 70% of the fluorescent signal from the Fast Red Violet LB (FRV) assay was AP-dependent, reflecting the high intrinsic fluorescence of the unreacted components. The ELF-97 phosphatase assay was also able to detect very low AP activity in chick chondrocytes that was undetectable by the azo dye method. CONCLUSIONS: The ELF-97 phosphatase assay was able to detect endogenous AP activity in fixed mammalian and avian cells by flow cytometry with superior sensitivity to previously described assays. This work also shows the applicability of ELF-97 to flow cytometry, supplementing its previously demonstrated histochemical applications. Copyright 1999 Wiley-Liss, Inc.

Identifying motor and sensory myelinated axons in rabbit peripheral nerves by histochemical staining for carbonic anhydrase and cholinesterase activities

NASA Technical Reports Server (NTRS)

Riley, Danny A.; Sanger, James R.; Matloub, Hani S.; Yousif, N. John; Bain, James L. W.

1988-01-01

Carbonic anhydrase (CA) and cholinesterase (CE) histochemical staining of rabbit spinal nerve roots and dorsal root ganglia demonstrated that among the reactive myeliated axons, with minor exceptions, sensory axons were CA positive and CE negative whereas motor axons were CA negative and CE positive. The high specificity was achieved by adjusting reaction conditions to stain subpopulations of myelinated axons selectively while leaving 50 percent or so unstained. Fixation with glutaraldehyde appeared necessary for achieving selectivity. Following sciatic nerve transection, the reciprocal staining pattern persisted in damaged axons and their regenerating processes which formed neuromas within the proximal nerve stump. Within the neuromas, CA-stained sensory processes were elaborated earlier and in greater numbers than CE-stained regenerating motor processes. The present results indicate that histochemical axon typing can be exploited to reveal heterogeneous responses of motor and sensory axons to injury.

Evidence that the ZNT3 protein controls the total amount of elemental zinc in synaptic vesicles

USGS Publications Warehouse

Linkous, D.H.; Flinn, J.M.; Koh, J.Y.; Lanzirotti, A.; Bertsch, P.M.; Jones, B.F.; Giblin, L.J.; Frederickson, C.J.

2008-01-01

The ZNT3 protein decorates the presynaptic vesicles of central neurons harboring vesicular zinc, and deletion of this protein removes staining for zinc. However, it has been unclear whether only histochemically reactive zinc is lacking or if, indeed, total elemental zinc is missing from neurons lacking the Slc30a3 gene, which encodes the ZNT3 protein. The limitations of conventional histochemical procedures have contributed to this enigma. However, a novel technique, microprobe synchrotron X-ray fluorescence, reveals that the normal 2- to 3-fold elevation of zinc concentration normally present in the hippocampal mossy fibers is absent in Slc30a3 knockout (ZNT3) mice. Thus, the ZNT3 protein evidently controls not only the "stainability" but also the actual mass of zinc in mossy-fiber synaptic vesicles. This work thus confirms the metal-transporting role of the ZNT3 protein in the brain. ?? The Histochemical Society, Inc.

Cadmium-Induced Hydrogen Sulfide Synthesis Is Involved in Cadmium Tolerance in Medicago sativa by Reestablishment of Reduced (Homo)glutathione and Reactive Oxygen Species Homeostases

PubMed Central

Cui, Weiti; Chen, Huiping; Zhu, Kaikai; Jin, Qijiang; Xie, Yanjie; Cui, Jin; Xia, Yan; Zhang, Jing; Shen, Wenbiao

2014-01-01

Until now, physiological mechanisms and downstream targets responsible for the cadmium (Cd) tolerance mediated by endogenous hydrogen sulfide (H2S) have been elusive. To address this gap, a combination of pharmacological, histochemical, biochemical and molecular approaches was applied. The perturbation of reduced (homo)glutathione homeostasis and increased H2S production as well as the activation of two H2S-synthetic enzymes activities, including L-cysteine desulfhydrase (LCD) and D-cysteine desulfhydrase (DCD), in alfalfa seedling roots were early responses to the exposure of Cd. The application of H2S donor sodium hydrosulfide (NaHS), not only mimicked intracellular H2S production triggered by Cd, but also alleviated Cd toxicity in a H2S-dependent fashion. By contrast, the inhibition of H2S production caused by the application of its synthetic inhibitor blocked NaHS-induced Cd tolerance, and destroyed reduced (homo)glutathione and reactive oxygen species (ROS) homeostases. Above mentioned inhibitory responses were further rescued by exogenously applied glutathione (GSH). Meanwhile, NaHS responses were sensitive to a (homo)glutathione synthetic inhibitor, but reversed by the cotreatment with GSH. The possible involvement of cyclic AMP (cAMP) signaling in NaHS responses was also suggested. In summary, LCD/DCD-mediated H2S might be an important signaling molecule in the enhancement of Cd toxicity in alfalfa seedlings mainly by governing reduced (homo)glutathione and ROS homeostases. PMID:25275379

Comperative studies with Culex pipiens egg rafts. Immunogenetic, electrophoretic and enzymatic analysis of unfertilized, compatible and incompatible fertilized eggs.

PubMed

Schumann, W

1974-01-01

By applying immunologic, electrophoretic and enzymatic methods, extracts of different raft types of Culex pipiens were analysed. Rafts of the crosses Pa x Pa and Ha x Ha contained four common antigens, while unfertilized rafts of Pa and Ha (no antisera were prepared against them) and rafts of the crosses Og x Og, Og x Pa, and Pa x Og shared three common antigens with the remaining raft extracts. Disk-electrophoresis of raft extracts in acrylamide gel resulted in different electropherograms. Ten protein bands were common to all these raft types. The unfertilized rafts of Pa and Ha yielded three more protein bands, the crosses Pa x Ha and Ha x Pa one more, the crosses Og x Og and Pa x Og three more, and Og x Pa two more. Many enzymes were demonstrated in the raft extracts after they were separated in acrylamide gel and incubated with the corresponding substrate solutions. All the raft types possessed one enzyme type for glutaminate-, lactate-, glucose-6-phosphate-dehydrogenase and catalase. Malate-dehydrogenase and leucine aminopeptidase occurred in each raft type as two isoenzymes. Alkaline phosphatase was observed as a single enzyme, but was lacking in rafts of the crosses Pa X Pa and Ha X Ha. While rafts of the crosses Og x Og and Og x Pa possessed two acid phosphatases, three could be demonstrated for the remaining raft types. Up to eight esterases appeared; rafts of the crosses Og x Og and Og x Pa possessed seven such activities. The results obtained by the Ouchterlony test, disk-electrophoresis and the histochemical enzyme tests are discussed in context and checked according to the phenomenon of incompatibility.

Breaking conceptual locks in modelling root absorption of nutrients: reopening the thermodynamic viewpoint of ion transport across the root

PubMed Central

Le Deunff, Erwan; Malagoli, Philippe

2014-01-01

Background The top-down analysis of nitrate influx isotherms through the Enzyme-Substrate interpretation has not withstood recent molecular and histochemical analyses of nitrate transporters. Indeed, at least four families of nitrate transporters operating at both high and/or low external nitrate concentrations, and which are located in series and/or parallel in the different cellular layers of the mature root, are involved in nitrate uptake. Accordingly, the top-down analysis of the root catalytic structure for ion transport from the Enzyme-Substrate interpretation of nitrate influx isotherms is inadequate. Moreover, the use of the Enzyme-Substrate velocity equation as a single reference in agronomic models is not suitable in its formalism to account for variations in N uptake under fluctuating environmental conditions. Therefore, a conceptual paradigm shift is required to improve the mechanistic modelling of N uptake in agronomic models. Scope An alternative formalism, the Flow-Force theory, was proposed in the 1970s to describe ion isotherms based upon biophysical ‘flows and forces’ relationships of non-equilibrium thermodynamics. This interpretation describes, with macroscopic parameters, the patterns of N uptake provided by a biological system such as roots. In contrast to the Enzyme-Substrate interpretation, this approach does not claim to represent molecular characteristics. Here it is shown that it is possible to combine the Flow-Force formalism with polynomial responses of nitrate influx rate induced by climatic and in planta factors in relation to nitrate availability. Conclusions Application of the Flow-Force formalism allows nitrate uptake to be modelled in a more realistic manner, and allows scaling-up in time and space of the regulation of nitrate uptake across the plant growth cycle. PMID:25425406

Profiling and functional classification of esterases in olive (Olea europaea) pollen during germination.

PubMed

Rejón, Juan D; Zienkiewicz, Agnieszka; Rodríguez-García, María Isabel; Castro, Antonio J

2012-10-01

A pollen grain contains a number of esterases, many of which are released upon contact with the stigma surface. However, the identity and function of most of these esterases remain unknown. In this work, esterases from olive pollen during its germination were identifided and functionally characterized. The esterolytic capacity of olive (Olea europaea) pollen was examined using in vitro and in-gel enzymatic assays with different enzyme substrates. The functional analysis of pollen esterases was achieved by inhibition assays by using specific inhibitors. The cellular localization of esterase activities was performed using histochemical methods. Olive pollen showed high levels of non-specific esterase activity, which remained steady after hydration and germination. Up to 20 esterolytic bands were identified on polyacrylamide gels. All the inhibitors decreased pollen germinability, but only diisopropyl fluorophosphate (DIFP) hampered pollen tube growth. Non-specific esterase activity is localized on the surface of oil bodies (OBs) and small vesicles, in the pollen intine and in the callose layer of the pollen tube wall. Acetylcholinesterase (AChE) activity was mostly observed in the apertures, exine and pollen coat, and attached to the pollen tube wall surface and to small cytoplasmic vesicles. In this work, for the first time a systematic functional characterization of esterase enzymes in pollen from a plant species with wet stigma has been carried out. Olive pollen esterases belong to four different functional groups: carboxylesterases, acetylesterases, AChEs and lipases. The cellular localization of esterase activity indicates that the intine is a putative storage site for esterolytic enzymes in olive pollen. Based on inhibition assays and cellular localization of enzymatic activities, it can be concluded that these enzymes are likely to be involved in pollen germination, and pollen tube growth and penetration of the stigma.

Evidence for salicylic acid signalling and histological changes in the defence response of Eucalyptus grandis to Chrysoporthe austroafricana

PubMed Central

Zwart, Lizahn; Berger, Dave Kenneth; Moleleki, Lucy Novungayo; van der Merwe, Nicolaas A.; Myburg, Alexander A.; Naidoo, Sanushka

2017-01-01

Eucalyptus species are cultivated for forestry and are of economic importance. The fungal stem canker pathogen Chrysoporthe austroafricana causes disease of varying severity on E. grandis. The Eucalyptus grandis-Chrysoporthe austroafricana interaction has been established as a model system for studying Eucalyptus antifungal defence. Previous studies revealed that the phytohormone salicylic acid (SA) affects the levels of resistance in highly susceptible (ZG14) and moderately resistant (TAG5) clones. The aims of this study were to examine histochemical changes in response to wounding and inoculation as well as host responses at the protein level. The anatomy and histochemical changes induced by wounding and inoculation were similar between the clones, suggesting that anatomical differences do not underlie their different levels of resistance. Tyloses and gum-like substances were present after inoculation and wounding, but cell death occurred only after inoculation. Hyphae of C. austroafricana were observed inside dead and living cells, suggesting that the possibility of a hemibiotrophic interaction requires further investigation. Proteomics analysis revealed the possible involvement of proteins associated with cell death, SA signalling and systemic resistance. In combination with previous information, this study forms a basis for future functional characterisation of candidate genes involved in resistance of E. grandis to C. austroafricana. PMID:28349984

Sieve-element differentiation and phloem sap contamination.

PubMed

Knoblauch, Michael; Peters, Winfried S; Bell, Karen; Ross-Elliott, Timothy J; Oparka, Karl J

2018-06-01

Sieve elements (SEs) degrade selected organelles and cytoplasmic structures when they differentiate. According to classical investigations, only smooth ER, mitochondria, sieve element plastids, and, in most cases, P-proteins remain in mature SEs. More recent proteomics and immuno-histochemical studies, however, suggested that additional components including a protein-synthesizing machinery and a fully developed actin cytoskeleton operate in mature SEs. These interpretations are at odds with conventional imaging studies. Here we discuss potential causes for these discrepancies, concluding that differentiating SEs may play a role by 'contaminating' phloem exudates. Copyright © 2017 Elsevier Ltd. All rights reserved.

Assessment of pancreas cells

NASA Technical Reports Server (NTRS)

Vanoss, C. J.

1978-01-01

Pancreatic islets were obtained from guinea pig pancreas by the collagenase method and kept alive in tissue culture prior to further studies. Pancreas cell morphology was studied by standard histochemical techniques using light microscopy. Preparative vertical electrophoresis-levitation of dispersed fetal guinea pig pancreas cells was conducted in phosphate buffer containing a heavy water (D20) gradient which does not cause clumping of cells or alter the osmolarity of the buffers. The faster migrating fractions tended to be enriched in beta-cell content. Alpha and delta cells were found to some degree in most fractions. A histogram showing the cell count distribution is included.

Alpha-lipoic acid treatment of acetaminophen-induced rat liver damage.

PubMed

Mahmoud, Y I; Mahmoud, A A; Nassar, G

2015-01-01

Acetaminophen (paracetamol) is a well-tolerated analgesic and antipyretic drug when used at therapeutic doses. Overdoses, however, cause oxidative stress, which leads to acute liver failure. Alpha lipoic acid is an antioxidant that has proven effective for ameliorating many pathological conditions caused by oxidative stress. We evaluated the effect of alpha lipoic acid on the histological and histochemical alterations of liver caused by an acute overdose of acetaminophen in rats. Livers of acetaminophen-intoxicated rats were congested and showed centrilobular necrosis, vacuolar degeneration and inflammatory cell infiltration. Necrotic hepatocytes lost most of their carbohydrates, lipids and structural proteins. Liver sections from rats pre-treated with lipoic acid showed fewer pathological changes; the hepatocytes appeared moderately vacuolated with moderate staining of carbohydrates and proteins. Nevertheless, alpha lipoic acid at the dose we used did not protect the liver fully from acetaminophen-induced acute toxicity.

Direct sectioning of unembedded cartilage: a simple method for microscopical and histochemical studies on chondrocytes and extracellular matrix.

PubMed

Stockert, J C; Del Castillo, P

1990-01-01

On account of the rigidity and compact structure of the hyaline cartilage, unfixed or formaldehyde fixed samples of this tissue can be directly sectioned by using a conventional ultramicrotome and a glass knife. This simple method allows to obtain microscopical sections from unembedded cartilage blocks, which show a well preserved histological structure and are very suitable to carry out morphological and histochemical studies on chondrocytes and cartilaginous matrix.

Electron microscopical and histochemical studies on the transverse striated muscles of birds after prolonged hypokinesis

NASA Technical Reports Server (NTRS)

Belak, M.; Kocisova, J.; Marcanik, J.; Boda, K.; Skarda, R.

1981-01-01

Studies of the gastrocnemius muscle were carried out in 4 month old cockerels of the laying hybrid after hypokinesis lasting 15 and 30 days. It was found that restricted movement resulted in dystrophic changes of myotibrils, enlargement of the sarcoplasmic reticulum and oedem of interfibrillar spaces. Histochemical studies revealed focuses of increased activity of non-specific esterase, decreased activity of dehydrogenase of lactic acid and a positive reaction of acid phosphatase.

Histological and histochemical study of the protective role of rosemary extract against harmful effect of cell phone electromagnetic radiation on the parotid glands.

PubMed

Ghoneim, Fatma M; Arafat, Eetmad A

2016-06-01

Electromagnetic fields (EMFs) are a class of non-ionizing radiation (NIR) that is emitted from mobile phone. It may have hazardous effects on parotid glands. So, we aimed to investigate the histological and histochemical changes of the parotid glands of rats exposed to mobile phone and study the possible protective role of rosemary against its harmful effect. Forty adult male albino rats were used in this study. They were classified into 4 equal groups. Group I (control), group II (control receiving rosemary), group III (mobile phone exposed group) and group IV (mobile exposed, rosemary treated group). Parotid glands were dissected out for histological and histochemical study. Moreover, measurement of oxidative stress markers; malondialdehyde (MDA) and total antioxidant capacity (TAC) was done. The results of this study revealed that rosemary has protective effect through improving the histological and histochemical picture of the parotid gland in addition of its antioxidant effect. It could be concluded from the current study, that exposure of parotid gland of rat models to electromagnetic radiation of mobile phone resulted in structural changes at the level of light and electron microscopic examination which could be explained by oxidative stress effect of mobile phone. Rosemary could play a protective role against this harmful effect through its antioxidant activity. Copyright © 2016 Elsevier GmbH. All rights reserved.

Prophylactic action of garlic on the histological and histochemical patterns of hepatic and gastric tissues in rats injected with a snake venom.

PubMed

Rahmy, T R; Hemmaid, K Z

2001-05-01

The present study aimed to examine the prophylactic action of oral administration of two doses of garlic on the histological and histochemical patterns of the gastric and hepatic tissues in rats envenomed with cobra snake. The study included the following groups: Group I contained control rats orally administered distilled water for ten days. Group II included rats orally administered daily for ten days with the equivalent therapeutic dose of garlic to rat (18 mg/kg body weight). Group III included rats orally administered daily for ten days with double the equivalent therapeutic dose of garlic to rat (36 mg/kg body weight). Group IV contained rats intramuscularly (i.m.) injected with 1/2 LD50 of cobra venom (0.0125 microg venom/gm body weight) and dissected after 6 hr from injection. Groups V and VI contained rats daily administered with the previous two doses of garlic for ten days, respectively, followed by a single i.m. injection of the above dose of cobra venom after 24 hr from the last garlic application. Rats of these two groups were dissected after 6 hr from venom injection. Administration of the therapeutic dose of garlic induced slight cytoplasmic granulation in some hepatic cells. However, administration of double the therapeutic dose caused swelling, necrosis, and damage of the gastric glandular epithelia together with signs of erosion, exfoliation, and necrosis of the surface mucosal cells. It also induced swelling and coalescence of the hepatic cells, loss of the normal arrangement of the hepatic cords, and hypertrophy of Kupffer cells. Injection with cobra venom caused loss of the normal characteristic appearance of the gastric glands and the epithelial lining cells of the gastric folds and the appearance of numerous inflammatory cells in the lamina properia. It also induced the occurrence of highly swollen hepatic cells, hepatic cellular necrosis and damage, as well as activated Kupffer cells. Nevertheless, pretreatment with the therapeutic dose of garlic for ten days induced a prophylactic activity against the pathogenic effects of the venom in both tissues, which appeared more or less normal except for very minor abnormalities. However, application of double the therapeutic dose of garlic for the same duration did not induce any prophylactic activity. Histochemically, slight alterations were noticed in the polysaccharide, protein, and nucleic acid contents of the gastric mucosa and the hepatic tissues due to administration of the therapeutic doses of garlic. However, severe depletions of these components were recorded in both tissues due to administration of double the therapeutic doses of garlic or injection of cobravenom or the application of both of them together. On the contrary, minor changes were noticed in the histochemical patterns of both tissues in rats pretreated with the therapeutic doses of garlic prior to venom application. It could be concluded that oral administration of the therapeutic dose of garlic for ten days has no serious side effects on gastric and hepatic tissues and could be used as a prophylactic tool against cobra snake envenomation.

Ameliorating effect of berbamine on hepatic key enzymes of carbohydrate metabolism in high-fat diet and streptozotocin induced type 2 diabetic rats.

PubMed

Sankaranarayanan, Chandrasekaran; Nishanthi, Ramajayam; Pugalendi, Pachaiappan

2018-07-01

Aberrations in the activities of key enzymes of carbohydrate metabolism is well documented in diabetes mellitus. Previous studies have shown that active ingredients in the extracts of Berberis aristata exhibits diverse pharmacological activities in animal models. The present study was undertaken to investigate whether berbamine (BBM), an alkaloid from the roots of Berberis aristata can ameliorate the altered activities of carbohydrate metabolic enzymes in high fat diet (HFD)/streptozotocin (STZ) induced diabetic rats. Supplementation of HFD for 4 weeks followed by intraperitonial administration of single low dose of STZ (40 mg/kg b.w.) to Sprague Dawley rats resulted in significant hyperglycemia with a decline in plasma insulin levels. The rats also exhibited decreased hemoglobin with an increase in glycated hemoglobin levels. The activities of hexokinase, glucose-6-phosphate dehydrogenase were decreased whereas increases in the activities of glucose-6-phosphatase and fructose-1,6-bisphosphatase were observed in the hepatic tissues of diabetic control rats. Glycogen content in the hepatic and skeletal muscle tissues were found to be decreased in diabetic rats. Oral administration of BBM for 56 days, dose dependently (50, 100, 200 mg/kg b.w.) improved insulin secretion in diabetic treated rats. Immunohistochemical studies on pancreas revealed a strong immunoreactivity to insulin in BBM treated rats. At the effective dose of 100 mg/kg b.w., BBM restored the altered activities of carbohydrate metabolic enzymes and also improved glycogen content in insulin dependent tissues. From the biochemical and histochemical data obtained in this study we conclude that BBM ameliorated the activities of metabolic enzymes and maintained glucose homeostasis in HFD/STZ induced diabetic rats and it can be used as a potential phytomedicine for the management of diabetes mellitus. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

Muscles within muscles: a tensiomyographic and histochemical analysis of the normal human vastus medialis longus and vastus medialis obliquus muscles

PubMed Central

Travnik, Ludvik; Djordjevič, Srdjan; Rozman, Sergej; Hribernik, Marija; Dahmane, Raja

2013-01-01

The aim of this study was to show the connection between structure (anatomical and histochemical) and function (muscle contraction properties) of vastus medialis obliquus (VMO) and vastus medialis longus (VML). The non-invasive tensiomyography (TMG) method was used to determine the contractile properties (contraction time; Tc) of VML and VMO muscle, as a reflection of the ratio between the slow and fast fibers in two groups of nine young men. VML and VMO significantly (P < 0.01) differ in the proportion of type 1 (59.6: 44%) and type 2b (6.3: 15%) fibers. The VML muscle is almost entirely composed of type 1 and type 2a fibers. In many samples of this muscle no type 2b fibers were found. The proportion of slow-twitch type 1 fibers is nearly twice as high as the proportion of fast-twitch type 2a fibers. These observations indicate that VML is a slower and more fatigue-resistant muscle than VMO muscle. These characteristics correspond to the different functions of the VML, which is an extensor of the knee, and to the VMO, which maintains the stable position of the patella in the femoral groove. Our results obtained by TMG provided additional evidence that muscle fibers within the segments of VM muscle were not homogenous with regard to their contractile properties, thereby confirming the histochemical results. Tc can be attributed to the higher percentage of slow-twitch fibers – type 1. The statistically shorter Tc (P ≤ 0.001) of VMO (22.8 ± 4.0 ms) compared with VML (26.7 ± 4.0 ms) in our study is consistent with previously found differences in histochemical, morphological and electrophysiological data. In conclusion, the results of this study provide evidence that the VML and VMO muscles are not only anatomically and histochemically different muscles, but also functionally different biological structures. PMID:23586984

[Histochemical detection of glycoproteins and glycosaminoglycans in the respiratory mucosa of albino rats during estrous cycle, pregnancy and puerperium].

PubMed

Pontes, P A; Simões, M J; Merzel, J

1989-11-01

In this work we attempted to detect, with histochemical methods, the possible modifications in the mucus of the respiratory mucosa of albino female rats during estral cycle, pregnancy and puerperium. Based on its results, it was possible to conclude that: a--There were no modifications in the nature of the epithelial and supraepithelial mucus during the studied periods: b--The Alcian Blue staining from lamina propria is absent during pregnancy and present during puerperium.

Synthesis of the signal molecule acetylcholine during the developmental cycle of Paramecium primaurelia (Protista, Ciliophora) and its possible function in conjugation.

PubMed

Delmonte Corrado, M U; Politi, H; Ognibene, M; Angelini, C; Trielli, F; Ballarini, P; Falugi, C

2001-06-01

We recently discovered, in mating-competent Paramecium primaurelia, the presence of functionally related molecules of the cholinergic system: the neurotransmitter acetylcholine (ACh), both its nicotinic and muscarinic receptors and its lytic enzyme acetylcholinesterase (AChE). Our results on the inhibition of mating-cell pairing in vivo in mating-competent cells treated with cholinomimetic drugs support the hypothesis that the cholinergic system plays a role in cell-to-cell adhesion. To investigate the possible function of the signal molecule ACh in conjugation in P. primaurelia, we attempted to detect the intracellular sites of ACh synthesis by localizing the ACh biosynthetic enzyme choline acetyltransferase (ChAT). Using immunocytochemical and histochemical methods, we have demonstrated the presence and activity of ChAT principally on the surface membrane of mating-competent cells and of mature but non-mating-competent cells. No evidence for ChAT activity was found in immature cells. Immunoblot analysis revealed the presence of immunoreactive bands, ranging in molecular mass from 42 to 133 kDa, as reported for ChAT isolated from higher organisms. In vivo experiments showed that inhibition of ChAT activity by Congo Red, known to be a potent competitive inhibitor of acetyl coenzyme A, did not affect mating-cell pairing. Conversely, inhibition of AChE with BW 284c51 or eserine, which block enzyme activity by reacting with a specific serine within the catalytic centre, significantly inhibited mating-cell pairing. Our results suggest that ACh has a negative modulating effect on conjugation in P. primaurelia.

Effect of Hypergravity on Carbonanhydrase Reactivity in inner Ear Ioncytes of developing Cichlid Fish

NASA Astrophysics Data System (ADS)

Beier, M.; Anken, R.; Rahmann, H.

It has been shown earlier that hypergravity slows down inner ear otolith growth in developing fish. Otolith growth in terms of mineralisation mainly depends on the enzyme carboanhydrase (CAH), which is responsible for the provision of the pH- value necessary for calcium carbonate deposition and thus also is presumed to play a prominent role in Ménière's disease (a sensory - motor disorder inducing vertigo and kinetosis). Larval siblings of cichlid fish (Oreochromis mossambicus) were subjected to hypergravity (3g; 6 hours) during development and separated into normally and kinetotically swimming individuals following the transfer to 1g (i.e., stopping the centrifuge; kinetotically behaving fish performed spinning movements). Subsequently, CAH was histochemically demonstrated in inner ear ionocytes (cells involved in the endolymphatic ion exchange) and enzyme reactivity was determined densitometrically. The results showed that CAH-reactivity was significantly increased in normally behaving hyper-g specimens as compared to controls kept at 1g, whereas no difference in enzyme reactivity was evident between the controls and kinetotically behaving fish. On the background of earlier studies, according to which (1) hypergravity induces a decrease of otolith growth and (2) the otolithic calcium incorporation (visualized using the calcium -tracer alizarin complexone) of kinetotically swimming hyper - g fish was lower as compared to normally behaving hyper - g animals, the present study strongly supports the concept that an increase in CAH-reactivity may result in a decrease of otolithic calcium deposition. The mechanism regulating CAH-activity hitherto remains to be determined. Acknowledgement: This work was financially supported by the German Aerospace Center (DLR) (FKZ: 50 WB 9997).

[Effects of androgens and gestagens on uterine myoma].

PubMed

Pashkova, V S; Erokhin, Iu A

1991-01-01

One hundred and sixty resected intramural myomas have been examined in 78 women with an intact menstrual cycle: 61 nodes from 31 patients of 37 to 57 years without a history of hormonal treatment, 52 nodes from 26 patients of 39 to 57 years with a history of androgen treatment and 47 nodes from 21 women of 30 to 50 years with a history of gestagen treatment. The histologic, histochemical, morphometric and biochemical studies showed that androgen and gestagen treatment stabilized and diminished the myomas, promoted secondary changes in the nodes and caused a statistically significant activation of sclerotic processes. Therapeutic effects of androgens were superior to those of gestagens.

Carbon tetrachloride-induced hepatotoxicity and its amelioration by Agaricus blazei Murrill extract in a mouse model.

PubMed

Chang, Jin-Biou; Wu, Ming-Fang; Yang, Yi-Yuan; Leu, Sy-Jye; Chen, Yung-Liang; Yu, Chun-Shu; Yu, Chieh-Chih; Chang, Shu-Jen; Lu, Hsu-Feng; Chung, Jing-Gung

2011-01-01

This study was conducted to evaluate the hepatoprotective effect of Agaricus blazei Murrill extract (ABM) against experimentally induced carbon tetrachloride (CCl(4)) toxicity in male BALB/c mice. The experiments included a normal group (no induction by CCl(4)), CCl(4-)induction group (with hepatotoxicity by CCl(4) and without treatment) and experimental groups with low dose (200 mg) or high dose (2,000 mg) of ABM extract (per kilogram mouse weight). All groups other than the normal group were treated with intraperitoneal injections of CCl(4) twice a week. Mice were tube-fed with experimental ABM extracts or double-distilled water, accordingly, on the remaining four days each week. The whole experimental protocol lasted 8 weeks; blood and liver samples were collected for biochemical and tissue histochemical analysis. Only administration of a high dose of ABM to treatment groups resulted in a significant abrogation of CCL(4)-induced increase of serum aspartate aminotransferase (AST) and alanine transaminase (ALT). Post-treatment with ABM also did not significantly reverse the alterations of glutathione peroxidase (GSHPx) and catalase. Both high- and low-dose ABM treatment reduced hepatic necrosis and fibrosis caused by CCl(4) in comparison with the CCl(4) control group in the histochemical analyses. Our results suggest that the ABM extract affects the levels of ALT and AST in mice.

Tissue Fixation and Processing for the Histological Identification of Lipids.

PubMed

Carriel, Víctor; Campos, Fernando; Aneiros-Fernández, José; Kiernan, John A

2017-01-01

Lipids are a heterogeneous group of substances characterized by their solubility in organic solvents and insolubility in water. Lipids can be found as normal components of different tissues and organs, and they can be affected by several pathological conditions. The histochemical identification of lipids plays an important role in histopathological diagnosis and research, but successful staining depends on adequate fixation and processing of the tissue. Here we describe methods to fix and process tissue samples for the histochemical identification of lipids in frozen or paraffin-embedded tissues.

Computer Image Analysis of Histochemically-Labeled Acetylcholinesterase.

DTIC Science & Technology

1984-11-30

image analysis on conjunction with histochemical techniques to describe the distribution of acetylcholinesterase (AChE) activity in nervous and muscular tissue in rats treated with organophosphates (OPs). The objective of the first year of work on this remaining 2 years. We began by adopting a version of the AChE staining method as modified by Hanker, which consistent with the optical properties of our video system. We wrote computer programs for provide a numeric quantity which represents the degree of staining in a tissue section. The staining was calibrated by

Corn fiber: structure, composition, and response to enzymes for fermentable sugars and coproducts.

PubMed

Akin, Danny E; Rigsby, Luanne L

2008-01-01

Corn (Zea mays L.) fiber, which is the seed coat and residual endosperm left after grain processing, is a low-value residue that contains carbohydrates and aromatic compounds that could provide value-added coproducts. Treatment of corn fiber with NaOH and assessment by gas chromatography indicated a prevalence of ferulic acid, with about 90% ester-linked in the cell walls. p-coumaric acid was much lower at about 10% of the amount of ferulic acid. Histochemical reactions employing acid phloroglucinol and diazotized sulfanilic acid indicated the presence of phenolic acids in cell walls of the pericarp and aleurone layer. Various protocols were tested using milled corn fiber and pretreatment with commercial ferulic acid esterases before cellulase treatment, and dry weight loss and sugars and phenolic acids released into the filtrate were evaluated. Ferulic acid esterases effectively degraded corn fiber and released substantial amounts of ferulic acid and sugars (e.g., glucose and xylose) in the incubation medium. Light microscopy showed that ferulic acid esterase substantially disrupted the aleurone layer but caused little visible damage to the lignified pericarp cell walls. Amounts of compounds released varied with protocols, and one study with various milling methods showed that esterase pretreatment followed by cellulase released about 2.8 to 4.4 and 1.5 to 2.9 times more ferulic acid and glucose, respectively, than cellulase alone. The highest levels for one lot of corn fiber with esterase pretreatment followed by cellulase were 3.9 and 218 mg/g of ferulic acid and glucose, respectively.

Methane alleviates copper-induced seed germination inhibition and oxidative stress in Medicago sativa.

PubMed

Samma, Muhammad Kaleem; Zhou, Heng; Cui, Weiti; Zhu, Kaikai; Zhang, Jing; Shen, Wenbiao

2017-02-01

Recent results discovered the protective roles of methane (CH 4 ) against oxidative stress in animals. However, the possible physiological roles of CH 4 in plants are still unknown. By using physiological, histochemical and molecular approaches, the beneficial role of CH 4 in germinating alfalfa seeds upon copper (Cu) stress was evaluated. Endogenous production of CH 4 was significantly increased in Cu-stressed alfalfa seeds, which was mimicked by 0.39 mM CH 4 . The pretreatment with CH 4 significantly alleviated the inhibition of seed germination and seedling growth induced by Cu stress. Cu accumulation was obviously blocked as well. Meanwhile, α/β amylase activities and sugar contents were increased, all of which were consistent with the alleviation of seed germination inhibition triggered by CH 4 . The Cu-triggered oxidative stress was also mitigated, which was confirmed by the decrease of lipid peroxidation and reduction of Cu-induced loss of plasma membrane integrity in CH 4 -pretreated alfalfa seedlings. The results of antioxidant enzymes, including ascorbate peroxidase (APX), superoxide dismutase (SOD), catalase (CAT), and guaiacol peroxidase (POD) total or isozymatic activities, and corresponding transcripts (APX1/2, Cu/Zn SOD and Mn-SOD), indicated that CH 4 reestablished cellular redox homeostasis. Further, Cu-induced proline accumulation was partly impaired by CH 4 , which was supported by the alternation of proline metabolism. Together, these results indicated that CH 4 performs an advantageous effect on the alleviation of seed germination inhibition caused by Cu stress, and reestablishment of redox homeostasis mainly via increasing antioxidant defence.

Indian Ginseng (Withania somnifera) supplementation ameliorates oxidative stress and mitochondrial dysfunctions in experimental model of stroke.

PubMed

Sood, Abhilasha; Mehrotra, Arpit; Dhawan, Devinder K; Sandhir, Rajat

2018-04-18

Stroke is an increasingly prevalent clinical condition and second leading cause of death globally. The present study evaluated the therapeutic potential of Indian Ginseng, also known as Withania somnifera (WS), supplementation on middle cerebral artery occlusion (MCAO) induced mitochondrial dysfunctions in experimental model of ischemic stroke. Stroke was induced in animals by occluding the middle cerebral artery, followed by reperfusion injury. Ischemia reperfusion injury resulted in increased oxidative stress indicated by increased reactive oxygen species and protein carbonyl levels; compromised antioxidant system; in terms of reduced superoxide dismutase and catalase activity, along with reduction in GSH levels and the redox ratio, impaired mitochondrial functions and enhanced expression of apoptosis markers. Ischemia reperfusion injury induced mitochondrial dysfunctions in terms of (i) reduced activity of the mitochondrial respiratory chain enzymes, (ii) reduced histochemical staining of complex-II and IV, (iii) reduced in-gel activity of mitochondrial complex-I to V, (iv) mitochondrial structural changes in terms of increased mitochondrial swelling, reduced mitochondrial membrane potential and ultrastructural changes. Additionally, an increase in the activity of caspase-3 and caspase-9 was also observed, along with altered expression of apoptotic proteins Bcl-2 and Bax in MCAO animals. MCAO animals also showed significant impairment in cognitive functions assessed using Y maze test. WS pre-supplementation, on the other hand ameliorated MCAO induced oxidative stress, mitochondrial dysfunctions, apoptosis and cognitive impairments. The results show protective effect of WS pre-supplementation in ischemic stroke and are suggestive of its potential application in stroke management.

AMP deaminase histochemical activity and immunofluorescent isozyme localization in rat skeletal muscle

NASA Technical Reports Server (NTRS)

Thompson, J. L.; Sabina, R. L.; Ogasawara, N.; Riley, D. A.

1992-01-01

The cellular distribution of AMP deaminase (AMPda) isozymes was documented for rat soleus and plantaris muscles, utilizing immunofluorescence microscopy and immunoprecipitation methods. AMPda is a ubiquitous enzyme existing as three distinct isozymes, A, B and C, which were initially purified from skeletal muscle, liver (and kidney), and heart, respectively. AMPda-A is primarily concentrated subsarcolemmally and intermyofibrillarly within muscle cells, while isozymes B and C are concentrated within non-myofiber elements of muscle tissue. AMPda-B is principally associated with connective tissues surrounding neural elements and the muscle spindle capsule, and AMPda-C is predominantly associated with circulatory elements, such as arterial and venous walls, capillary endothelium, and red blood cells. These specific localizations, combined with documented differences in kinetic properties, suggest multiple functional roles for the AMPda isozymes or temporal segregation of similar AMPda functions. Linkage of the AMPda substrate with adenosine production pathways at the AMP level and the localization of isozyme-C in vascular tissue suggest a regulatory role in the microcirculation.

Prostatic acid phosphatase is an ectonucleotidase and suppresses pain by generating adenosine

PubMed Central

Zylka, Mark J.; Sowa, Nathaniel A.; Taylor-Blake, Bonnie; Twomey, Margaret A.; Herrala, Annakaisa; Voikar, Vootele; Vihko, Pirkko

2008-01-01

SUMMARY Thiamine monophosphatase (TMPase, also known as Fluoride-Resistant Acid Phosphatase) is a classic histochemical marker of small-diameter dorsal root ganglia neurons. The molecular identity of TMPase is currently unknown. We found that TMPase is identical to the transmembrane isoform of Prostatic Acid Phosphatase (PAP), an enzyme with unknown molecular and physiological functions. We then found that PAP knockout mice have normal acute pain sensitivity but enhanced sensitivity in chronic inflammatory and neuropathic pain models. In gain-of-function studies, intraspinal injection of PAP protein has potent anti-nociceptive, anti-hyperalgesic and anti-allodynic effects that last longer than the opioid analgesic morphine. PAP suppresses pain by functioning as an ecto-5’-nucleotidase. Specifically, PAP dephosphorylates extracellular adenosine monophosphate (AMP) to adenosine and activates A1-adenosine receptors in dorsal spinal cord. Our studies reveal molecular and physiological functions for PAP in purine nucleotide metabolism and nociception and suggest a novel use for PAP in the treatment of chronic pain. PMID:18940592

Nitric Oxide Signaling in Hypergravity-Induced Neuronal Plasticity

NASA Technical Reports Server (NTRS)

Holstein, Gay R.

2003-01-01

The goal of this research project was to identify the neurons and circuits in the vestibular nuclei and nucleus prepositus hypoglossi that utilize nitric oxide (NO) for intercellular signaling during gravity-induced plasticity. This objective was pursued using histochemical and immunocytochemical approaches to localize NO-producing neurons and characterize the fine morphology of the cells in ground-based studies of normal rats, rats adapted to hypergravity, and rats adapted to hypergravity and then re-adapted to the 1G environment. NO-producing neurons were identified and studied using four methodologies: i) immunocytochemistry employing polyclonal antibodies directed against neuronal nitric oxide synthase (nNOS), to provide an indication of the capacity of a cell for NO production; ii) immunocytochemistry employing a monoclonal antibody directed against L-citrulline, to provide an indirect index of the enzyme's activity; iii) histochemistry based on the NADPH-diaphorase reaction, for fuI1 cytological visualization of neurons; and iv) double immunofluorescence to co-localize nNOS and L-citrulline in individual vestibular nuclei (VN) and neurons.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Vanderlaan, M.; Fong, S.; King, E.B.

To improve identification of preneoplastic bladder cancer cells, we have studied two enzyme histochemical changes in bladder tumors induced in male Fisher 344 rats by the carcinogen N-butyl-N-(4-hydroxybutyl)-nitrosamine. In early areas of focal nodular hyperplasia there was a dramatic increase in staining for NADH:menadione oxidoreductase (diaphorase) activity. In nonfocal areas as well, there were many individual cells with intense staining, while the controls were of uniform moderate staining. Large papillomas and carcinomas often showed heterogeneous staining, ..gamma..-Glutamyltranspeptidase (GGT) was absent from normal urothelium and from all tumors except the most advanced carcinomas and large papillomas. In old, carcinogen-exposed animals, GGTmore » activity was seen in the luminal surface of tumors and in the interlesion urothelium. In newborn rats and in rats with regenerative hyperplasia following wounding of the urothelium, the diaphorase staining was less than that in the untreated adult. Our findings suggest that increased diaphorase activity may serve to identify early islands of carcinogen-induced, enzymatically altered bladder cells, while GGT will not.« less

Apple (Malus x domestica).

PubMed

Dandekar, Abhaya M; Teo, Gianni; Uratsu, Sandra L; Tricoli, David

2006-01-01

Apple (Malus x domestica) is one of the most consumed fruit crops in the world. The major production areas are the temperate regions, however, because of its excellent storage capacity it is transported to distant markets covering the four corners of the earth. Transformation is a key to sustaining this demand - permitting the potential enhancement of existing cultivars as well as to investigate the development of new cultivars resistant to pest, disease, and storage problems that occur in the major production areas. In this paper we describe an efficient Agrobacterium tumefaciens-mediated transformation protocol that utilizes leaf tissues from in vitro grown plants. Shoot regeneration is selected with kanamycin using the selectable kanamycin phosphotransferase (APH(3)II) gene and the resulting transformants confirmed using the scorable uidA gene encoding the bacterial beta-glucuronidase (GUS) enzyme via histochemical staining. Transformed shoots are propagated, rooted to create transgenic plants that are then introduced into soil, acclimatized and transferred to the greenhouse from where they are taken out into the orchard for field-testing.

Exogenous Nitric Oxide (NO) Interferes with Lead (Pb)-Induced Toxicity by Detoxifying Reactive Oxygen Species in Hydroponically Grown Wheat (Triticum aestivum) Roots

PubMed Central

Kaur, Gurpreet; Singh, Harminder Pal; Batish, Daizy R.; Mahajan, Priyanka; Kohli, Ravinder Kumar; Rishi, Valbha

2015-01-01

Nitric Oxide (NO) is a bioactive signaling molecule that mediates a variety of biotic and abiotic stresses. The present study investigated the role of NO (as SNP [sodium nitroprusside]) in ameliorating lead (Pb)-toxicity in Triticum aestivum (wheat) roots. Pb (50 and 250 μM) alone and in combination with SNP (100 μM) was given to hydroponically grown wheat roots for a period of 0–8 h. NO supplementation reduced the accumulation of oxidative stress markers (malondialdehyde, conjugated dienes, hydroxyl ions and superoxide anion) and decreased the antioxidant enzyme activity in wheat roots particularly up to 6 h, thereby suggesting its role as an antioxidant. NO ameliorated Pb-induced membrane damage in wheat roots as evidenced by decreased ion-leakage and in situ histochemical localization. Pb-exposure significantly decreased in vivo NO level. The study concludes that exogenous NO partially ameliorates Pb-toxicity, but could not restore the plant growth on prolonged Pb-exposure. PMID:26402793

Width of thermal damage after using the YAG contact laser for cutting biological tissue: animal experimental investigation.

PubMed

Mecke, H; Schünke, M; Schnaidt, S; Freys, I; Semm, K

1991-01-01

At the University Women's Clinic in Kiel, the YAG contact laser has been used as a cutting instrument in pelviscopic operations since 1987. When the laser cuts, it produces only a scant amount of mechanical trauma. The determining factor is the amount of thermal damage produced along the wound margins and in direct neighboring tissue. The extent of the tissue change seen in the uterus and liver parenchyma of rats and the striated muscle of rabbits after application of the YAG contact laser was demonstrated using various staining techniques and stains. Liver parenchyma proved to be the most sensitive to thermal damage. In the uterine horn, enzyme-histochemical ATPase and alkaline phosphatase demonstrations showed a significantly wider zone of thermal damage after laser incision than did hematoxylin-eosin and Goldner staining techniques. A good understanding of the extent of thermal damage is essential for atraumatic pelviscopic operations using the YAG contact laser and also for the preventing of complications.

Histochemical Characterization of Oocytes in the Pink Cuskeel (Genypterus blacodes).

PubMed

Cohen, Stefanía; Petcoff, Gladys; Freijo, Roberto O; Portiansky, Enrique L; Barbeito, Claudio G; Macchi, Gustavo J; Díaz, Alcira O

2015-08-01

In the present study we histochemically and lectinhistochemically characterized the growing oocytes of the pink cuskeel (Genypterus blacodes). We used histochemical methods for the localization and characterization of glycoconjugates (GCs) and lectin histochemical techniques for the identification of specific sugar residues. We analyzed presence and distribution of GCs in the different structures of the growing follicles (cortical alveoli, globules, yolk granules and zona radiata). During the initial stage of vitellogenesis, the oocytes presented small yolk granules composed of GCs that gradually increased during exogenous vitellogenesis. These GCs contained moderate quantities of α-D-mannose, D-glucose, N-acetylglucosamine and N-acetyl-neuraminic acid. The cortical alveoli contained both neutral and carboxylated GCs, and lectin techniques detected N-acetylgalactosamine, galactose and L-fucose. The zona radiata showed a strong positive reaction to PAS and it reacted weakly with more specific techniques, such as KOH/PA*S and PA/Bh/KOH/PAS. This structure showed GCs with oxidizable vicinal diols, O-acyl sugars and sialic acid residues with different substitution types and presented N-acetylgalactosamine and L-fucose specific residues. The oocytes follicular envelope evidenced neutral and acidic non-sulfated GCs and high concentrations of α-D-mannose, D-glucose, galactose and N-acetylgalactosamine. The intergranular cytoplasmic GCs were mainly rich in α-D-mannose, D-glucose, N-acetylgalactosamine, N-acetylglucosamine and N-acetyl-neuraminic acid. These results enhance the comprehension of the structure and functionality of the pink cuskeel ovarian follicles, and provide a useful tool for the study of this tissue in other teleost species.

A rapid increase in macrophage-derived versican and hyaluronan in infectious lung disease.

PubMed

Chang, Mary Y; Tanino, Yoshinori; Vidova, Veronika; Kinsella, Michael G; Chan, Christina K; Johnson, Pamela Y; Wight, Thomas N; Frevert, Charles W

2014-02-01

The goals of this study were to characterize the changes in chondroitin sulfate proteoglycans and hyaluronan in lungs in acute response to gram-negative bacterial infection and to identify cellular components responsible for these changes. Mice were treated with intratracheal (IT) live Escherichia coli, E. coli lipopolysaccharide (LPS), or PBS. Both E. coli and LPS caused rapid selective increases in mRNA expression of versican and hyaluronan synthase (Has) isoforms 1 and 2 associated with increased immunohistochemical and histochemical staining for versican and hyaluronan in the lungs. Versican was associated with a subset of alveolar macrophages. To examine whether macrophages contribute to versican and hyaluronan accumulation, in vitro studies with primary cultures of bone marrow-derived and alveolar macrophages were performed. Unstimulated macrophages expressed very low levels of versican and hyaluronan synthase mRNA, with no detectible versican protein or hyaluronan product. Stimulation with LPS caused rapid increases in versican mRNA and protein, a rapid increase in Has1 mRNA, and concomitant inhibition of hyaluronidases 1 and 2, the major hyaluronan degrading enzymes. Hyaluronan could be detected following chloroquine pre-treatment, indicating rapid turnover and degradation of hyaluronan by macrophages. In addition, the effects of LPS, the M1 macrophage classical activation agonist, were compared to those of IL-4/IL-13 or IL-10, the M2a and M2c alternative activation agonists, respectively. Versican and Has1 increased only in response to M1 activation. Finally, the up-regulation of versican and Has1 in the whole lungs of wild-type mice following IT LPS was completely abrogated in TLR-4(-/-) mice. These findings suggest that versican and hyaluronan synthesis may play an important role in the innate immune response to gram-negative lung infection. Copyright © 2014 International Society of Matrix Biology. Published by Elsevier B.V. All rights reserved.

Histochemical study of the elastic fibers in pathologic human temporomandibular joint discs.

PubMed

Leonardi, R; Villari, L; Bernasconi, G; Caltabiano, M

2001-10-01

This study investigated histochemically the elastic fibers in human temporomandibular joint (TMJ) discs with varying degrees of tissue degeneration/regeneration to determine whether there are differences that correlate with the histologic findings. Ten diseased human TMJ discs and 2 control specimens were studied histochemically by staining with Weigert's resorcin-fuchsin after oxidation with peracetic acid. This technique selectively stains elastic, elaunin, (pre-elastic), and oxytalan fibers. In TMJ discs with an abnormal collagen fiber arrangement, an increased number of oxytalan fibers could be observed, contrary to discs with scar-like tissue transformation in which oxytalan fibers were decreased in number. In discs showing tears and clefts, the oxytalan fibers run perpendicular to the defects, whereas elaunin and elastic fibers were mainly circumferentially arranged. In discs with chondroid metaplasia, elastic, elaunin, and oxytalan fibers were extensively detected. It is hypothesized that the elastic, elaunin, and oxytalan fibers found in severely damaged discs appear to ensure biomechanical compliance by reinforcing regions devoid of collagen bundles and thus function as shock absorbers of stretch and compression. Copyright 2001 American Association of Oral and Maxillofacial Surgeons

Ultrastructural and histochemical study on the interrenal cells of the male stickleback (Gasterosteus aculeatus, Teleostea), in relation to the reproductive annual cycle

PubMed Central

CIVININI, ANNALENA; PADULA, DANIELA; GALLO, VALENTINA P.

2001-01-01

The steroidogenic interrenal cells in the adrenal homologue of the male stickleback (Gasterosteus aculeatus) were studied in relation to the reproductive cycle by means of histological and ultrastructural observations, and using histochemical methods for the localisation of 3β-hydroxysteroid dehydrogenase (3βHSD) and 17β-hydroxysteroid dehydrogenase (17βHSD). To determine the various stages of the reproductive cycle, the testes were also examined by histological and histochemical methods (3βHSD). The results indicate that in this teleost the interrenal cells can undergo a cycle in which phases characterised by different cytological aspects are observed. During this cycle there is a renewal of organelles, in particular mitochondria and SER. Periodic degenerative processes are also found. Organelle cytology showed that the cell cycle has at least 3 different aspects during the year. An analogy with some cytological aspects of the adrenal zonation in mammals is possible. It is postulated that the interrenal gland activity could substitute or supplement androgen production by the testes. PMID:11554507

Histochemical detection of lead in plant tissues

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tung, G.; Temple, P.J.

1996-06-01

A histochemical staining technique using sodium rhodizonate was developed for detecting lead in living or preserved plant tissues. Sodium rhodizonate formed a bright scarlet-red precipitate with lead at pH 3.0, but showed no significant color responses with other metals. The precipitation of lead by this staining technique was confirmed by detection of lead in the red-stained precipitate with electron microscopy X-ray analysis. This histochemical technique for lead provided rapid, quantifiable, and unambiguous evidence for the accumulation and localization of lead in plant tissues. Soil-borne lead accumulated primarily in the roots, although at high concentrations, lead also accumulated at the endsmore » of transpirational streams, particularly at hydathodes, trichomes, and the termini of xylem streams. Lead deposited from the atmosphere accumulated on the surface of conifer foliage and also appeared in or on cell walls of various internal cells and tissues. Lead concentrations in foliage and the color intensity of the stained deposits in spruce foliage decreased with distance from the lead source and increased with age of needles. No evidence of lead deposition inside cell contents was observed by this stain.« less

Histochemical and ultrastructural study of the chicken salivary palatine glands.

PubMed

Samar, María E; Avila, Rodolfo E; Esteban, Francisco J; Olmedo, Luis; Dettin, Luis; Massone, Adriana; Pedrosa, Juan A; Peinado, María A

2002-01-01

Salivary glands are a good model to investigate the relationship between cell secretion and glandular structure. Most studies of this organ deal with mammals, but we are interested in a morphofunctional characterization of these glands in poultry in relation with particular feeding habits. For this purpose, conventional and lectin histochemical methods as well as ultrastructural methods have been applied to the chicken lateral and medial palatine salivary glands. It was found that periodic acid-Schiff (PAS)-positive, alcianophilic, and metachromatic or orthochromatic cells were present with a more homogeneous distribution pattern in lateral glands than in medial palatine glands. Lectin staining depended on the lectin type that was applied, but also on the glandular part both in lateral and medial glands. Ultrastructural studies showed cytoplasmic membranous structures with a scattered granular or filamentous content depending on the secretory cell. In conclusion, morphofunctional characteristics of salivary glands of chicken suggest that their products are involved in lubrication and humidification of food ingested, and probably in protection of the oral surface, as has been previously described for other animals showing similar histochemical staining patterns.

Ectopic pinealoma: an unusual clinical presentation and a histochemical comparison with a seminoma of the testis

PubMed Central

Beeley, J. M.; Daly, J. J.; Timperley, W. R.; Warner, J.

1973-01-01

A patient with ectopic pinealoma first presented with apparent anorexia nervosa and hypernatraemic coma. A history of diabetes insipidus two months previously was not known on admission to hospital. The diabetes insipidus was unmasked by the administration of steroids. Neuroendocrinal and neuropathological aspects of the case are discussed with reference to the march of symptoms due to the growth of the tumour. Histochemical evidence is presented supporting the similarity between ectopic pinealoma and seminoma which suggests that they may more properly be referred to as atypical teratomas. Images PMID:4356733

Central core disease. A correlated genetic, histochemical, ultramicroscopic, and biochemical study.

PubMed Central

Isaacs, H; Heffron, J J; Badenhorst, M

1975-01-01

Two patients suffering from central core disease are presented. The condition is associated with musculoskeletal abnormalities which have been traced back over five generations. In addition to the typical histochemical findings, electronmicroscopic study has revealed the presence of both structured and non-structured cores in adjacent areas. The calcium uptake by the sarcoplasmic reticulum was reduced to one-third of normal. Phosphorylase activity was normal in the one case and reduced to 63% in the other. Actomyosin Mg2+-activated ATPase activity was decreased, as was the Ca2+-dependent ATPase of the sarcoplasmic reticulum. Images PMID:130467

Biological effects caused by low-power laser light in the treatment of the dentition, periodontium, and mucosa of oral cavity, and lip diseases

NASA Astrophysics Data System (ADS)

Kunin, Anatoly A.; Erina, Stanislava V.; Kashuba, Victor A.; Pankova, Svetlana N.; Stepanov, Nicolay N.; Kazmina, Svetlana G.; Dergunova, Elvira I.; Buerger, F.; Herdt, Alexander; Podolskaya, Elana E.; Shumilovitch, Bogdan R.; Ippolitov, Yu. A.; Tchernov, V. I.

1997-12-01

Nowadays low-power therapy is one of the leading trends in a combined treatment of the oral cavity and lips diseases. The present paper sums up the results of the investigation into the biological effects caused by low-power laser light (LPLL) during its interaction with hard and soft tissues of the oral cavity and lips. A research on the effect of LPLL upon the remineralization processes in the hard dental tissues in the stage in the stage of an initial caries was carried out in 150 patients. The biological effects caused by an interaction of LPLL with the parodontium tissues in the process of treatment of medium degree disease of the parodontium were studied in 140 patients; the effects of the above mentioned character which generated in lips tissues during treatment of a post-radiation chilitis were analyzed in 32 patients. Immunological, biochemical histochemical, morphological, stomatoscopic, bacteriological and other methods were employed while studying the bioeffects caused by LPLL in the parodontium, lips tissues and hard tissues of the tooth.

Ecto-5′-Nucleotidase Activity in Human T Cell Subsets. DECREASED NUMBERS OF ECTO-5′-NUCLEOTIDASE POSITIVE CELLS FROM BOTH OKT4+ AND OKT8+ CELLS IN PATIENTS WITH HYPOGAMMAGLOBULINEMIA

PubMed Central

Thompson, Linda F.; Saxon, Andrew; O'Connor, Richard D.; Fox, Robert I.

1983-01-01

T lymphocytes from control subjects were separated into subsets using monoclonal antibodies of the OKT series and complement lysis and analyzed for ecto-5′-nucleotidase activity both by quantitative radiochemical assay and a histochemical stain. T cells from 15 control subjects contained 54±4% OKT4+ (helper/inducer) cells and 32±3% OKT8+ (cytotoxic/suppressor) cells. Total T cell ecto-5′-nucleotidase activity was 10.9±2.1 nmol/h per 106 cells with 25±7% positive by histochemical stain. Ecto-5′-nucleotidase activity in OKT4-enriched populations was 5.43±1.8 nmol/h per 106 cells with 14±2% positive by histochemical stain; that in OKT8-enriched populations was 17.1±5.9 nmol/h per 106 cells with 35±8% positive by histochemical stain. Two of four patients with congenital agammaglobulinemia and four of seven patients with common variable immunodeficiency had decreased proportions of OKT4+ T cells with corresponding increases in the proportions of OKT8+ T cells (OKT4/OKT8 = 0.60 to 1.0 as compared with 1.7±0.2 for control subjects). All four patients with congenital agammaglobulinemia, and three of seven patients with common variable immunodeficiency also had low T cell ecto-5′-nucleotidase activity (<5.5 nmol/h per 106 cells). Ecto-5′-nucleotidase activity in OKT4- enriched populations isolated from four patients with low total T cell activity was 2.85±0.90 nmol/h per 106 cells with 10±4% positive by histochemical stain; that in OKT8-enriched populations was 6.82±1.7 nmol/h per 106 cells with 7.5±3% positive by histochemical stain. Thus, the number of ecto-5′-nucleotidase positive cells is decreased, especially in the OKT8+ subpopulation, and the low total T cell ecto-5′-nucleotidase activity seen in these patients is due to fewer positive cells rather than to substantially less activity per cell. Our data indicate that ecto-5′-nucleotidase activity defines two subpopulations of T lymphocytes (ecto-5′-nucleotidase positive and negative), the proportions of which are markedly altered in many patients with hypogammaglobulinemia. In preliminary studies with seven patients, increased numbers of ecto-5′-nucleotidase negative T cells appeared to correlate with increased suppressor T cell activity toward in vitro immunoglobulin synthesis. Therefore, ecto-5′-nucleotidase may be a useful cell surface marker in the study of imbalances of regulatory T cell subsets in patients with antibody synthesis disorders. PMID:6300192

A Fischer rat substrain deficient in dipeptidyl peptidase IV activity makes normal steady-state RNA levels and an altered protein. Use as a liver-cell transplantation model.

PubMed Central

Thompson, N L; Hixson, D C; Callanan, H; Panzica, M; Flanagan, D; Faris, R A; Hong, W J; Hartel-Schenk, S; Doyle, D

1991-01-01

Dipeptidyl peptidase IV (DPPIV) is a serine exoproteinase expressed at high levels in epithelial cells of kidney, liver and small intestine. Recently Watanabe, Kohima & Fujimoto [(1987) Experientia 43, 400-401] and Gossrau et al. [(1990) Histochem. J. 22, 172-173] reported that Fischer 344 rats are deficient in this enzyme. We have examined DPPIV expression in Fischer 344 rats available from U.S. and German suppliers and find that livers of the U.S. Fischer rats, in contrast with their German counterparts, express active DPPIV (D+). Northern analysis of liver RNA showed comparable levels of 3.4 kb and 5.6 kb DPPIV transcripts in both D+ rats from the U.S. and German (D-) rats. Monoclonal antibody (MAb) 236.3 to DPPIV immunoprecipitated at 150 kDa enzymically active (105 kDa, denatured) protein from surface-labelled D+ hepatocytes and reacted with canalicular and sinusoidal membranes (as shown by immunofluorescence microscopy). MAb 236.3 failed to immunoprecipitate a labelled peptide from D- cell extract or to stain D- liver sections. Polyclonal antibody (PAb) specific for DPPIV immunoprecipitated an enzymically active peptide from D+ hepatocyte extracts and a smaller, inactive peptide from D- hepatocyte extracts. Peptide maps of DPPIV immunoprecipitated from D+ extracts with MAb 236.3 and PAb were identical, but differed from that of the D- hepatocyte component recognized by PAb. The molecular basis of the DPPIV deficiency in the D- rats thus appears to be the translation of an enzymically inactive protein missing the epitope recognized by MAb 236.3. We have exploited these D- rats as hosts for syngeneic transplantation of liver cells from D+ Fischer rats. DPPIV expression is stable in the transplanted cells and allows them to be readily distinguished from the surrounding D- tissue. Images Fig. 1. Fig. 2. Fig. 3. Fig. 4. PMID:1705112

Profiling and functional classification of esterases in olive (Olea europaea) pollen during germination

PubMed Central

Rejón, Juan D.; Zienkiewicz, Agnieszka; Rodríguez-García, María Isabel; Castro, Antonio J.

2012-01-01

Background and Aims A pollen grain contains a number of esterases, many of which are released upon contact with the stigma surface. However, the identity and function of most of these esterases remain unknown. In this work, esterases from olive pollen during its germination were identifided and functionally characterized. Methods The esterolytic capacity of olive (Olea europaea) pollen was examined using in vitro and in-gel enzymatic assays with different enzyme substrates. The functional analysis of pollen esterases was achieved by inhibition assays by using specific inhibitors. The cellular localization of esterase activities was performed using histochemical methods. Key Results Olive pollen showed high levels of non-specific esterase activity, which remained steady after hydration and germination. Up to 20 esterolytic bands were identified on polyacrylamide gels. All the inhibitors decreased pollen germinability, but only diisopropyl fluorophosphate (DIFP) hampered pollen tube growth. Non-specific esterase activity is localized on the surface of oil bodies (OBs) and small vesicles, in the pollen intine and in the callose layer of the pollen tube wall. Acetylcholinesterase (AChE) activity was mostly observed in the apertures, exine and pollen coat, and attached to the pollen tube wall surface and to small cytoplasmic vesicles. Conclusions In this work, for the first time a systematic functional characterization of esterase enzymes in pollen from a plant species with wet stigma has been carried out. Olive pollen esterases belong to four different functional groups: carboxylesterases, acetylesterases, AChEs and lipases. The cellular localization of esterase activity indicates that the intine is a putative storage site for esterolytic enzymes in olive pollen. Based on inhibition assays and cellular localization of enzymatic activities, it can be concluded that these enzymes are likely to be involved in pollen germination, and pollen tube growth and penetration of the stigma. PMID:22922586

Effect of biologically synthesized copper oxide nanoparticles on metabolism and antioxidant activity to the crop plants Solanum lycopersicum and Brassica oleracea var. botrytis.

PubMed

Singh, Ajey; Singh, N B; Hussain, Imtiyaz; Singh, Himani

2017-11-20

Study on the ecological effect of metal oxide nanomaterials (NMs) has quickly amplified over the precedent years because it is assumed that these NMs will sooner or later be released into the environment. The present study deals with biologically oriented process for the green synthesis of copper oxide nanoparticles (CuO NPs) by using Morus alba leaf extract as reducing agent. Powder X-ray diffraction (XRD) and transmission electron microscope (TEM) analysis revealed the monoclinic phase and 20-40nm size respectively. The presence of reducing and capping agents revealed by Fourier transform infrared (FTIR) spectroscopy. The seedlings of Brassica oleracea var. botrytis and Solanum lycopersicum were exposed to 10, 50, 100, and 500mgL -1 concentrations of CuO NPs in the sand medium. Bioaccumulation of Cu was also investigated by atomic absorption spectroscopy (AAS). Plant exposure to 100 and 500mgL -1 of CuO NPs has resulted in significant reduction of total chlorophyll and sugar content in the two test plants while 10mgL -1 of NPs slightly increased the pigment and sugar content in tomato plants only. Augmentation of lipid peroxidation, electrolyte leakage, and antioxidant enzyme activity was observed in a dose dependent manner upon plants exposure to CuO NPs. Deposition of lignin in roots of both plants treated with the highest concentration of CuO NPs was observed. Histochemical analysis of leaves of treated plant with nitroblue tetrazolium and 3 ' 3 ' diaminobenzidine showed a concentration dependent increase in superoxide and hydrogen peroxide formation in leaves. The green synthesis of CuO NPs was carried out by using Morus alba leaf extract. Accumulation of NPs more actively by tomato plants as compared to cauliflower was possibly due to the difference in root morphology. The histochemical visualization highlights the spatial organization of oxidant biochemistry occurring in response to metal stress. Copyright © 2017 Elsevier B.V. All rights reserved.

Remarkable heterogeneity in myosin heavy-chain composition of the human young masseter compared with young biceps brachii.

PubMed

Osterlund, Catharina; Lindström, Mona; Thornell, Lars-Eric; Eriksson, Per-Olof

2012-10-01

Adult human jaw muscles differ from limb and trunk muscles in enzyme-histochemical fibre type composition. Recently, we showed that the human masseter and biceps differ in fibre type pattern already at childhood. The present study explored the myosin heavy-chain (MyHC) expression in the young masseter and biceps muscles by means of gel electrophoresis (GE) and immuno-histochemical (IHC) techniques. Plasticity in MyHC expression during life was evaluated by comparing the results with the previously reported data for adult muscles. In young masseter, GE identified MyHC-I, MyHC-IIa MyHC-IIx and small proportions of MyHC-fetal and MyHC-α cardiac. Western blots confirmed the presence of MyHC-I, MyHC-IIa and MyHC-IIx. IHC revealed in the masseter six isomyosins, MyHC-I, MyHC-IIa, MyHC-IIx, MyHC-fetal, MyHC α-cardiac and a previously not reported isoform, termed MyHC-IIx'. The majority of the masseter fibres co-expressed two to four isoforms. In the young biceps, both GE and IHC identified MyHC-I, MyHC-IIa and MyHC-IIx. MyHC-I predominated in both muscles. Young masseter showed more slow and less-fast and fetal MyHC than the adult and elderly masseter. These results provide evidence that the young masseter muscle is unique in MyHC composition, expressing MyHC-α cardiac and MyHC-fetal isoforms as well as hitherto unrecognized potential spliced isoforms of MyHC-fetal and MyHC-IIx. Differences in masseter MyHC expression between young adult and elderly suggest a shift from childhood to adulthood towards more fast contractile properties. Differences between masseter and biceps are proposed to reflect diverse evolutionary and developmental origins and confirm that the masseter and biceps present separate allotypes of muscle.

Polar localization of plasma membrane Ca2+/Mg2+ ATPase correlates with the pattern of steady ionic currents in eggs ofLymnaea stagnalis andBithynia tentaculata (Mollusca).

PubMed

Zivkovic, Danica; Créton, Robbert; Zwaan, Gideon; de Bruijn, Willem C; Dohmen, M René

1990-11-01

During extrusion of the first polar body in eggs ofLymnaea stagnalis andBithynia tentaculata a localized Ca 2+ /Mg 2+ ATPase activity was detected, using Ando's enzyme-cytochemical method for electron microscopy [Ando et al. (1981) Acta Histochem Cytochem 14:705-726]. The enzyme activity was distributed in a polar fashion, along the cytoplasmic face of the plasma membrane. In the eggs ofLymnaea it was found only in the vegetal hemisphere, whereas inBithynia eggs it was localized both in the vegetal hemisphere and at the animal pole. This pattern of enzyme activity corresponds to the polar pattern of transcellular ionic currents measured with the vibrating probe, which we showed to be partially carried or regulated by calcium [Zivkovic and Dohmen (1989) Biol Bull (Woods Hole) 176 (Suppl):103-109]. The characteristics of the ATPase were studied using a variety of approaches such as ion and substrate depletions and substitutions, addition of specific inhibitors of ATPase activity, treatment with EDTA/EGTA and electron energy-loss spectrometry. The results indicate that, inLymnaea, there are at least two enzymatic entities. The first one is a Ca 2+ /Mg 2+ ATPase localized along the membrane and in the cortex of the vegetal hemisphere. The second one is a Ca 2+ -stimulated ATPase (calcium pump of the plasma membrane) localized in a small region of the membrane at the vegetal pole. We speculate that in the eggs ofLymnaea andBithynia a functional relationship exists between the plasma-membrane-associated ATPase activity and the transcellular ionic currents measured in the same region.

Painful unilateral temporalis muscle enlargement: reactive masticatory muscle hypertrophy.

PubMed

Katsetos, Christos D; Bianchi, Michael A; Jaffery, Fizza; Koutzaki, Sirma; Zarella, Mark; Slater, Robert

2014-06-01

An instance of isolated unilateral temporalis muscle hypertrophy (reactive masticatory muscle hypertrophy with fiber type 1 predominance) confirmed by muscle biopsy with histochemical fiber typing and image analysis in a 62 year-old man is reported. The patient presented with bruxism and a painful swelling of the temple. Absence of asymmetry or other abnormalities of the craniofacial skeleton was confirmed by magnetic resonance imaging and cephalometric analyses. The patient achieved symptomatic improvement only after undergoing botulinum toxin injections. Muscle biopsy is key in the diagnosis of reactive masticatory muscle hypertrophy and its distinction from masticatory muscle myopathy (hypertrophic branchial myopathy) and other non-reactive causes of painful asymmetric temporalis muscle enlargement.

Gold nanoparticles induced cloudy swelling to hydropic degeneration, cytoplasmic hyaline vacuolation, polymorphism, binucleation, karyopyknosis, karyolysis, karyorrhexis and necrosis in the liver.

PubMed

Abdelhalim, Mohamed Anwar K; Jarrar, Bashir M

2011-09-22

Nanoparticles (NPs) can potentially cause adverse effects on organ, tissue, cellular, subcellular and protein levels due to their unusual physicochemical properties. Advances in nanotechnology have identified promising candidates for many biological and biomedical applications. The aim of the present study was to investigate the particle-size, dose and exposure duration effects of gold nanoparticles (GNPs) on the hepatic tissue in an attempt to cover and understand the toxicity and their potential therapeutic and diagnostic use. A total of 70 healthy male Wistar-Kyoto rats were exposed to GNPs received 50 or 100 ul of GNPs infusion of size (10, 20 and 50 nm for 3 or 7 days) to investigate particle-size, dose and exposure duration effects of GNPs on the hepatic tissue. In comparison with respective control rats, exposure to GNPs doses has produced alterations in the hepatocytes, portal triads and the sinusoids. The alterations in the hepatocytes were mainly vacuolar to hydropic degeneration, cytopasmic hyaline vacuolation, polymorphism, binucleation, karyopyknosis, karyolysis, karyorrhexis and necrosis. The hepatocytes swelling might be exhibited as a result of disturbances of membranes function that lead to massive influx of water and Na+ due to GNPs effects accompanied by leakage of lysosomal hydrolytic enzymes that lead to cytoplasmic degeneration and macromolecular crowding. Hydropic degeneration is a result of ion and fluid homestasis that lead to an increase of intracellular water. The vacuolated swelling of the cytoplasm of the hepatocytes of the GNPs treated rats might indicate acute and subacute liver injury induced by the GNPs. Binucleation represents a consequence of cell injury and is a sort of chromosomes hyperplasia which is usually seen in regenerating cells. The induced histological alterations might be an indication of injured hepatocytes due to GNPs toxicity that became unable to deal with the accumulated residues resulting from metabolic and structural disturbances caused by these NPs. These alterations were size-dependent with smaller ones induced the most effects and related with time exposure of GNPs. The appearance of hepatocytes cytoplasmic degeneration and nuclear destruction may suggest that GNPs interact with proteins and enzymes of the hepatic tissue interfering with the antioxidant defense mechanism and leading to reactive oxygen species (ROS) generation which in turn may induce stress in the hepatocytes to undergo atrophy and necrosis. More histomorphologcal, histochemical and ultrastrucural investigations are needed in relation of the application of GNPs with their potential role as a therapeutic and diagnostic tool.

Gold nanoparticles induced cloudy swelling to hydropic degeneration, cytoplasmic hyaline vacuolation, polymorphism, binucleation, karyopyknosis, karyolysis, karyorrhexis and necrosis in the liver

PubMed Central

2011-01-01

Background Nanoparticles (NPs) can potentially cause adverse effects on organ, tissue, cellular, subcellular and protein levels due to their unusual physicochemical properties. Advances in nanotechnology have identified promising candidates for many biological and biomedical applications. The aim of the present study was to investigate the particle-size, dose and exposure duration effects of gold nanoparticles (GNPs) on the hepatic tissue in an attempt to cover and understand the toxicity and their potential therapeutic and diagnostic use. Methods A total of 70 healthy male Wistar-Kyoto rats were exposed to GNPs received 50 or 100 ul of GNPs infusion of size (10, 20 and 50 nm for 3 or 7 days) to investigate particle-size, dose and exposure duration effects of GNPs on the hepatic tissue. Results In comparison with respective control rats, exposure to GNPs doses has produced alterations in the hepatocytes, portal triads and the sinusoids. The alterations in the hepatocytes were mainly vacuolar to hydropic degeneration, cytopasmic hyaline vacuolation, polymorphism, binucleation, karyopyknosis, karyolysis, karyorrhexis and necrosis. Conclusions The hepatocytes swelling might be exhibited as a result of disturbances of membranes function that lead to massive influx of water and Na+ due to GNPs effects accompanied by leakage of lysosomal hydrolytic enzymes that lead to cytoplasmic degeneration and macromolecular crowding. Hydropic degeneration is a result of ion and fluid homestasis that lead to an increase of intracellular water. The vacuolated swelling of the cytoplasm of the hepatocytes of the GNPs treated rats might indicate acute and subacute liver injury induced by the GNPs. Binucleation represents a consequence of cell injury and is a sort of chromosomes hyperplasia which is usually seen in regenerating cells. The induced histological alterations might be an indication of injured hepatocytes due to GNPs toxicity that became unable to deal with the accumulated residues resulting from metabolic and structural disturbances caused by these NPs. These alterations were size-dependent with smaller ones induced the most effects and related with time exposure of GNPs. The appearance of hepatocytes cytoplasmic degeneration and nuclear destruction may suggest that GNPs interact with proteins and enzymes of the hepatic tissue interfering with the antioxidant defense mechanism and leading to reactive oxygen species (ROS) generation which in turn may induce stress in the hepatocytes to undergo atrophy and necrosis. More histomorphologcal, histochemical and ultrastrucural investigations are needed in relation of the application of GNPs with their potential role as a therapeutic and diagnostic tool. PMID:21939512

Renal denervation using focused infrared fiber lasers: a potential treatment for hypertension.

PubMed

Alexander, Vinay V; Shi, Zhennan; Iftekher, Fariha; Welsh, Michael J; Gurm, Hitinder S; Rising, Gail; Yanovich, Amber; Walacavage, Kim; Islam, Mohammed N

2014-11-01

Renal denervation has recently become of great interest as a potential treatment for resistant hypertension. Denervation techniques using radio frequency (RF) or ultrasound energy sources have already been explored in literature. In this study, we investigate the use of lasers as a potential energy source for renal denervation. In vitro studies are performed in porcine/ovine renal arteries with focused laser beams at 980 nm, 1210 nm, and 1700 nm to study the ability to damage renal nerves without causing injury to non-target tissue structures like the endothelium. Then, a 980 nm laser catheter prototype is built and used to demonstrate in vivo renal denervation in ovine renal arteries. This study utilizes fiber coupled infrared lasers at 980 nm, 1210 nm, and 1700 nm. In vitro laser denervation studies at 980 nm are performed in both porcine and ovine renal arteries to study the ability of focused laser beams to damage renal nerves without injuring the endothelium. In vitro studies using lasers close to the lipid absorption lines at 1210 nm and 1700 nm are also performed in porcine renal arteries to study the possibility of selectively damaging the renal nerves by targeting the lipid myelin sheaths surrounding the nerves. Then, a laser catheter prototype is designed and built for in vivo renal denervation in ovine renal arteries using the 980 nm laser (powers ranging from 2 to 4 W, 5 seconds per exposure). Histochemical evaluations of the frozen sections are performed using methylthiazolyldiphenyl-tetrazolium bromide (MTT) assay. Histochemical analysis of in vitro laser treatments at 980 nm in porcine and ovine renal arteries show clear evidence of laser-induced renal nerve damage without injury to the endothelium and part of the media. No evidence of selective nerve damage is observed using the 1210 nm and 1700 nm lasers with the current treatment parameters. Histochemical analysis of in vivo laser treatments in ovine renal arteries using a focused 980 nm laser show clear evidence of renal nerve damage with depths of damage extending > 1.5 mm from the artery wall. Sections with laser-induced damage to the media/adventitia at depths of > 1 mm without injury to the endothelium are also observed. We demonstrate the use of focused lasers as an attractive energy source for causing renal nerve damage without injury to the artery wall and thus, may have potential therapeutic applications for conditions such as resistant hypertension, where renal denervation has been shown to be a promising form of treatment. © 2014 Wiley Periodicals, Inc.

[Histochemical study of the digestive organs of rats after a flight on "Kosmos-605"].

PubMed

Shubich, M G; Goriacheva, L L; Dudetskiĭ, V I; Lutsenko, N M; Mogil'naia, G M

1977-01-01

The histochemical study of the stomach, small and large intestines and pancreas of rats flown aboard the biosatellite Cosmos-605 as well as of synchronous and vivarium controls demonstrated a significant decline in the mucine producing capacity of epithelial cells of the stomach of the flight rats on the R + 1 day. The study showed an increased content of sialo- and sulphosaccharides in goblet cells of cryptae of large intestine and a reduced content of free cation protein in the acinar cells of the pancreas of flight rats. The changes were transient and disappeared by the R + 26 day.

Electrophysiological, histochemical, and hormonal adaptation of rat muscle after prolonged hindlimb suspension

NASA Astrophysics Data System (ADS)

Kourtidou-Papadeli, Chrysoula; Kyparos, Antonios; Albani, Maria; Frossinis, Athanasios; Papadelis, Christos L.; Bamidis, Panagiotis; Vivas, Ana; Guiba-Tziampiri, Olympia

2004-05-01

The perspective of long-duration flights for future exploration, imply more research in the field of human adaptation. Previous studies in rat muscles hindlimb suspension (HLS), indicated muscle atrophy and a change of fibre composition from slow-to-fast twitch types. However, the contractile responses to long-term unloading is still unclear. Fifteen adult Wistar rats were studied in 45 and 70 days of muscle unweighting and soleus (SOL) muscle as well as extensor digitorum longus (EDL) were prepared for electrophysiological recordings (single, twitch, tetanic contraction and fatigue) and histochemical stainings. The loss of muscle mass observed was greater in the soleus muscle. The analysis of electrophysiological properties of both EDL and SOL showed significant main effects of group, of number of unweighting days and fatigue properties. Single contraction for soleus muscle remained unchanged but there was statistically significant difference for tetanic contraction and fatigue. Fatigue index showed a decrease for the control rats, but increase for the HLS rats. According to the histochemical findings there was a shift from oxidative to glycolytic metabolism during HLS. The data suggested that muscles atrophied, but they presented an adaptation pattern, while their endurance in fatigue was decreased.

Salivary gland oncocytes in African hedgehogs (Atelerix albiventris) mimicking cytomegalic inclusion disease.

PubMed

Brunnert, S R; Hensley, G T; Citino, S B; Herron, A J; Altman, N H

1991-07-01

The salivary glands from three African hedgehogs contained multiple foci of cytomegalic cells, which occasionally had a mild to moderate infiltrate of lymphocytes at the periphery. The cytomegalic cells were 35 to 40 microns in diameter with abundant acidophilic granular to hyalin cytoplasm. The nuclei were enlarged with clumped marginalized chromatin and a large, (6 to 8 microns in diameter) central, brightly eosinophilic nucleolus that had the appearance of an inclusion body by light microscopy. Histochemically most of the cytomegalic cells contained cytoplasmic metachromatic granules with Feyrter's thionine inclusion stain. Scattered cells at the periphery of the cytomegalic foci contained periodic acid-Schiff-positive cytoplasmic granules. Ultrastructurally the cytomegalic cells contained numerous tightly-packed, often bizarre, enlarged mitochondria that completely filled the cytoplasm. The nucleus consisted of a dense central core of chromatin associated with the nucleolus and the remaining chromatin was clumped and marginalized. Nuclear and cytoplasmic virions consistent with cytomegalovirus were not present. Histochemical stains of the nucleus for heavy metals were negative. The ultrastructural and histochemical findings of the cytomegalic cells were consistent with oncocytes. Previous reports in the literature of similar cells in the salivary glands of insectivores appear to have been erroneously described as cytomegalovirus infections.

The carbohydrate deposits detected by histochemical methods in the molecular layer of the dentate gyrus in the hippocampal formation of patients with schizophrenia, Down's syndrome and dementia, and aged person.

PubMed

Nishimura, A; Ikemoto, K; Satoh, K; Yamamoto, Y; Rand, S; Brinkmann, B; Nishi, K

2000-11-01

Post-mortem brain tissue was obtained from 28 patients with brain disorders, of which 15 had clinically diagnosed schizophrenia, 6 Alzheimer type dementia, 5 dementia with tangles and 2 cases of Down's syndrome. The controls were 22 cases from autopsies without brain disorders or with no known episodes of brain disorder. The tissues were stained for the detection of carbohydrate deposits in the hippocampal formation, using lectin, immunohistochemical and conventional staining methods. The staining revealed the existence of spherical deposits in the inner and middle molecular layers of the dentate gyrus in the hippocampal formation which contained fucose, galactose, N-acetyl galactosamine, N-acetyl glucosamine, sialic acid, mannose and chondroitin sulfate. The number of the deposits was higher in patients with brain disorder such as schizophrenia, Alzheimer type dementia, dementia with tangles or Down's syndrome, and in some aged individuals, in comparison to those in younger individuals. No deposits were detected in a few younger or aged individuals. Spherical deposits 3-10 microm in diameter may be an immature form of the corpora amylacea, since they were similar in the histochemical characteristics with lectin, immunohistochemical and conventional staining methods. However, differing staining ability by hematoxylin, periodic acid Schiff's reagent and antibodies against the intracellular degraded proteins such as ubiquitin and tau-protein was observed. The antibodies against ubiquitin and tau-protein showed clear reactivity with the corpora amylacea and no reactivity with spherical deposits, indicating that the corpora amylacea has an intracellular origin and spherical deposits an extracellular matrix origin. The results obtained in this study indicate that not only neuronal degeneration but also unusual glycometabolism in neurons may disturb the neuronal function and cause brain disorders, and that spherical deposits may cause dysfunction of the neuronal network in the dentate gyrus of the hippocampus which is closely linked with recognition and memory functions.

Copper/zinc superoxide dismutase insufficiency impairs progesterone secretion and fertility in female mice.

PubMed

Noda, Yoshihiro; Ota, Kuniaki; Shirasawa, Takuji; Shimizu, Takahiko

2012-01-01

Copper/zinc superoxide dismutase (CuZn-SOD, SOD1) is one of the major antioxidant enzymes, and is localized in the cytoplasm to scavenge superoxide. To investigate the physiological role of SOD1 in the ovaries, we analyzed the fertility of Sod1-deficient female mice. To evaluate their hormonal metabolism, we measured pituitary and ovarian hormone levels in the plasma of the mutant mice. Plasma follicle-stimulating hormone, luteinizing hormone, and estradiol were not altered in the mutant compared to the wild-type females, while the plasma progesterone level was significantly reduced in the mutant females. Furthermore, the mutant mice showed decreased progesterone secretion under the condition of superovulation. In a histochemical analysis, we observed a remarkable reduction in the corpus luteum area in the mutant ovaries without atrophic changes. The mutant mice also displayed enhanced superoxide generation in the region surrounding the corpora lutea, which was associated with increased apoptotic cells and suppressed vasculature. These results suggested that SOD1 deficiency dysregulated luteal formation because of increased superoxide generation in the ovary. In vitro fertilization experiments showed no abnormal fertilization of Sod1-deficient oocytes. In addition, when Sod1-deficient embryos were transferred into the oviducts of wild-type females, mutant embryos developed at a normal rate, indicating that SOD1 deficiency in embryos did not cause miscarriage in the uterus of wild-type females. These results indicated that increased intracellular ROS impaired luteal formation and progesterone production in the mutant females, thus suggesting that SOD1 plays a crucial role in both the luteal function and the maintenance of fertility in female mice.

De novo transcriptome sequencing of black pepper (Piper nigrum L.) and an analysis of genes involved in phenylpropanoid metabolism in response to Phytophthora capsici.

PubMed

Hao, Chaoyun; Xia, Zhiqiang; Fan, Rui; Tan, Lehe; Hu, Lisong; Wu, Baoduo; Wu, Huasong

2016-10-21

Piper nigrum L., or "black pepper", is an economically important spice crop in tropical regions. Black pepper production is markedly affected by foot rot disease caused by Phytophthora capsici, and genetic improvement of black pepper is essential for combating foot rot diseases. However, little is known about the mechanism of anti- P. capsici in black pepper. The molecular mechanisms underlying foot rot susceptibility were studied by comparing transcriptome analysis between resistant (Piper flaviflorum) and susceptible (Piper nigrum cv. Reyin-1) black pepper species. 116,432 unigenes were acquired from six libraries (three replicates of resistant and susceptible black pepper samples), which were integrated by applying BLAST similarity searches and noted by adopting Kyoto Encyclopaedia of Genes and Gene Ontology (GO) genome orthology identifiers. The reference transcriptome was mapped using two sets of digital gene expression data. Using GO enrichment analysis for the differentially expressed genes, the majority of the genes associated with the phenylpropanoid biosynthesis pathway were identified in P. flaviflorum. In addition, the expression of genes revealed that after susceptible and resistant species were inoculated with P. capsici, the majority of genes incorporated in the phenylpropanoid metabolism pathway were up-regulated in both species. Among various treatments and organs, all the genes were up-regulated to a relatively high degree in resistant species. Phenylalanine ammonia lyase and peroxidase enzyme activity increased in susceptible and resistant species after inoculation with P. capsici, and the resistant species increased faster. The resistant plants retain their vascular structure in lignin revealed by histochemical analysis. Our data provide critical information regarding target genes and a technological basis for future studies of black pepper genetic improvements, including transgenic breeding.

Interrupted orthodontic force results in less root resorption than continuous force in human premolars as measured by microcomputed tomography.

PubMed

Sawicka, Monika; Bedini, Rossella; Wierzbicki, Piotr M; Pameijer, Cornelis H

2014-01-01

Root resorption is an undesirable but very frequently occurring sequel of orthodontic treatment. The aim of this study was to compare root resorption caused by either continuous (CF) or interrupted (IF) orthodontic force. The study was performed on human subjects on 30 first upper and lower premolars scheduled for extraction for orthodontic reasons. During four weeks before extraction 12 teeth were subjected to either CF or IF. The force was generated by a segmental titanium-molybdenum alloy cantilever spring that was activated in buccal direction. Initially a force of 60 CentiNewton was used in both CF and IF groups, the force in the former, however, was reactivated every week for 4 weeks. There was no reactivation of force in the IF group after initial application. A morphometric analysis of root resorption was performed by microcomputed tomography and the extent of tooth movement was measured on stone casts. Furthermore, a Tartarate-Resistant Acidic Phosphatase activity (TRAP), the marker enzyme of osteoclasts and cementoclasts, was determined by histochemical method. The Mann-Whitney U test was used to compare the difference in measured parameters between treatment and control tooth groups. The number of resorption craters was significantly higher and their average volume almost twice as large in the CF compared to the IF group (p < 0.05). However, the distance of tooth displacement was similar for both groups. Cementoclasts were detected with the TRAP technique on the surface of two teeth only; both were subjected to continuous force. The use of IF leads to less destruction of root structure as opposed to continuous force while the same tooth movement was achieved.

Depletion of intracellular zinc from neurons by use of an extracellular chelator in vivo and in vitro.

PubMed

Frederickson, Christopher J; Suh, Sang W; Koh, Jae-Young; Cha, Yoo K; Thompson, Richard B; LaBuda, Christopher J; Balaji, Rengarajan V; Cuajungco, Math P

2002-12-01

The membrane-impermeable chelator CaEDTA was introduced extracellularly among neurons in vivo and in vitro for the purpose of chelating extracellular Zn(2+). Unexpectedly, this treatment caused histochemically reactive Zn(2+) in intracellular compartments to drop rapidly. The same general result was seen with intravesicular Zn(2+), which fell after CaEDTA infusion into the lateral ventricle of the brain, with perikaryal Zn(2+) in Purkinje neurons (in vivo) and with cortical neurons (in vitro). These findings suggest either that the volume of zinc ion efflux and reuptake is higher than previously suspected or that EDTA can enter cells and vesicles. Caution is therefore warranted in attempting to manipulate extracellular or intracellular Zn(2+) selectively.

Studies of Intercellular Communication and Intracellular Metabolic Responses by Bone Cells to Simulated Weightlessness

NASA Technical Reports Server (NTRS)

Doty, Stephen B.

1997-01-01

Spaceflight affects the weight bearing skeletal tissues by reducing the rate of new bone formation. This effect on the long bones of flown rats has been quantitated but the effect at the cellular level and the mechanism(s) involved are not understood. We are applying electron microscopy, coupled with histochemistry and immunocytochemistry to determine the cellular functions most affected by spaceflight. The emphasis for study of these samples from SLS-1, a 9-day mission, is on the histochemical and structural changes of the endosteal and perivascular osteoblasts found in diaphyseal bone of femur and tibia. Work is still in progress but some findings are described: (1) An expected decrease in alkaline phosphatase activity in osteoblasts from flight animals, but an increase in enzyme activity in the stromal stem cells adjacent to the osteoblast. (2) An increase in osteoclastic TRAP activity in the trabecular bone region in response to spaceflight. (3) A large increase in procollagen containing secretory granules in osteoblasts in the recovery group, and a significant decrease in granule numbers in the flight group.

[Metabolism of thyroid gland cells as affected by prolactin and emotional-physical stress].

PubMed

Strizhkov, V V

1991-01-01

A study was made of the role of prolactin (PRL) in the regulation of thyroid function in intact animals and in those exposed to stress (swimming was used as physical exercise). A single daily dose of 125 micrograms of PRL per 100 g of body mass was injected subcutaneously in 0.5 ml of saline solution during a week to male rats (control: intact rats; injection of 0.5 ml of saline solution subcutaneously). Redox enzymes; succinate dehydrogenase, lactate dehydrogenase, glucose-6-phosphate dehydrogenase, NAD.H2 and NADP.H2, ATPase and monoamine oxidase, total protein, RNA and glycogen in glandular cells were investigated histochemically 24 h after the last injection of PRL or saline, 30 min., 1, 2, 3, 5 and 7 hours after swimming or right after complete fatigue (in the presence of experimental hyperprolactinemia). A conclusion has been made that one of the most important mechanisms of the adaptive effect of PRL is its ability to suppress thyroid function, thus decreasing the metabolism level, which results in reduction of oxygen consumption and improves body tolerance to stress.

Melanosome degradation: fact or fiction.

PubMed

Borovanský, Jan; Elleder, Milan

2003-06-01

Our mini review summarizes what is known about the (bio)degradation of melanosomes. Unlike melanosome biogenesis where our knowledge enables us to explain it in molecular terms posing many interesting questions on the relation between lysosomes and melanosomes, melanosome degradation has remained 'terra incognita'. Observations at optical and ultrastructural levels describe the disintegration of melanosomes in the lysosomal compartment (in auto- and heterophagosomes). Histochemical studies suggest the participation of acid hydrolases in the process of melanosome degradation. Biochemical data confirm the ability of lysosomal hydrolases to degrade melanosome constituents except the melanin moiety. The similarity of melanin structure to that of polycyclic aromatic hydrocarbons suggests that melanin should be sensitive mainly, if not exclusively, to oxidative breakdown. In vitro melanin can indeed be decomposed by an oxidative attack and the degradation is accompanied by fluorescence and decreasing absorbance. From enzymes engaged in the biotransformation of polycyclic hydrocarbons only phagosomal NADPH oxidase meets the criteria (particularly as for compartmental and catalytic properties) to be involved in melanin biodegradation. The in vivo biodegradation of melanin has so far been clearly demonstrated in Aspergillus and fungi melanins.

[Morphological changes in the thyroid gland of rats during various phases of the estral cycle].

PubMed

Pliner, L I; Ledovskaia, S M

1975-08-01

The functional state of the thyroid gland and the concentration of thyroid hormones in the peripheral blood were studied in 20 mature female albino rats during their estral cycle. Evaluation of the thyroid functional state was made according to data of histological, morphological (the diameter of folliculi, the height of the thyroid epithelium) and histochemical analysis (determination of NAD and NADP-dehydrogenase, succinatedehydrogenase, lactate dehydrogenase, peroxydase, acid and alkaline phosphatase) as well as biochemical determination of iodine bound with protein (IBP) in the blood plasma and investigation of the ratio of the parameters in question under conditions of the sex cycle. The cyclic changes of the morphological state of the thyroid gland attended by the phases of the estral cycle were revealed. The activation of the organ was observed in proestrus and estrus which was evidenced by high levels of activity of the enzymes under study, high concentration of IBP in the blood and increased height of thyreocytes. A decreased function of the thyroid parenchyma was observed at the period of metaestrus-diestrus.

FINE STRUCTURAL LOCALIZATION OF ADENOSINETRIPHOSPHATASE ACTIVITY IN HEART MUSCLE MYOFIBRILS

PubMed Central

Tice, Lois W.; Barrnett, Russell J.

1962-01-01

Activity of myofibrillar adenosinetriphosphatase was demonstrated histochemically at a fine structural level in isolated, unfixed or hydroxyadipaldehyde-fixed cardiac myofibrils in the rat, using a lead precipitation technique and either Ca++ or Mg++ as activating ion. Activity in relaxed myofibrils was found in the A band, but not the H, I, or Z bands. Deposits of final product frequently exhibited an axial periodicity of near 365 A, and bore a close relationship to filaments within the A band. Several patterns of distribution occurred in contracted myofibrils. In myofibrils which had shortened to the point of disappearance of the I band, final product was distributed throughout the sarcomere, except for the unreactive Z band. A second type of distribution occurred in strongly contracted fibers in which there was intensification of activity in the center of the sarcomere. These findings are discussed in the light of the recent morphological evidence and it is suggested that the distribution of final product is consistent with localization of enzyme activity to the cross-bridges between the thick and thin filaments. PMID:13981351

Effect of an estrogen-deficient state and alendronate therapy on bone loss resulting from experimental periapical lesions in rats.

PubMed

Xiong, Haofei; Peng, Bin; Wei, Lili; Zhang, Xiaolei; Wang, Li

2007-11-01

The aim of the research was to evaluate the impact of an estrogen-deficient state and alendronate (ALD) therapy on bone loss resulting from experimental periapical lesions in rats. Periapical lesions were induced on ovariectomized (OVX) and sham-ovariectomized (Sham) rats. After sample preparation, histologic and radiographic examination for periapical bone loss area and an enzyme histochemical test for tartrate-resistant acid phosphatase (TRAP) were performed. The results showed that OVX significantly increased bone loss resulting from periradicular lesions. After daily subcutaneous injection of ALD, the bone loss area and the number of TRAP-positive cells (osteoclasts) were reduced. These findings suggested that alendronate may protect against increased bone loss from experimental periapical lesions in estrogen-deficient rats. Given recent recognition of adverse effects of bisphosphonates, including an increased risk for osteonecrosis, the findings from this study should not be interpreted as a new indication for ALD treatment. However, they may offer insight into understanding and predicting outcomes in female postmenopausal patients already on ALD therapy for medical indications.

The extensor digitorum brevis: histological and histochemical aspects

PubMed Central

Jennekens, F. G. I.; Tomlinson, B. E.; Walton, J. N.

1972-01-01

Samples of the extensor digitorum brevis muscle (EDB) obtained at necropsy from 26 subjects without known neuromuscular disease were examined histologically and histochemically. In the two youngest subjects, aged 2 months and 8 years, a mosaic distribution of type I and type II fibres was present. From the second decade onwards, increasing with age, the mosaic pattern was gradually replaced by groups of type I and type II fibres and areas of grouped fibre atrophy appeared. It is suggested that these findings may be explained by a slow process of denervation and reinnervation. This process does not seem to occur to the same extent in three other distal limb muscles from which specimens were also examined. Images PMID:4260286

Histochemical studies on protease formation in the cotyledons of germinating bean seeds.

PubMed

Yomo, H; Taylor, M P

1973-03-01

Protease formation in Phaseolus vulgaris L. cotyledons during seed germination was studied histochemically using a gelatin-film-substrate method. Protease activity can be detected by this method on the 5th day of germination, at approximately the same time that a rapid increase of activity was observed by a test-tube assay with casein as a substrate. At the early stage of germination, protease activity was observed throughout the cotyledon except in two or three cell layers below the cotyledon surface and in several cell layers around the vascular bundles. A highly active cell layer surrounding the protease-inactive cells near the vascular bundles is suggested to be a source of the protease.

The effect of calorie restriction on the presence of apoptotic ovarian cells in normal wild type mice and low-plasma-IGF-1 Laron dwarf mice

PubMed Central

2013-01-01

Background It is known that caloric restriction extends lifespan and can minimize age-related dysfunction of the reproductive system. We became interested in how caloric restriction influences apoptosis, which is a crucial process that maintains ovarian cell homeostasis. Methods We examined ovarian cells in: 2.5-year-old wild type mice on caloric restriction (CR) or fed ad libitum (AL) and Laron dwarf mice (GHR-KO) at the same ages on CR or fed AL. Apoptosis was assessed by histochemical analysis on paraffin sections of ovarian tissue. Results Morphological and histochemical analysis revealed that CR improved reproductive potential in 2.5-year-old WT littermates and GHR-KO female mice, as indicated by the increased number of ovarian follicles. The level of apoptosis in ovarian tissue was higher in WT mice on a CR diet compared with WT mice on the AL diet. In GHR-KO mice, the level of apoptosis in ovaries was similar for mice on CR and on AL diets and bigger than in WT mice on CR. Conclusions Morphological and histochemical analysis revealed a younger biological age of the ovaries in 2-year-old WT littermates and GHR-KO female mice on CR compared with animals fed AL. PMID:24063422

Mucinous (colloid) adenocarcinomas secrete distinct O-acylated forms of sialomucins: a histochemical study of gastric, colorectal and breast adenocarcinomas.

PubMed

Sáez, C; Japón, M A; Poveda, M A; Segura, D I

2001-12-01

Mucinous (colloid) adenocarcinomas represent a distinct group of tumours defined by the presence of large amounts of extracellular mucins. By using histochemical methods, we analysed mucins secreted by mucinous versus non-mucinous adenocarcinomas and looked for differential secretion profiles. Sixty-four adenocarcinomas were studied (23 colorectal, 17 gastric, and 24 breast tumours). Thirty-two tumours were of the colloid type. The following methods were applied to paraffin tissue sections: (i) Alcian blue (pH 2.5) and periodic acid-Schiff (PAS); (ii) high iron diamine and Alcian blue (pH 2.5); (iii) periodic acid borohydride, potassium hydroxide, and PAS; (iv) periodic acid-thionine Schiff, potassium hydroxide, and PAS; and (v) periodic acid-borohydride and PAS. Most adenocarcinomas secreted acidic mucins, with sialomucins predominating over sulfomucins, except for non-mucinous adenocarcinomas of the breast which showed predominant neutral mucins. All mucinous adenocarcinomas contained C9-O-acyl sialic acid as mono, di(C8,C9)-, or tri(C7,C8,C9)-O-acyl forms. Acidic mucins secreted by the majority of non-colloid adenocarcinomas consisted of non-O-acylated sialomucins. C9-O-acylation of sialic acid is a characteristic feature of mucinous adenocarcinomas and can be readily detected by histochemical methods.

On the future contents of a small journal of histochemistry.

PubMed

Pellicciari, C

2012-12-10

In the last three years, more than 70,000 scientific articles have been published in peer reviewed journals on the application of histochemistry in the biomedical field: most of them did not appear in strictly histochemical journals, but in others dealing with cell and molecular biology, medicine or biotechnology. This proves that histochemistry is still an active and innovative discipline with relevance in basic and applied biological research, but also demonstrates that especially the small histochemical Journals should likely reconsider their scopes and strategies to preserve their authorship. A review of the last three years volumes of the European Journal of Histochemistry, taken as an example of a long-time established small Journal, confirmed that the published articles were widely heterogeneous in their topics and experimental models, as in this Journal's tradition. This strongly suggests that a Journal of histochemistry should keep its role as a forum open to an audience as broad as possible, publishing papers on cell and tissue biology in a wide variety of models. This will improve knowledge of the basic mechanisms of development and differentiation, while helping to increase the number of potential authors since scientists who generally do not use histochemistry in their research will find hints for the applications of histochemical techniques to novel still unexplored subjects.

Extrafloral nectaries of four varieties of Chamaecrista ramosa (Vogel) H.S.Irwin & Barneby (Fabaceae): anatomy, chemical nature, mechanisms of nectar secretion, and elimination.

PubMed

da Silva Pereira, Priscila; de Almeida Gonçalves, Letícia; da Silva, Marcos José; Rezende, Maria Helena

2018-04-27

Considering the importance of extrafloral nectaries (EFNs) in Fabaceae, the objectives of this research were to analyze (1) the anatomical and histochemical characteristics of the EFNs of Chamaecrista ramosa var. ramosa, C. ramosa var. curvifoliola, C. ramosa var. parvifoliola, and C. ramosa var. lucida and (2) the ultrastructure of the EFNs of C. ramosa var. ramosa. Standard techniques in plant anatomy and transmission electron microscopy were used. The anatomical analyses confirmed the characteristics described for extrafloral nectaries, evidencing three well-defined regions: epidermis, nectariferous, and subnectariferous parenchymas. Carbohydrates, proteins, pectins/mucilages, and lipids were detected by histochemical analyzes in all varieties. The ultrastructure of the EFNs of C. ramosa var. ramosa allowed the observation of microchannels at the external periclinal cell walls of the epidermis covering the secretory region. The nectariferous and subnectariferous parenchyma cells have periplasmic spaces, large plastids containing starch grains and plastoglobules, mitochondria, developed endoplasmic reticulum, large vacuoles with electron-dense contents, and membrane residues may be associated with the vacuole, suggesting the occurrence of autophagic processes. The anatomical, histochemical, and ultrastructural patterns revealed characteristics that confirm the glands of C. ramosa as extrafloral nectaries and suggest the eccrine mechanism of secretion.

On the future contents of a small journal of histochemistry

PubMed Central

Pellicciari, C.

2012-01-01

In the last three years, more than 70,000 scientific articles have been published in peer reviewed journals on the application of histochemistry in the biomedical field: most of them did not appear in strictly histochemical journals, but in others dealing with cell and molecular biology, medicine or biotechnology. This proves that histochemistry is still an active and innovative discipline with relevance in basic and applied biological research, but also demonstrates that especially the small histochemical journals should likely reconsider their scopes and strategies to preserve their authorship. A review of the last three years volumes of the European Journal of Histochemistry, taken as an example of a long-time established small journal, confirmed that the published articles were widely heterogeneous in their topics and experimental models, as in this journal's tradition. This strongly suggests that a journal of histochemistry should keep its role as a forum open to an audience as broad as possible, publishing papers on cell and tissue biology in a wide variety of models. This will improve knowledge of the basic mechanisms of development and differentiation, while helping to increase the number of potential authors since scientists who generally do not use histochemistry in their research will find hints for the applications of histochemical techniques to novel still unexplored subjects. PMID:23361247

Pigmented Cells in the Pineal Gland of Female Viscacha (Lagostomus maximus maximus): A Histochemical and Ultrastructural Study

PubMed Central

Busolini, Fabricio Ivan; Rodríguez, Graciela Beatriz; Filippa, Verónica Palmira

2017-01-01

The presence of pigment has been demonstrated in different nervous structures such as those of retina, substantia nigra, and locus coeruleus. These pigments have also been described in the pineal gland of different mammal species. Histochemical and ultrastructural studies of the pineal gland of female viscacha (Lagostomus maximus maximus) were performed to analyze the presence of pigmented cells under natural conditions and to evaluate a probable relation between pigment content and glandular activity during pregnancy. The following techniques were applied: hematoxylin-eosin, phosphotungstic acid-hematoxylin, Masson-Fontana silver, DOPA histochemistry, Schmorl's reaction and toluidine blue. Estradiol and progesterone serum levels were determined by RIA. The ultrastructural features of the pineal pigment granules were also analyzed. Pigment granules were observed in a random distribution, but the pigmented cells were frequently found near blood vessels. The pineal pigment was histochemically identified as melanin. Differences in the amount of pigmented cells were found between pregnant and nonpregnant viscachas. The ultrastructural analysis revealed the presence of premelanosomes and melanosomes. Estradiol and progesterone levels vary during pregnancy. In conclusion, the changes in the amount of pigment content and hormone levels may indicate that the pineal gland of female viscacha is susceptible to endocrine variations during pregnancy. PMID:29391866

Immunohistochemical/histochemical double staining method in the study of the columnar metaplasia of the oesophagus.

PubMed

Cabibi, D; Giannone, A G; Mascarella, C; Guarnotta, C; Castiglia, M; Pantuso, G; Fiorentino, E

2014-03-05

Intestinal metaplasia in Barrett's oesophagus (BO) represents an important risk factor for oesophageal adenocarcinoma. Instead, few and controversial data are reported about the progression risk of columnar-lined oesophagus without intestinal metaplasia (CLO), posing an issue about its clinical management. The aim was to evaluate if some immunophenotypic changes were present in CLO independently of the presence of the goblet cells. We studied a series of oesophageal biopsies from patients with endoscopic finding of columnar metaplasia, by performing some immunohistochemical stainings (CK7, p53, AuroraA) combined with histochemistry (Alcian-blue and Alcian/PAS), with the aim of simultaneously assess the histochemical features in cells that shows an aberrant expression of such antigens. We evidenced a cytoplasmic expression of CK7 and a nuclear expression of Aurora A and p53,  both in goblet cells of BO and in non-goblet cells of CLO, some of which showing mild dysplasia. These findings suggest that some immunophenotypic changes are present in CLO and they can precede the appearance of the goblet cells or can be present independently of them, confirming the conception of BO as the condition characterized by any extention of columnar epithelium. This is the first study in which a combined immunohistochemical/histochemical method has been applied to Barrett pathology.

Effect of sex on histological and histochemical structures of different parts of the kidney in Japanese quail.

PubMed

Mobini, Behzad; Abdollahi, MohammadHossein

2016-09-01

The aim of the present study was to investigate the effect of gender on the histological and histochemical structures of different anatomical regions of the kidney in Japanese quail (Coturnix japonica). Tissue samples from cranial, middle and caudal divisions of each kidney were obtained from 20 male and 20 female adult, healthy Japanese quail. The sections stained with hematoxylin & eosin ( H & E: ), Masson's trichrome, Verhoeff's, Alcian blue (pH 2.5), Periodic acid-Schiff, and Gomori's method for reticulum. Unusual findings of the kidney in Japanese quail were the presence of three types of nephrons, all the connective tissue fibers in capsule and interlobular septa and AB-reactions of the proximal convoluted cells. No significant sex-based differences were found. The various histological structures of the kidney showed no significant differences among different divisions of the left and right kidneys. It is concluded that the general histological and histochemical properties of the kidney in Japanese quail were similar to those of chickens and some other species, but that there were also some differences. One of the major differences was brush border of interdigitating microvilli on luminal surface of collecting ducts in Japanese quail. © 2016 Poultry Science Association Inc.

Histochemical carbonic anhydrase in rat inner medullary collecting duct

NASA Technical Reports Server (NTRS)

Kleinman, J. G.; Bain, J. L.; Fritsche, C.; Riley, D. A.

1992-01-01

Rat inner medullary collecting duct (IMCD) secretes substantial amounts of H+. However, carbonic anhydrase (CA), a concomitant of H+ secretion, has been generally reported absent in this segment. To reexamine this problem, we investigated CA and the morphological phenotypes of cells comprising the IMCD by CA histochemistry, using a modified Hansson technique with light and electron microscopy. Throughout the medulla, tubule cells exhibit histochemical CA activity. In the initial third of the inner medulla, a small proportion have features of intercalated cells and demonstrate some degree of CA activity. However, the majority population in the early portions of the IMCD appears to consist of principal cells. These also show CA staining of widely variable intensity, both among and within cells. A third cell type, previously called "IMCD cells", appears in the middle portion of the IMCD and is the only cell type present near the papilla tip. In contrast to previous reports, these "IMCD cells" have histochemical CA staining, also of highly variable intensity. These results demonstrate that stainable carbonic anhydrase to support acidification is present throughout the rat IMCD, both in intercalated cells and in some cells clearly not of this type. Therefore, the presence of CA is not specific for the intercalated cell type and suggests that other cell types may participate in acid secretion in IMCD.

Pollutant-induced cell death and reactive oxygen species accumulation in the aerial roots of Chinese banyan (Ficus microcarpa)

NASA Astrophysics Data System (ADS)

Liu, Nan; Cao, Ce; Sun, Zhongyu; Lin, Zhifang; Deng, Rufang

2016-11-01

Industrial pollutants induce the production of toxic reactive oxygen species (ROS) such as O2.-, H2O2, and ·OH in plants, but they have not been well quantified or localized in tissues and cells. This study evaluated the pollutant- (HSO3-, NH4NO3, Al3+, Zn2+, and Fe2+) induced toxic effects of ROS on the aerial roots of Chinese banyan (Ficus microcarpa). Root cell viability was greatly reduced by treatment with 20 mM NaHSO3, 20 mM NH4NO3, 0.2 mM AlCl3, 0.2 mM ZnSO4, or 0.2 mM FeSO4. Biochemical assay and histochemical localization showed that O2.- accumulated in roots in response to pollutants, except that the staining of O2.- under NaHSO3 treatment was not detective. Cytochemical localization further indicated that the generated O2.- was present mainly in the root cortex, and pith cells, especially in NH4NO3- and FeSO4-treated roots. The pollutants also caused greatly accumulated H2O2 and ·OH in aerial roots, which finally resulted in lipid peroxidation as indicated by increased malondialdehyde contents. We conclude that the F. microcarpa aerial roots are sensitive to pollutant-induced ROS and that the histochemical localization of O2.- via nitrotetrazolium blue chloride staining is not effective for detecting the effects of HSO3- treatment because of the treatment’s bleaching effect.

Loud Noise Exposure Produces DNA, Neurotransmitter and Morphological Damage within Specific Brain Areas.

PubMed

Frenzilli, Giada; Ryskalin, Larisa; Ferrucci, Michela; Cantafora, Emanuela; Chelazzi, Silvia; Giorgi, Filippo S; Lenzi, Paola; Scarcelli, Vittoria; Frati, Alessandro; Biagioni, Francesca; Gambardella, Stefano; Falleni, Alessandra; Fornai, Francesco

2017-01-01

Exposure to loud noise is a major environmental threat to public health. Loud noise exposure, apart from affecting the inner ear, is deleterious for cardiovascular, endocrine and nervous systems and it is associated with neuropsychiatric disorders. In this study we investigated DNA, neurotransmitters and immune-histochemical alterations induced by exposure to loud noise in three major brain areas (cerebellum, hippocampus, striatum) of Wistar rats. Rats were exposed to loud noise (100 dBA) for 12 h. The effects of noise on DNA integrity in all three brain areas were evaluated by using Comet assay. In parallel studies, brain monoamine levels and morphology of nigrostriatal pathways, hippocampus and cerebellum were analyzed at different time intervals (24 h and 7 days) after noise exposure. Loud noise produced a sudden increase in DNA damage in all the brain areas under investigation. Monoamine levels detected at 7 days following exposure were differently affected depending on the specific brain area. Namely, striatal but not hippocampal dopamine (DA) significantly decreased, whereas hippocampal and cerebellar noradrenaline (NA) was significantly reduced. This is in line with pathological findings within striatum and hippocampus consisting of a decrease in striatal tyrosine hydroxylase (TH) combined with increased Bax and glial fibrillary acidic protein (GFAP). Loud noise exposure lasting 12 h causes immediate DNA, and long-lasting neurotransmitter and immune-histochemical alterations within specific brain areas of the rat. These alterations may suggest an anatomical and functional link to explain the neurobiology of diseases which prevail in human subjects exposed to environmental noise.

A novel histochemical method for the visualization of thrombin activity in the nervous system.

PubMed

Bushi, D; Gera, O; Kostenich, G; Shavit-Stein, E; Weiss, R; Chapman, J; Tanne, D

2016-04-21

Although thrombin has an important role in both central and peripheral nerve diseases, characterization of the anatomical distribution of its proteolytic activity has been limited by available methods. This study presents the development, challenges, validation and implementation of a novel histochemical method for visualization of thrombin activity in the nervous system. The method is based on the cleavage of the substrate, Boc-Asp(OBzl)-Pro-Arg-4MβNA by thrombin to liberate free 4-methoxy-2-naphthylamine (4MβNA). In the presence of 5-nitrosalicylaldehyde, free 4MβNA is captured, yielding an insoluble yellow fluorescent precipitate which marks the site of thrombin activity. The sensitivity of the method was determined in vitro using known concentrations of thrombin while the specificity was verified using a highly specific thrombin inhibitor. Using this method we determined the spatial distribution of thrombin activity in mouse brain following transient middle cerebral artery occlusion (tMCAo) and in mouse sciatic nerve following crush injury. Fluorescence microscopy revealed well-defined thrombin activity localized to the right ischemic hemisphere in cortical areas and in the striatum compared to negligible thrombin activity contralaterally. The histochemical localization of thrombin activity following tMCAo was in good correlation with the infarct areas per triphenyltetrazolium chloride staining and to thrombin activity measured biochemically in tissue punches (85 ± 35 and 20 ± 3 mU/ml, in the cortical and striatum areas respectively, compared to 7 ± 2 and 13 ± 2 mU/ml, in the corresponding contralateral areas; mean ± SEM; p<0.05). In addition, 24 h following crush injury, focal areas of highly elevated thrombin activity were detected in teased sciatic fibers. This observation was supported by the biochemical assay and western blot technique. The histochemical method developed in this study can serve as an important tool for studying the role of thrombin in physiological and pathological conditions. Copyright © 2016 IBRO. Published by Elsevier Ltd. All rights reserved.

Fibrinogen storage disease in a Chinese boy with de novo fibrinogen Aguadilla mutation: Incomplete response to carbamazepine and ursodeoxycholic acid.

PubMed

Zhang, Mei-Hong; Knisely, A S; Wang, Neng-Li; Gong, Jing-Yu; Wang, Jian-She

2016-08-12

Fibrinogen storage disease (FSD) is a rare autosomal-dominant disorder caused by mutation in FGG, encoding the fibrinogen gamma chain. Here we report the first Han Chinese patient with FSD, caused by de novo fibrinogen Aguadilla mutation, and his response to pharmacologic management. Epistaxis and persistent clinical-biochemistry test-result abnormalities prompted liver biopsy in a boy, with molecular study of FGG in him and his parents. He was treated with the autophagy enhancer carbamazepine, reportedly effective in FSD, and with ursodeoxycholic acid thereafter. Inclusion bodies in hepatocellular cytoplasm stained immune-histochemically for fibrinogen. Selective analysis of FGG found the heterozygous mutation c.1201C > T (p.Arg401Trp), absent in both parents. Over more than one year's follow-up, transaminase and gamma-glutamyl transpeptidase activities have lessened but not normalized. This report expands the epidemiology of FSD and demonstrates idiosyncrasy in response to oral carbamazepine and/or ursodeoxycholic acid in FSD.

Refractory Cushing's disease caused by multinodular ACTH-cell hyperplasia.

PubMed

McKeever, P E; Koppelman, M C; Metcalf, D; Quindlen, E; Kornblith, P L; Strott, C A; Howard, R; Smith, B H

1982-09-01

A patient with pituitary-dependent hypercortisolism, unresponsive to resection of nodules in the anterior lobe, is described. Histochemical stains of the nodules showed multiple, focal, cellular expansions of the fibrovascular stroma. Transitions between normal and expanded adenohypophysial acini were present. Immunoperoxidase stains for ACTH and other pituitary hormones revealed that these multiple foci contained an excess of ACTH-positive cells. Less than 10% of the cells in these foci were negative for ACTH and positive for other hormones. Serial sections showed that these foci of predominantly ACTH-producing acini were not connected. Clinical, morphological, and immunohistochemical data indicated that ACTH-cell hyperplasia caused Crushing's disease in this patient. Pathologic study of individual cases should concentrate on determining whether hyperplasia or adenoma exist at the time of surgical exploration of the pituitary gland, since this determination is important to proper treatment. Tentative criteria to recognize ACTH-cell hyperplasia are: 1. Multiple foci of ACTH laden cells. 2. A minor subpopulation of cells of alternate hormone series. 3. Expansion without destruction of acini in the adenohypophysis.

[Metastasis revealing malignant peritoneum mesothelioma: About the difficulty to identify the primary tumors].

PubMed

Bretagne, Charles-Henri; Petitjean, Alain; Felix, Sophie; Bedgedjian, Isabelle; Algros, Marie-Paule; Delabrousse, Eric; Valmary-Degano, Séverine

2016-04-01

Peritoneal malignant mesothelioma is a rare and extremely aggressive tumor that is sometimes difficult to diagnose. We report two cases of metastatic malignant peritoneal mesothelioma. In one case, malignant metastatic cells were identified in cervical lymph nodes while in the other case, the cells were found in the liver. In both cases, metastases were identified before discovering the primary tumor. This led to the misdiagnosis of carcinoma of unknown origin. Nevertheless, the histological and immuno-histochemical patterns were typical of malignant mesothelioma. Regarding metastasis of unknown origin, a differentiation of epithelioid peritoneal malignant mesothelioma and adenocarcinoma proved to be difficult. Therefore, we discuss the diagnostic usefulness of immuno-histochemical mesothelioma markers. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

Nasal and paranasal adenocarcinomas with neuroendocrine differentiation in dogs.

PubMed

Ninomiya, F; Suzuki, S; Tanaka, H; Hayashi, S; Ozaki, K; Narama, I

2008-03-01

Tumors of the nasal cavity or paranasal sinuses of 18 dogs were examined histopathologically, immunohistochemically, and histochemically. The tumors were classified histologically as 13 adenocarcinomas, 3 transitional carcinomas, 1 squamous cell carcinoma, and 1 adenosquamous carcinoma. Tumor cells were strongly immunoreactive for broad-spectrum cytokeratins in all cases, for cytokeratin 8/18 in 16 cases, and for cytokeratin 19 in 17 cases. None of the 18 carcinomas had cytologic or histologic features indicative of neuroendocrine differentiation, yet tumor cells in 5 of the 13 adenocarcinomas were argyrophilic and immunohistochemically positive for synaptophysin and chromogranin A. Results of this study indicate that neuroendocrine markers may be detected immunohistochemically and histochemically in canine nasal or paranasal adenocarcinomas despite the lack of typical histologic features of neuroendocrine differentiation.

Zygomatic gland adenoma in a dog: histochemical and immunohistochemical evaluation.

PubMed

Giudice, Chiara; Marco, Rondena; Mirko, Radice; Luca, Mertel; Giorgio, Cammarata

2005-01-01

Orbital epithelial tumors in dogs are rare and most frequently malignant. Distinguishing their origin from the lacrimal or zygomatic gland is often challenging and is based mostly on tumor location. A case of adenoma involving the orbit in a 13-year-old, female, standard Schnauzer is reported. Histologically, the neoplasm was characterized by nests and cords of epithelial cells mostly forming small glandular structures. The origin of the tumor from the zygomatic gland was determined by histochemical characteristics (alcian blue pH 1 positive staining) of a small remnant of normal gland included within the tumor capsule. The benign nature of our finding was confirmed by follow-up information: 2 years after complete surgical removal of the mass no tumor recurrence or metastases was recorded.

Roles of zinc and metallothionein-3 in oxidative stress-induced lysosomal dysfunction, cell death, and autophagy in neurons and astrocytes.

PubMed

Lee, Sook-Jeong; Koh, Jae-Young

2010-10-26

Zinc dyshomeostasis has been recognized as an important mechanism for cell death in acute brain injury. An increase in the level of free or histochemically reactive zinc in astrocytes and neurons is considered one of the major causes of death of these cells in ischemia and trauma. Although zinc dyshomeostasis can lead to cell death via diverse routes, the major pathway appears to involve oxidative stress.Recently, we found that a rise of zinc in autophagic vacuoles, including autolysosomes, is a prerequisite for lysosomal membrane permeabilization and cell death in cultured brain cells exposed to oxidative stress conditions. The source of zinc in this process is likely redox-sensitive zinc-binding proteins such as metallothioneins, which release zinc under oxidative conditions. Of the metallothioneins, metallothionein-3 is especially enriched in the central nervous system, but its physiologic role in this tissue is not well established. Like other metallothioneins, metallothionein-3 may function as metal detoxicant, but is also known to inhibit neurite outgrowth and, sometimes, promote neuronal death, likely by serving as a source of toxic zinc release. In addition, metallothionein-3 regulates lysosomal functions. In the absence of metallothionein-3, there are changes in lysosome-associated membrane protein-1 and -2, and reductions in certain lysosomal enzymes that result in decreased autophagic flux. This may have dual effects on cell survival. In acute oxidative injury, zinc dyshomeostasis and lysosomal membrane permeabilization are diminished in metallothionein-3 null cells, resulting in less cell death. But over the longer term, diminished lysosomal function may lead to the accumulation of abnormal proteins and cause cytotoxicity.The roles of zinc and metallothionein-3 in autophagy and/or lysosomal function have just begun to be investigated. In light of evidence that autophagy and lysosomes may play significant roles in the pathogenesis of various neurological diseases, further insight into the contribution of zinc dynamics and metallothionein-3 function may help provide ways to effectively regulate these processes in brain cells.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Weeks, J.M.; Svendsen, C.

A simple subcellular histochemical staining technique employing the lysosomal probe neutral red has been developed for use with the epiendogeic earthworm Lumbricus rubellus Hoffmeister. Coelomocytes extracted from the coelomic cavity of earthworms into an isotonic earthworm Ringer solution were allowed to adhere to a microscope slide for 30 s before the application of a neutral red dye. This red dye was rapidly accumulated within the lysosomes. Observation of the loss of this dye from these lysosomes into the surrounding cytosol has enabled the quantification of the degree of lysosomal damage caused to earthworms with exposure to an increasing range ofmore » soil copper concentrations, in both laboratory and mesocosm studies. This simple in vitro biomarker has potential for the rapid assessment of the toxic effects to earthworms from soils contaminated with heavy metals and metalloids.« less

Salt tolerance and activity of antioxidative enzymes of transgenic finger millet overexpressing a vacuolar H(+)-pyrophosphatase gene (SbVPPase) from Sorghum bicolor.

PubMed

Anjaneyulu, Ediga; Reddy, Palle Surender; Sunita, Merla Srilakshmi; Kishor, Polavarapu B Kavi; Meriga, Balaji

2014-06-15

A vacuolar proton pyrophosphatase cDNA clone was isolated from Sorghum bicolor (SbVPPase) using end-to-end gene-specific primer amplification. It showed 80-90% homology at the nucleotide and 85-95% homology at the amino acid level with other VPPases. The gene was introduced into expression vector pCAMBIA1301 under the control of the cauliflower mosaic virus 35S (CaMV35S) promoter and transformed into Agrobacterium tumifaciens strain LBA4404 to infect embryogenic calli of finger millet (Eleusine coracana). Successful transfer of SbVPPase was confirmed by a GUS histochemical assay and PCR analysis. Both, controls and transgenic plants were subjected to 100 and 200mM NaCl and certain biochemical and physiological parameters were studied. Relative water content (RWC), plant height, leaf expansion, finger length and width and grain weight were severely reduced (50-70%), and the flowering period was delayed by 20% in control plants compared to transgenic plants under salinity stress. With increasing salt stress, the proline and chlorophyll contents as well as the enzyme activities of superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX), guaiacol peroxidase (GPX) and glutathione reductase (GR) increased by 25-100% in transgenics, while malondialdehyde (MDA) showed a 2-4-fold decrease. The increased activities of antioxidant enzymes and the reduction in the MDA content suggest efficient scavenging of reactive oxygen species (ROS) in transgenics and, as a consequence, probably alleviation of salt stress. Also, the leaf tissues of the transgenics accumulated 1.5-2.5-fold higher Na(+) and 0.4-0.8-fold higher K(+) levels. Together, these results clearly demonstrate that overexpression of SbVPPase in transgenic finger millet enhances the plant's performance under salt stress. Copyright © 2014 Elsevier GmbH. All rights reserved.

Interaction between organophosphate compounds and cholinergic functions during development.

PubMed

Aluigi, M G; Angelini, C; Falugi, C; Fossa, R; Genever, P; Gallus, L; Layer, P G; Prestipino, G; Rakonczay, Z; Sgro, M; Thielecke, H; Trombino, S

2005-12-15

Organophosphate (OP) compounds exert inhibition on cholinesterase (ChE) activity by irreversibly binding to the catalytic site of the enzymes. For this reason, they are employed as insecticides for agricultural, gardening and indoor pest control. The biological function of the ChE enzymes is well known and has been studied since the beginning of the XXth century; in particular, acetylcholinesterase (AChE, E.C. 3.1.1.7) is an enzyme playing a key role in the modulation of neuromuscular impulse transmission. However, in the past decades, there has been increasing interest concerning its role in regulating non-neuromuscular cell-to-cell interactions mediated by electrical events, such as intracellular ion concentration changes, as the ones occurring during gamete interaction and embryonic development. An understanding of the mechanisms of the cholinergic regulation of these events can help us foresee the possible impact on environmental and human health, including gamete efficiency and possible teratogenic effects on different models, and help elucidate the extent to which OP exposure may affect human health. The chosen organophosphates were the ones mainly used in Europe: diazinon, chlorpyriphos, malathion, and phentoate, all of them belonging to the thionophosphate chemical class. This research has focused on the comparison between the effects of exposure on the developing embryos at different stages, identifying biomarkers and determining potential risk factors for sensitive subpopulations. The effects of OP oxonisation were not taken into account at this level, because embryonic responses were directly correlated to the changes of AChE activity, as determined by histochemical localisation and biochemical measurements. The identified biomarkers of effect for in vitro experiments were: cell proliferation/apoptosis as well as cell differentiation. For in vivo experiments, the endpoints were: developmental speed, size and shape of pre-gastrula embryos; developmental anomalies on neural tube, head, eye, heart. In all these events, we had evidence that the effects are mediated by ion channel activation, through the activation/inactivation of acetylcholine receptors (AChRs).

Histochemical and immunohistochemical characterization of chordoma in ferrets.

PubMed

Yui, Takeshi; Ohmachi, Tetsuo; Matsuda, Kazuya; Okamoto, Minoru; Taniyama, Hiroyuki

2015-04-01

Chordomas of the tip of the tail in 6 ferrets were examined using histopathological, histochemical and immunohistochemical procedures. Histopathologically, round neoplastic cells containing numerous cytoplasmic vacuoles of varying sizes, categorized as "physaliphorous cells", were observed in the amorphous eosinophilic or pale basophilic myxoid stroma. Physaliphorous cells were arranged in lobules and in a "chordoid" or "cobblestone" manner. The neoplasms were diagnosed as benign chordoma without local invasion and metastasis. Histochemically, the cytoplasm of small neoplastic cells was positive for periodic acid-Schiff stain and alcian blue (AB) pH 2.5 and pH 1.0 stains, but negative for hyaluronidase digestion-AB pH 2.5 stain. All neoplastic cells were strongly stained with colloidal ion, negative for high iron diamine AB pH 2.5 and toluidine blue pH 2.5 stains, and positive for Mayer's mucicarmine stain. Immunohistochemistry using antibodies directed against low-molecular-weight cytokeratins (CK18, CK19 and CK20), vimentin and mucin core protein (MUC5AC) revealed that neoplastic cells had both epithelial and mesenchymal elements. The expression of low-molecular-weight cytokeratins suggests that neoplastic cells acquired the properties of glandular epithelial cells and produced epithelial mucus. Furthermore, the expression of cytokeratins, vimentin, S100 protein, brachyury and epithelial membrane antigen indicates that the neoplasms were equivalent to the classic type of human chordoma. Therefore, immunohistochemistry using these antibodies can be useful for the characterization of ferret chordoma.

Indications for laser therapy in diverse models of periodontitis

NASA Astrophysics Data System (ADS)

Kunin, Anatoly A.; Erina, Stanislava V.; Sokolova, Irina A.; Pankova, Svetlana N.; Ippolitov, Yu. A.; Lepechina, L. I.; Malinovskaya, L. A.; Chitrina, L. L.

1996-11-01

Parodontal diseases have an immunological pathogenic mechanism leading to various manifestations and can not be referred to as a common inflammation. The home and foreign research points at active and immunological reaction with the following distraction surrounding tissues of the tooth. Histochemical and biochemical examinations show metabolic disturbances of parodontal tissues. A total sample size of 604 people suffering from average height of chronic generalized parodontitis was examined in the survey. Immunological and histochemical tests were taken before and after a course of laser therapy with the use of helium-neon laser 'YAGODA', an inhibitory and stimulating dosage irradiations and anti-inflammatory dosage irradiations with infrared laser 'UZOR'. We selected a group of patients with the decreased local immunological status on the ground of immunological tests. Histochemical tests shaped the next group with the passive and active forms of parodontitis pathology. The tests data resulted in a method of laser therapy. The investigations confirm that the chronic generalized parodontitis has a shift in tissue immunity of the oral cavity and cell-bound metabolic disturbance of gum epithelium. It is expedient to use the anti-inflammatory dosage irradiations with infrared laser 'UZOR' to correct immunity, and in case of and active process to realize the DNA and RNA synthesis by means of increasing the irradiation with the apparatus 'YAGODA'. The irradiation decreases in case of a passive process.

[The structure of vegetative organs, and saponins histochemical localization and content comparization in Polygala sibirica L].

PubMed

Teng, Hong Mei; Fang, Min Feng; Hu, Zheng Hai

2009-02-01

Anatomical, histochemical and phytochemistry methods were used to investigate the structure of vegetative organs, and saponins localization and dynamic changes in Polygala sibirica L. The root consisted of developed periderm and secondary vascular. The secondary phloem was thick, and mainly composed of parenchyma. There were well-developed vessels and fibers in the secondary xylem. The stem was composed of epidermis, cortex and vascular bundle. The ring of sclerenchymatous cells lied between cortex and phloem might be the apoplastic protective screen which could protect the stem from drought. The leaf was bifacial one. The root and stem possessed characteristics adapting to arid environment. Histochemical localization results showed that saponins distributed in secondary phloem and phelloderm of root, in epidermis, cortex and phloem of stem, mainly in mesophyll of leaf. It displayed that saponins accumulated mainly in parenchyma cells of vegetative organs, among of which, the secondary phloem was the main storage site. The HPLC results also showed that the saponins accumulated in all the vegetative organs of Polygala sibirica L., with higher content in roots and lower content in the aerial part that included stems and leaves. The study indicated the aerial part of Polygala sibirica L. also had medicinal value. The saponins content had dynamic variance at the developmental stage, the crude drug should be gathered at period from April to May.

Enzyme

MedlinePlus

Enzymes are complex proteins that cause a specific chemical change in all parts of the body. For ... use them. Blood clotting is another example of enzymes at work. Enzymes are needed for all body ...

The beginning of Space Life Science in China exploration rockets for biological experiment during 1960's

NASA Astrophysics Data System (ADS)

Jiang, Peidong; Zhang, Jingxue

The first step of space biological experiment in China was a set of five exploration rockets launched during 1964 to 1966, by Shanghai Institute of Machine and Electricity, and Institute of Biophysics of The Chinese Academy of Sciences. Three T-7AS1rockets for rats, mice and other samples in a biological cabin were launched and recovered safely in July of 1964 and June of 1965. Two T-7AS2rockets for dog, rats, mice and other samples in a biological cabin were launched and recovered safely in July of 1966. Institute of Biophysics in charged of the general design of biological experiments, telemetry of physiological parameters, and selection and training of experiment animals. The samples on-board were: rats, mice, dogs, and test tubes with fruit fly, enzyme, bacteria, E. Coli., lysozyme, bacteriaphage, RNAase, DNAase, crystals of enzyme, etc. Physiological, biochemical, bacte-riological, immunological, genetic, histochemical studies had been conducted, in cellular and sub cellular level. The postures of rat and dog were monitored during flight and under weight-lessness. Physiological parameters of ECG, blood pressure, respiration rate, body temperature were recorded. A dog named"Xiao Bao"was flight in 1966 with video monitor, life support system and conditioned reflex equipment. It flighted for more than 20 minutes and about 70km high. After 40 years, the experimental data recorded of its four physiological parameters during the flight process was reviewed. The change of 4 parameters during various phase of total flight process were compared, analyzed and discussed.

Distribution of cholinesterases in insects*

PubMed Central

Booth, G. M.; Lee, An-Horng

1971-01-01

The study of toxicology and other related fields has been largely based on in vitro techniques. These methods have provided quantitative information on the effects of inhibitors on enzymes, but none on the localized effects of inhibitors on selected sites of action within the animal. Histochemical study of frozen sections does provide data on the site of action of toxicants. The utility of histochemistry in conjunction with in vitro methods is discussed. The substrates acetylthiocholine and phenyl thioacetate were utilized in demonstrating cholinesterase. Neither substrate penetrated well into freshly dissected nerve cord preparations, but both compounds were hydrolysed by sectioned tissue. The leaving group of phenyl thioacetate was demonstrated to be benzenethiol. In general, acetylthiocholine was hydrolysed slightly more rapidly by insect cholinesterases. A unique cholinesterase was found in motor end-plates of cricket muscle, which hydrolyses acetylthiocholine and which was inhibited by physostigmine. No other insect muscle preparation showed this activity. Topical application of insecticides showed that a vital site of action in flies is the peripheral area of the thoracic ganglia and that in crickets the brain and nerve cord are involved at knock-down. Kinetic data indicate that acetylthiocholine has a greater affinity than does phenyl thioacetate for a variety of enzyme sources. Ultrastructural evidence shows that cholinesterases that hydrolyse acetylthiocholine are membrane-bound. Phenyl thioacetate was found to be useful as a model in designing new insecticides. ImagesFig. 5Fig. 6Fig. 7Fig. 8Fig. 13Fig. 14Fig. 15Fig. 16Fig. 9Fig. 10Fig. 11Fig. 12Fig. 1Fig. 2Fig. 3Fig. 4Fig. 17Fig. 18Fig. 19 PMID:5315359

AgNOR histochemical expression in benign prostatic hyperplasia and prostatic adenocarcinoma

NASA Astrophysics Data System (ADS)

Rita, R.; Delyuzar; Laksmi, L. I.

2018-03-01

Benign prostatic hyperplasia and prostatic adenocarcinoma were common diseases and usually occurred after the 5th decade of life. The problem in diagnosing using Hematoxylin and Eosin staining was how to differentiate whether it is benign or malignant zone. Therefore, proliferating markers, such as AgNOR, could be helping to over this difficulty. A descriptive study using consecutive sampling as the method of sample recruiting was conducted to describe AgNOR histochemical expression in benign prostatic hyperplasia and prostatic adenocarcinoma. AgNOR staining was done in 13 benign prostatic hyperplasia samples and 7 prostatic adenocarcinoma samples, which have been confirmed using p63 immunohistochemical staining before. Benign prostatic hyperplasia usually showed lower AgNOR proliferating activity while all of theprostatic adenocarcinoma (100%) had high AgNOR proliferating activity.

A lectin histochemical study on carbohydrate moieties of the gonadotropin-like substance in the epithelial cells of Hatschek's pit of Branchiostoma belcheri

NASA Astrophysics Data System (ADS)

Fang, Y. Q.; Welsch, U.

1997-03-01

The present light microscopic lectin, histochemical study suggests for the first time that the vertebrate gonadotropin-like substance in the basal part of the epithelial cells of Hatschek's pit is a sialic acid-containing glycoprotein. The binding intensity of the epithelial cells in Hatschek's pit to 6 lectins ( Limulus polyphemus agglutinin (LPA), Wheat germ agglutinin (WGA), Helix pomatia agglutinin (HPA), Concanavalin A (Con A), Ulex europaeus agglutinin I (UEA I) and Ricinus communis agglutinin I (RCA I)) indicate that the carbohydrate composition of the gonadotrophic glycoprotein is similar to that of mammals and fish, and that N-acetyl-D-galactosamine, sialic acid, glucosamine, D-mannose and L-fucose are components of the carbohydrate portion.

Certification procedures for nuclear fast red (Kernechtrot), CI 60760.

PubMed

Frank, M; Dapson, Rw; Wickersham, Tw; Kiernan, Ja

2007-02-01

Nuclear fast red (CI 60760), also known as Kernechtrot, is commonly used in conjunction with an excess of aluminum ions as a red nuclear counterstain following histochemical procedures that yield blue products. The dye has also been used as a histochemical and colorimetric reagent for calcium. Unsatisfactory samples of nuclear fast red are encountered occasionally, and confusion has resulted from applying the name of the dye to neutral red (CI 50040), an unrelated compound with different properties. Tests for the identity and performance of nuclear fast red have been developed in the laboratory of the Biological Stain Commission. The Commission will now accept samples submitted by vendors for certification. We describe here the spectrophotometric, chromatographic and biological staining methods that are used to identify and test nuclear fast red.

Staining Methods for Normal and Regenerative Myelin in the Nervous System.

PubMed

Carriel, Víctor; Campos, Antonio; Alaminos, Miguel; Raimondo, Stefania; Geuna, Stefano

2017-01-01

Histochemical techniques enable the specific identification of myelin by light microscopy. Here we describe three histochemical methods for the staining of myelin suitable for formalin-fixed and paraffin-embedded materials. The first method is conventional luxol fast blue (LFB) method which stains myelin in blue and Nissl bodies and mast cells in purple. The second method is a LBF-based method called MCOLL, which specifically stains the myelin as well the collagen fibers and cells, giving an integrated overview of the histology and myelin content of the tissue. Finally, we describe the osmium tetroxide method, which consist in the osmication of previously fixed tissues. Osmication is performed prior the embedding of tissues in paraffin giving a permanent positive reaction for myelin as well as other lipids present in the tissue.

Effect of inhibition of microsomal Ca(2+)-ATPase on cytoplasmic calcium and enzyme secretion in pancreatic acini.

PubMed

Metz, D C; Pradhan, T K; Mrozinski, J E; Jensen, R T; Turner, R J; Patto, R J; Gardner, J D

1994-01-13

We used thapsigargin (TG), 2,5-di-tert-butyl-1,4-benzohydroquinone (BHQ) and cyclopiazonic acid (CPA), each of which inhibits microsomal Ca(2+)-ATPase, to evaluate the effects of this inhibition on cytoplasmic free calcium ([Ca2+]i) and secretagogue-stimulated enzyme secretion in rat pancreatic acini. Using single-cell microspectrofluorimetry of fura-2-loaded acini we found that all three agents caused a sustained increase in [Ca2+]i by mobilizing calcium from inositol-(1,4,5)-trisphosphate-sensitive intracellular calcium stores and by promoting influx of extracellular calcium. Concentrations of all three agents that increased [Ca2+]i potentiated the stimulation of enzyme secretion caused by secretagogues that activate adenylate cyclase but inhibited the stimulation of enzyme secretion caused by secretagogues that activate phospholipase C. With BHQ, potentiation of adenylate cyclase-mediated enzyme secretion occurred immediately whereas inhibition of phospholipase C-mediated enzyme secretion occurred only after several min of incubation. In addition, the effects of BHQ and CPA on both [Ca2+]i and secretagogue-stimulated enzyme secretion were reversed completely by washing whereas the actions of TG could not be reversed by washing. Concentrations of BHQ in excess of those that caused maximal changes in [Ca2+]i inhibited all modes of stimulated enzyme secretion by a mechanism that was apparently unrelated to changes in [Ca2+]i. Finally, in contrast to the findings with TG and BHQ, CPA inhibited bombesin-stimulated enzyme secretion over a range of concentrations that was at least 10-fold lower than the range of concentrations over which CPA potentiated VIP-stimulated enzyme secretion.

Concentration effects of the UV filter oxybenzone in Cyperus alternifolius: assessment of tolerance by stress-related response.

PubMed

Chen, Feiran; Schnick, Sandrine; Schröder, Peter

2018-06-01

Phytoremediation has been proposed to reduce the load of the sunscreen oxybenzone (OBZ) in the aquatic environment. Despite the proven removal efficiency of this compound, little is known about its influence, particularly oxidative stress on plants. In this study, a short-term incubation of macrophytic Cyperus alternifolius was performed to prove the plant's ability to withstand the stress. Detached shoots were immersed in medium spiked with different concentrations of OBZ (50, 100, and 500 μM) for 2, 4, and 7 days, respectively. Increased formation of O 2 - and H 2 O 2 in Cyperus treated with OBZ was characterized by intense colorization following histochemical staining. Alterations of enzyme activities involved in the antioxidative defense system indicate an adaptive response of C. alternifolius to this xenobiotic stress. Quantification of lipid peroxidation reveals that no significant membrane damage occurred during incubation with OBZ. Overall, 50 μM OBZ (tenfold higher than the amount frequently detected in the environment) exhibited low toxic effects. Accordingly, this pilot study provides information on the potential use of Cyperus to remove emerging sunscreen contaminants from water bodies.

Myalgia as the revealing symptom of multicore disease and fibre type disproportion myopathy

PubMed Central

Sobreira*, C; Marques, W; Barreira, A

2003-01-01

Objective: To report the occurrence of myalgia as the revealing symptom of multicore disease and fibre type disproportion myopathy. Methods: The clinical cases of three patients with fibre type disproportion myopathy and one with multicore disease are described. Skeletal muscle biopsies were processed for routine histological and histochemical studies. Results: The clinical picture was unusual in that the symptoms were of late onset and the predominant complaint was muscle pain exacerbated by exercise. Muscle weakness was found in only a single patient, the mother of a patient with fibre type disproportion myopathy. Physical examination was unremarkable in the other patients. Muscle biopsies from patients 1 and 2 contained type I fibres that were considerably smaller than the type II fibres, supporting the diagnosis of fibre type disproportion myopathy. Skeletal muscle of patient 4 showed multiple areas, predominantly but not exclusively in the type I fibres, from which oxidative enzyme activities were absent, as seen in multicore disease. Conclusions: Muscle pain was the main clinical manifestation in our patients. Recognition of the broader clinical expression of these myopathies is important for prognostic reasons and for genetic counselling of the family members. PMID:12933945

Adenosine triphosphatase activity of cutaneous nerve fibers.

PubMed

Idé, C; Saito, T

1980-02-01

The histochemical study of Mg++-activated adenosine triphosphatase (Mg++-ATPase) activity was carried out on the peripheral nerves of mouse digital skin by light and electron microscopy. Under the light microscope, the ATPase activity was clearly demonstrated on the nerve fibers as a fine network in the subepidermal regions. Under the electron microscope, the reaction product of enzyme activity was located in the interspace between axolemma and the surrounding Schwann cells of the unmyelinated nerve fibers. No reaction product was observed in the space between the axolemma and the Schwann cells associated with myelinated nerve fibers. Demonstrable activity was absent at the nodes of Ranvier as well as on the para- and internodal regions of these myelinated axons. The part of the axolemma lacking a Schwann cell sheath failed to show a reaction product. The perineural epithelial cells surrounding the nerve fibers displayed reaction product in the caveolae. These results suggest a functional difference in the axon-Schwann interface of myelinated as compared to unmyelinated nerve fibers. The function of the perineural epithelial cell would be expected to be a regulatory one in transferring materials across the epithelium to keep the proper humoral environment around nerve fibers.

MELAS syndrome in a patient with a point mutation in MTTS1.

PubMed

Lindberg, C; Moslemi, A-R; Oldfors, A

2008-02-01

BACKGROUND, OBJECTIVE AND METHODS: We describe a female patient with a mitochondrial encephalopathy, lactic acidosis and stroke-like episodes syndrome. As a child, she developed epilepsy and stroke-like episodes giving cognitive impairment and ataxia but no hearing impairment. At the age of 44 years, she suffered a cerebral sinus thrombosis which was warfarin treated. One month later, she developed an episode of severe acidosis associated with encephalopathy and myelopathy. She was found to harbour a 7512T>C mutation in the mitochondrial encoded tRNA(Ser(UCN)) gene (MTTS1). The mutation load was 91% in muscle and 24% in blood. Enzyme histochemical analysis of the muscle tissue showed numerous cytochrome c oxidase (COX)-negative fibres. Restriction fragment length polymorphism (RFLP) analysis of single muscle fibres showed significantly higher level (median 97%, range: 94-99%) of the mutation in the COX-negative fibres compared with COX-positive fibres (median 36%, range: 12-91%), demonstrating the pathogenic effect of the mutation. Different levels of heteroplasmy (range 34-61%) were detected in hair shafts analysed by RFLP. This case adds to the spectrum of clinical presentations, i.e. sinus thrombosis, in patients having MTTS1 mutations.

Functional Utrastructure of Genlisea (Lentibulariaceae) Digestive Hairs

PubMed Central

Płachno, Bartosz Jan; Kozieradzka-Kiszkurno, Małgorzata; Świątek, Piotr

2007-01-01

Background and Aims Digestive structures of carnivorous plants produce external digestive enzymes, and play the main role in absorption. In Lentibulariaceae, the ultrastructure of digestive hairs has been examined in some detail in Pinguicula and Utricularia, but the sessile digestive hairs of Genlisea have received very little attention so far. The aim of this study was to fill this gap by expanding their morphological, anatomical and histochemical characterization. Methods Several imaging techniques were used, including light, confocal and electron microscopy, to reveal the structure and function of the secretory hairs of Genlisea traps. This report demonstrates the application of cryo-SEM for fast imaging of whole, physically fixed plant secretory structures. Key Results and Conclusion The concentration of digestive hairs along vascular bundles in subgenus Genlisea is a primitive feature, indicating its basal position within the genus. Digestive hairs of Genlisea consist of three compartments with different ultrastructure and function. In subgenus Tayloria the terminal hair cells are transfer cells, but not in species of subgenus Genlisea. A digestive pool of viscous fluid occurs in Genlisea traps. In spite of their similar architecture, the digestive-absorptive hairs of Lentibulariaceae feature differences in morphology and ultrastructure. PMID:17550910

The distribution of cholinesterases in the cat carotid body.

PubMed

Biscoe, T J; Silver, A

1966-03-01

1. The distribution of acetyl- and butyrylcholinesterase in the carotid body of the cat has been examined histochemically. Studies were made on normal carotid bodies and on carotid bodies from cats in which certain nerves had been cut some time previously. The nerves sectioned were the sinus nerve, the post-ganglionic sympathetic branch of the superior cervical ganglion or the preganglionic cervical sympathetic trunk.2. It was confirmed that more butyrylcholinesterase than acetylcholinesterase is present. Both enzymes are found in three sites: (i) as strands, (ii) as plexuses, (iii) inside a few cells.3. The distribution is unaffected by cutting the sinus nerve or preganglionic cervical sympathetic nerves. Disorganization and depletion of the cholinesterases in the strands and plexuses occurs when the post-ganglionic branch of the superior cervical ganglion is cut. The cholinesterase in cells is unaffected.4. In carotid bodies in which vessels were filled with red blood cells or in which the vascular bed was injected with carmine-gelatine, it was seen that strands and plexuses are associated with blood vessels, and with blood vessels and cells respectively.5. It is suggested that a cholinergic pathway controlling carotid body blood vessels runs in the post-ganglionic cervical sympathetic.

Direct visualization of clay microfabric signatures driving organic matter preservation in fine-grained sediment

NASA Astrophysics Data System (ADS)

Curry, Kenneth J.; Bennett, Richard H.; Mayer, Lawrence M.; Curry, Ann; Abril, Maritza; Biesiot, Patricia M.; Hulbert, Matthew H.

2007-04-01

We employed direct visualization of organic matter (OM) sequestered by microfabric signatures in organo-clay systems to study mechanisms of OM protection. We studied polysaccharides, an abundant class of OM in marine sediments, associated with the nano- and microfabric of clay sediment using a novel application of transmission electron microscopy, histochemical staining (periodic acid-thiosemicarbazide-silver proteinate), and enzymatic digestion techniques. We used two experimental organo-clay sediment environments. First, laboratory-consolidated sediment with 10% chitin (w/w) added was probed for chitin before and after digestion with chitinase. Second, fecal pellets from the polychaete Heteromastus filiformis were used as a natural environment rich in clay and polysaccharides. Sections of this material were probed with silver proteinate for polysaccharides before and after digestion with a mixture of enzymes (amylase, cellulase, chitinase, dextranase, and pectinase). In both environments, chitin or other polysaccharides were found within pores, bridging clay domains, and attached to clay surfaces in undigested samples. Digested samples showed chitin or polysaccharides more closely associated with clay surfaces and in small pores. Our results imply protective roles for both sorption to clay surfaces and encapsulation within clay microfabric signatures.

[Comparative study of diagnostic methods for visceral leishmaniasis in dogs from Ilha Solteira, SP].

PubMed

de Assis, Juliana; de Queiroz, Nina Marí Gual Pimenta; da Silveira, Rita de Cássia Vieira; Nunes, Cáris Maroni; Oliveira, Trícia Maria Ferreira de Sousa; Junior, Antonio Carlos Faconti de Noronha; Neves, Maria Francisca; Machado, Rosangela Zacarias; Buzetti, Wilma Aparecida Starke

2010-01-01

The purpose of the present work was a comparative study of diagnostic methods for Canine Visceral Leishmaniasis (CVL) using serological methods, enzyme-linked immunosorbent assay (ELISA) and indirect fluorescent antibody test (IFAT), histochemical (HE) and immunohistochemical (IMHC) tests using spleen, lymph node and liver canine tissues. In addition, Polymerase Chain Reaction (PCR) was done in blood and in tissues in order to compare and confirm no conclusive and negative diagnosis by the methods above. For this study, 34 dogs were divided according to clinical signs in asymptomatic, oligosymptomatic and polisymptomatic Leishmania-infected dogs euthanized by Zoonotic Disease Control Center (CCZ) from Ilha Solteira, SP, Brazil. The positivism indexes of ELISA, IMHC, IFAT and HE were 65.0, 62.0, 56.0 and 56.0%, respectively with the highest numbers of positive dogs in polisymptomatic (92.0%) followed by oligosymptomatic (57.0%) and asymptomatic dogs (12.5%). Furthermore, PCR confirmed the positive results and detected DNA in tissues from 100% of negative dogs and 89.0% suspects raising the animal positivism index up to 97.0%. In conclusion, PCR was the most sensitive and a valuable method for a definitive CVL diagnosis.

The blood-brain barrier of the chick glycogen body (corpus gelatinosum) and its functional implications.

PubMed

Möller, Wilhelm; Kummer, Wolfgang

2003-07-01

Among recent vertebrates only birds possess a glycogen body (corpus gelatinosum), located in the rhomboidal sinus of the lumbosacral region of the spinal cord and separated from the neural tissue proper. Because of the specific topographical situation of this circumventricular organ, the structure of its vascular system is of special interest with respect to the still unsolved functional problems. The existence of a blood-brain barrier is demonstrated by the exclusion of intravascularly injected tracer (horseradish peroxidase), and immunocytochemical demonstration of glucose transporter-1 as a functional marker and of neurothelin, occludin and ZO-1 as structural markers. Alkaline phosphatase and gamma-glutamyltransferase activities, two enzyme reactions frequently used for demonstration of an established blood-brain barrier in vitro, were localized histochemically on the plasmalemma of glycogen body cells and were absent from the endothelium. In addition, local enlargements of the intercellular space were observed by transmission and scanning electron microscopy. In accordance with the concept of a third circulation the cerebrospinal fluid may be the vehicle for distributing substances originating in the glycogen body to the CNS, while the vascular endothelium maintains the internal milieu by virtue of its dynamic barrier functions.

Conversion of rat muscle fiber types. A time course study.

PubMed

Oakley, C R; Gollnick, P D

1985-01-01

Rats were used in this study to determine the time course of conversion of muscle fiber types. The right or left gastrocnemius muscle was removed thereby causing an overload on the ipsilateral soleus and plantaris muscles. The contralateral limb served as a control. The type II to type I fiber conversion was followed histochemically in the soleus and plantaris muscles for one to six weeks following surgery. Muscle sections were stained for myofibrillar actomyosin ATPase and NADH tetrazolium reductase. The type I population in the soleus muscle was 99.3% six weeks after synergist removal. The plantaris muscle underwent a two fold increase in the percentage of type I fibers after six weeks. Transitional fibers were prominent in the plantaris muscle and reached their peak at 4% (P less than 0.05) of the total population, four weeks after surgery.

The study of the relationship between aberrant expression of hot shock protein 70 (HSP70) and spontaneous abortion.

PubMed

Peng, Y-B; Liu, H; Huang, S-H; Lai, H; Zhou, Q; Luo, Y; Zhang, Z-Y; Xi, B-R; Ouyang, X

2017-02-01

The present study is aimed to explore the relationship between aberrant expression of heat shock protein 70 (HSP) and spontaneous abortion. 50 patients with spontaneous abortion and 50 patients with induced abortion were continuously selected based on the nearest matching principle, and the proportion of age and gestational age was 1:1. The decidual tissues were obtained, and the cell apoptosis was determined by TUNEL assay. Further, the expression of HSP70 was assayed by immune-histochemical staining, and the expression of HSP70 mRNA was detected by the RT-PCR approach. Apoptosis rate, HSP70 expression and HSP70 mRNA expression in the observation group were significantly higher than the control group. HSP70 might induce apoptosis so as to cause spontaneous abortion.

Respiratory disease caused by synthetic fibres: a new occupational disease.

PubMed Central

Pimentel, J C; Avila, R; Lourenço, A G

1975-01-01

Seven patients exposed to the inhalation of synthetic fibres presented with various bronchopulmonary diseases, such as asthma, extrinsic allergic alveolitis, chronic bronchitis with bronchiectasis, spontaneous pneumothorax, and chronic pneumonia. The histological features are described and an attempt has been made to set up immunological techniques for the diagnosis. A series of histochemical techniques, based on textile chemistry, are proposed for the identification of the inclusions found in bronchopulmonary lesions. The results of the experimental production of the disease in guinea-pigs by the inhalation of synthetic fibre dusts are presented. The prognosis of these cases is good in the acute or recently established cases but is poor when widespread and irreversible fibrosis has set in. The authors consider that pulmonary disease due to inhaled particles is probably set off by an individual factor, possibly immunological. Images PMID:1179318

Aging and unusual catecholamine-containing structures in the mouse brain.

PubMed

Masuoka, D T; Jonsson, G; Finch, C E

1979-06-22

Brains of C57BL/6J mice, aged 4, 8 and 20--29 months, were examined by the Falck-Hillarp histochemical fluorescence technique. Numerous large, intensely fluorescent green to yellow-green spots (LIFS) were observed in the brains of senescent mice. LIFS were generally round to ovoid in shape and ranged in size from about 10 micrometer to about 30 micrometer. Histochemical and pharmacological procedures and spectral analysis indicated that the formaldehyde-induced fluorescence of the LIFS was due to the presence of catecholamines (CA) rather than aging pigment. Their distribution in the brain suggests an association with nerve axons or terminals rather than cell bodies. The number of LIFS in the hypothalamus increased progressively during aging. It is proposed that LIFS may represent age-related, unusual CA accumulation in enlargements proximal to axonal or terminal portions undergoing spontaneous degeneration.

Microwave-Assisted Tissue Preparation for Rapid Fixation, Decalcification, Antigen Retrieval, Cryosectioning, and Immunostaining

PubMed Central

2016-01-01

Microwave irradiation of tissue during fixation and subsequent histochemical staining procedures significantly reduces the time required for incubation in fixation and staining solutions. Minimizing the incubation time in fixative reduces disruption of tissue morphology, and reducing the incubation time in staining solution or antibody solution decreases nonspecific labeling. Reduction of incubation time in staining solution also decreases the level of background noise. Microwave-assisted tissue preparation is applicable for tissue fixation, decalcification of bone tissues, treatment of adipose tissues, antigen retrieval, and other special staining of tissues. Microwave-assisted tissue fixation and staining are useful tools for histological analyses. This review describes the protocols using microwave irradiation for several essential procedures in histochemical studies, and these techniques are applicable to other protocols for tissue fixation and immunostaining in the field of cell biology. PMID:27840640

[Histological and histochemical characteristics of pancreas of deer at the Altay].

PubMed

Riadinskaia, N I; Siraziev, R Z

2008-01-01

Season changes in the pancreas from animals belonging to genuine deer subfamily have been investigated by histological, histochemical and biometric methods. Glycogen is not found in the pancreas cells throughout the seasons pointing to high functional activity of glandular cells, since glycogen is consumed for carbohydrate biopolymer synthesis and not accumulated. Depending upon the season, cytoplasm of pancreacells, cells of excretory ducts and pancreas islets showed different intensity of pyroninophilous reaction indicating RNA presence. These data coupled with the presence of protein in these cells demonstrate protein-synthesizing ability of the gland adapted to biorhythm. Changes in quantity and types of web cells as well as in functional activity of pancrea cells and pancreas islets revealed season regularity and reflected functional lability of the cells and their constant involvement in many of vital important process.

Zinc treatment of the digestive gland of the slug Arion after L. 2. Sublethal effects at the histological level

DOE Office of Scientific and Technical Information (OSTI.GOV)

Recio, A.; Marigomez, J.A.; Angulo, E.

1988-12-01

Several authors have pointed out the necessity when measuring the level of pollution within a biological system on an assessment of the biological effects of pollutants. Within this field, histological parameters for the indication of environmentally induced stress response (i.e., destabilization of the lysosomal systems, reduction of the mean epithelia thickness of digestive gland cells-MET) could offer an initial approach to determine the impact of a pollutant on the biota. The present work is the completion of the histochemical study on cellular distribution of zinc in the digestive gland of the slug Arion ater. Thus, the histochemical and planimetric studymore » of the effects of zinc-exposure on A. ater is presented as the other aspect in which histology is concerned in pollution research.« less

Yaba Tumor Poxvirus Synthesis In Vitro I. Cytopathological, Histochemical, and Immunofluorescent Studies

PubMed Central

Yohn, David S.; Haendiges, Victoria A.; Grace, James T.

1966-01-01

Yohn, David S. (Roswell Park Memorial Institute, Buffalo, N.Y.), Victoria A. Haendiges, and James T. Grace, Jr. Yaba tumor poxvirus synthesis in vitro. I. Cytopathological, histochemical, and immunofluorescent studies. J. Bacteriol. 91:1977–1985. 1966.—Yaba tumor poxvirus synthesis in BSC-1 cell culture was followed sequentially with light microscopy, immunofluorescent microscopy, and various histochemical stains. The first evidence of infection was detected at 24 hr when nucleoli became hypertrophic, reflecting enhanced ribonucleic acid (RNA) synthesis. At 36 hr, deoxyribonucleic acid synthesis was detected in the cytoplasm. This was immediately followed by or associated with antigen synthesis at paranuclear sites and enhanced RNA synthesis in the cytoplasm. Cytoplasmic inclusions were readily apparent at 4 days in initially infected cells. Contiguous spread of virus was judged to have occurred around the third day of infection. The time required to complete the synthetic cycle from time of infection to production of infectious progeny was estimated to be of the order of 60 hr. This cycle is 6 to 10 times longer than for vaccinia virus. By light microscopy, cytopathic effects (CPE) were detectable in 5 days in heavily infected cultures. With 100 units or less of infectious virus, CPE was not readily detected until the 10th to 14th day. At this time, focal areas of infection classified either as microtumors or microplaques were present. Secondary foci appeared during the 4th week of incubation. Images PMID:4287080

Anatomical and phenological implications of the relationship between Schinus polygama (Cav.) (Cabrera) and the galling insect Calophya rubra (Blanchard).

PubMed

Guedes, L M; Aguilera, N; Ferreira, B G; Becerra, J; Hernández, V; Isaias, R M S

2018-05-01

The success of galling insects could be determined by synchronisation with host plant phenology and climate conditions, ensuring suitable oviposition sites for gall induction and food resources for their survival. The anatomical, histochemical and phenological synchronisation strategies between Calophya rubra (Blanchard) (Hemiptera: Psylloidea) and its host, the evergreen plant Schinus polygama (Cav.) (Cabrera) (Anacardiaceae), in the Mediterranean climate of southern Chile was evaluated and compared to that of the congeneric C. cf. duvauae (Scott) from Brazil and closely related host plant S. engleri in a subtropical climate. Anatomical, histometric, histochemical and vegetative phenology studies of the stem and galls were conducted from June 2015 to December 2016. Based on the anatomical, histometric and histochemical analysis, the conical stem gall traits imply gains over the non-galled stem toward the galling insect survival, but the maintenance of phellem, secretory ducts and pith indicate conservative developmental traits that cannot be manipulated by C. rubra. Our results indicate that the conditions of the Mediterranean climate zone limit C. rubra immature activity during unfavourable periods, probably determining a diapause period and a univoltine life cycle, which are peculiarities of the S. polygama- C. rubra system. The synchronisation between development and seasonality confers peculiarities to the S. polygama- C. rubra system in the Mediterranean climate zone. © 2018 German Society for Plant Sciences and The Royal Botanical Society of the Netherlands.

Histochemical detection of lead and zinc in plant tissues

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tung, G.; Temple, P.J.

1975-01-01

Histochemical studies on uptake and localization of lead and zinc in plant tissues were carried out. A histochemical stain technique was developed to differentiate zinc from lead. Lead was detected in plant tissues by soaking fresh plant materials in freshly prepared sodium rhodizonate stain (0.2% Na rhodizonate acidified to pH3 with glacial acetic acid). Samples were evacuated 5 min and soaked for 30 min before embedding in the congealed stain, then sectioned with a cryostat and examined under a light microscope. Lead particles in plant tissues were stained scarlet-red. Gelatinous, proteinaceous or saccharic embedding materials normally used to prepare plantmore » sampled for sectioning in the cryostat interfered with the color reaction. Sectioning plant samples without staining whole tissues resulted in a weakened response to the stain. Color of stained sample materials were retained for several months if stored in a frozen condition. This technique was used to detect lead both inside and on the surface of plant samples collected in the vicinity of highway and industrial lead sources and to trace the pathways of lead uptake from the air or from contaminated soils. A sodium rhodizonate technique was also developed to be specific for zinc in plant tissues. Plant samples were soaked in a neutral Na-rhodizonate in phosphate buffer at pH 7.5 for observation. The color of zinc developed to produce a purplish or reddish-brown color.« less

The Catalytic Function of Enzymes.

ERIC Educational Resources Information Center

Splittgerber, Allan G.

1985-01-01

Discusses: structure of the enzyme molecule; active site; reaction mechanism; transition state; factors affecting enzyme reaction rates, concentration of enzyme; concentration of substrate; product concentration; temperature effects and pH effects; factors causing a lowering of activation energy; proximity and orientation effects; substrate strain…

Studying modification of aminoglycoside antibiotics by resistance-causing enzymes via microarray.

PubMed

Disney, Matthew D

2012-01-01

Widespread bacterial resistance to antibiotics is a significant public health concern. To remain a step ahead of evolving bacteria, new methods to study resistance to antibacterials and to uncover novel antibiotics that evade resistance are urgently needed. Herein, microarray-based methods that have been developed to study aminoglycoside modification by resistance-causing enzymes are reviewed. These arrays can also be used to study the binding of aminoglycoside antibiotics to a mimic of their therapeutic target, the rRNA aminoacyl site (A-site), and how modification by resistance-causing enzymes affects their abilities to bind RNA.

Differential Phytotoxic Impact of Plant Mediated Silver Nanoparticles (AgNPs) and Silver Nitrate (AgNO3) on Brassica sp.

PubMed Central

Vishwakarma, Kanchan; Shweta; Upadhyay, Neha; Singh, Jaspreet; Liu, Shiliang; Singh, Vijay P.; Prasad, Sheo M.; Chauhan, Devendra K.; Tripathi, Durgesh K.; Sharma, Shivesh

2017-01-01

Continuous formation and utilization of nanoparticles (NPs) have resulted into significant discharge of nanosized particles into the environment. NPs find applications in numerous products and agriculture sector, and gaining importance in recent years. In the present study, silver nanoparticles (AgNPs) were biosynthesized from silver nitrate (AgNO3) by green synthesis approach using Aloe vera extract. Mustard (Brassica sp.) seedlings were grown hydroponically and toxicity of both AgNP and AgNO3 (as ionic Ag+) was assessed at various concentrations (1 and 3 mM) by analyzing shoot and root length, fresh mass, protein content, photosynthetic pigments and performance, cell viability, oxidative damage, DNA degradation and enzyme activities. The results revealed that both AgNPs and AgNO3 declined growth of Brassica seedlings due to enhanced accumulation of AgNPs and AgNO3 that subsequently caused severe inhibition in photosynthesis. Further, the results showed that both AgNPs and AgNO3 induced oxidative stress as indicated by histochemical staining of superoxide radical and hydrogen peroxide that was manifested in terms of DNA degradation and cell death. Activities of antioxidants, i.e., ascorbate peroxidase (APX) and catalase (CAT) were inhibited by AgNPs and AgNO3. Interestingly, damaging impact of AgNPs was lesser than AgNO3 on Brassica seedlings which was due to lesser accumulation of AgNPs and better activities of APX and CAT, which resulted in lesser oxidative stress, DNA degradation and cell death. The results of the present study showed differential impact of AgNPs and AgNO3 on Brassica seedlings, their mode of action, and reasons for their differential impact. The results of the present study could be implied in toxicological research for designing strategies to reduce adverse impact of AgNPs and AgNO3 on crop plants. PMID:29075270

The glycan keratan sulfate in inner ear crystals

NASA Technical Reports Server (NTRS)

Fermin, C. D.; Martin, D. S.; Li, Y. T.; Li, S. C.

1995-01-01

The otoconial matrix (OM) of chicks (Gallus domesticus) inner ear was analyzed. Histochemically the OM was reacted with phosphotungstic acid (PTA) and immunohistochemically with the monoclonal antibody antikeratan sulfate (antiKS). The OM was digested with the enzyme endo-beta-galactosidase (E beta Galase) or separated by 1D and 2D gel electrophoresis. PTA which reacts with glycoproteins precipitated the OM, suggesting that the OM contains glycoproteins. A central core in each crystal had no PTA staining, suggesting that the core lacked glycoproteins. Anti KS antibody stained the OM with increased density in older embryos as determined by color thresholding. E beta Galase, which cleaves the lactosamine repeating units in KS, decreased the immunostain by 30% in the OM and by 20% in the cartilage. The OM from the utricle, saccule and macula lagena contained similar molecular weight bands. Five dense bands in the OM were less dense in tissue and blood controls, suggesting that such bands are enriched in the OM. Isoelectric focusing of the OM showed a negatively charged high molecular weight smear not present in blood and faint in tissue controls. The high affinity of the OM for the cationic PTA stain, the strong immunohistochemical reaction of the OM with anti KS antibody and high molecular weight negative smear in 2D gels taken together suggest that: a) the OM contains large amounts of glycoproteins and glycans, one of which is keratan sulfate, because its immuno stain with antiKS antibody was decreased by the enzyme E beta Galase, b) the utricle, saccule and macula lagena may have similar composition, and c) the concentration of KS may increase gradually until complete mineralization of the OM is reached.

PAP and NT5E inhibit nociceptive neurotransmission by rapidly hydrolyzing nucleotides to adenosine

PubMed Central

2011-01-01

Background Prostatic acid phosphatase (PAP) and ecto-5'-nucleotidase (NT5E, CD73) produce extracellular adenosine from the nucleotide AMP in spinal nociceptive (pain-sensing) circuits; however, it is currently unknown if these are the main ectonucleotidases that generate adenosine or how rapidly they generate adenosine. Results We found that AMP hydrolysis, when measured histochemically, was nearly abolished in dorsal root ganglia (DRG) neurons and lamina II of spinal cord from Pap/Nt5e double knockout (dKO) mice. Likewise, the antinociceptive effects of AMP, when combined with nucleoside transport inhibitors (dipyridamole or 5-iodotubericidin), were reduced by 80-100% in dKO mice. In addition, we used fast scan cyclic voltammetry (FSCV) to measure adenosine production at subsecond resolution within lamina II. Adenosine was maximally produced within seconds from AMP in wild-type (WT) mice but production was reduced >50% in dKO mice, indicating PAP and NT5E rapidly generate adenosine in lamina II. Unexpectedly, we also detected spontaneous low frequency adenosine transients in lamina II with FSCV. Adenosine transients were of short duration (<2 s) and were reduced (>60%) in frequency in Pap-/-, Nt5e-/- and dKO mice, suggesting these ectonucleotidases rapidly hydrolyze endogenously released nucleotides to adenosine. Field potential recordings in lamina II and behavioral studies indicate that adenosine made by these enzymes acts through the adenosine A1 receptor to inhibit excitatory neurotransmission and nociception. Conclusions Collectively, our experiments indicate that PAP and NT5E are the main ectonucleotidases that generate adenosine in nociceptive circuits and indicate these enzymes transform pulsatile or sustained nucleotide release into an inhibitory adenosinergic signal. PMID:22011440

Effect of repeated fasting/refeeding on obesity development and health complications in rats arising from reduced nest.

PubMed

Mozeš, Štefan; Šefčíková, Zuzana; Raček, Ľubomír

2015-02-01

Overnutrition during postnatal life represents a risk factor for later obesity and associated metabolic disorders. We investigated the interaction between postnatal and later-life nutrition on body composition, blood pressure and the jejunal enzyme activities in male Sprague-Dawley rats. From birth, we adjusted the number of pups in the nest to 4 (small litters-SL; overfeeding) or to 10 pups (normal litters-NL; controls), and from day 50 until 70, the SL (SL-R) and NL (NL-R) rats were subjected to 1 day fasting and 1 day refeeding cycles (RFR). Their body composition was determined by magnetic resonance imaging, and enzyme activity was assayed histochemically. At 50 and 70 days, SL rats were found to be overweight (p < 0.001), with higher adiposity (p < 0.001) and blood pressure (p < 0.01). Moreover, despite significantly decreased daily food intake during RFR (SL-R 39 %, NL-R 23 %), higher fat deposition (p < 0.001) and blood pressure (p < 0.05) was detected in SL-R rats. Activity of alkaline phosphatase (AP) functionally involved in lipid absorption was significantly higher in SL than NL rats (p < 0.001) but substantially decreased in RFR groups (SL-R p < 0.001, NL-R p < 0.01). However, despite these enzymatic adaptations to reduced food intake, the SL-R rats displayed significantly higher AP activity in comparison with NL-R rats (p < 0.01) on day 70. Our results demonstrate that postnatal overfeeding predisposes the ontogeny of intestinal function, which may promote the probability of obesity risk. Accordingly, in these animals, efficient fat deposition and elevated blood pressure were not diminished in response to dietary restrictions in later life.

Clinical expression of haemochromatosis in Irish C282Y homozygotes identified through family screening.

PubMed

Gleeson, F; Ryan, E; Barrett, S; Crowe, J

2004-09-01

In Ireland, the homozygote frequency of the C282Y mutation in the HFE gene is 1/83. The biochemical expression of this mutation is high in haemochromatosis (HH) individuals identified through family screening, but the clinical expression of the mutation in Irish HH subjects to date has not been investigated fully. To determine the clinical, biochemical and histological penetrance of the C282Y mutation in Irish C282Y homozygotes identified through family screening. Two hundred and nine C282Y homozygous individuals comprising of 172 first-degree relatives, 31 second-degree relatives and four unrelated individuals were identified following HFE mutation analysis of 167 families. The following variables were analysed: age at identification, gender, fasting transferrin saturation, fasting serum ferritin, liver enzymes, clinical symptomatology, liver histopathology and histochemical iron staining. An elevated transferrin saturation in combination with an elevated ferritin was present in 43.4% of males and 23.3% of females. Abnormal liver enzymes were found in 32.3% of males. Diabetes, a haemochromatosis-specific association, was noted in 2.8% of males. Of those individuals requiring liver histopathology evaluation, 38% had moderate-to-severe iron staining, and 42% had fibrosis; 2.8% of the biopsied cohort had cirrhosis. Thus, HH cirrhotics were identified in less than 1% of the screened population. Although the homozygote frequency in Ireland is very high, the prevalence of advanced liver disease was less than 1% of the family members screened. Nevertheless, 42% of biopsied patients had histological evidence of iron overload-related architectural change and 2.8% had cirrhosis. This cohort of young people had previously unrecognized biochemical iron overload and histopathological change. This emphasizes the importance and value of both genetic and biochemical screening in first-degree relatives of identified homozygotes.

Assessment of RNAi-induced silencing in banana (Musa spp.).

PubMed

Dang, Tuong Vi T; Windelinckx, Saskia; Henry, Isabelle M; De Coninck, Barbara; Cammue, Bruno P A; Swennen, Rony; Remy, Serge

2014-09-18

In plants, RNA- based gene silencing mediated by small RNAs functions at the transcriptional or post-transcriptional level to negatively regulate target genes, repetitive sequences, viral RNAs and/or transposon elements. Post-transcriptional gene silencing (PTGS) or the RNA interference (RNAi) approach has been achieved in a wide range of plant species for inhibiting the expression of target genes by generating double-stranded RNA (dsRNA). However, to our knowledge, successful RNAi-application to knock-down endogenous genes has not been reported in the important staple food crop banana. Using embryogenic cell suspension (ECS) transformed with ß-glucuronidase (GUS) as a model system, we assessed silencing of gusAINT using three intron-spliced hairpin RNA (ihpRNA) constructs containing gusAINT sequences of 299-nt, 26-nt and 19-nt, respectively. Their silencing potential was analysed in 2 different experimental set-ups. In the first, Agrobacterium-mediated co-transformation of banana ECS with a gusAINT containing vector and an ihpRNA construct resulted in a significantly reduced GUS enzyme activity 6-8 days after co-cultivation with either the 299-nt and 19-nt ihpRNA vectors. In the second approach, these ihpRNA constructs were transferred to stable GUS-expressing ECS and their silencing potential was evaluated in the regenerated in vitro plants. In comparison to control plants, transgenic plants transformed with the 299-nt gusAINT targeting sequence showed a 4.5 fold down-regulated gusA mRNA expression level, while GUS enzyme activity was reduced by 9 fold. Histochemical staining of plant tissues confirmed these findings. Northern blotting used to detect the expression of siRNA in the 299-nt ihpRNA vector transgenic in vitro plants revealed a negative relationship between siRNA expression and GUS enzyme activity. In contrast, no reduction in GUS activity or GUS mRNA expression occurred in the regenerated lines transformed with either of the two gusAINT oligo target sequences (26-nt and 19-nt). RNAi-induced silencing was achieved in banana, both at transient and stable level, resulting in significant reduction of gene expression and enzyme activity. The success of silencing was dependent on the targeted region of the target gene. The successful generation of transgenic ECS for second transformation with (an)other construct(s) can be of value for functional genomics research in banana.

Microscopic and histochemical manifestations of hyaline cartilage dynamics.

PubMed

Malinin, G I; Malinin, T I

1999-01-01

Structure and function of hyaline cartilages has been the focus of many correlative studies for over a hundred years. Much of what is known regarding dynamics and function of cartilage constituents has been derived or inferred from biochemical and electron microscopic investigations. Here we show that in conjunction with ultrastructural, and high-magnification transmission light and polarization microscopy, the well-developed histochemical methods are indispensable for the analysis of cartilage dynamics. Microscopically demonstrable aspects of cartilage dynamics include, but are not limited to, formation of the intracellular liquid crystals, phase transitions of the extracellular matrix and tubular connections between chondrocytes. The role of the interchondrocytic liquid crystals is considered in terms of the tensegrity hypothesis and non-apoptotic cell death. Phase transitions of the extracellular matrix are discussed in terms of self-alignment of chondrons, matrix guidance pathways and cartilage growth in the absence of mitosis. The possible role of nonenzymatic glycation reactions in cartilage dynamics is also reviewed.

Primary adenocarcinomas of the human urinary bladder: histochemical, immunological and ultrastructural studies.

PubMed

Alroy, J; Roganovic, D; Banner, B F; Jacobs, J B; Merk, F B; Ucci, A A; Kwan, P W; Coon, J S; Miller, A W

1981-01-01

Neoplastic and non-neoplastic tissue specimens from ten patients with primary adenocarcinoma of the urinary bladder were examined. Most of these tumors were associated with either foci of transitional cell carcinoma and/or with glandular metaplasia of the bladder epithelium. The mucin produced by the neoplastic cells was PAS, alcian blue, mucicarmine, PB/KOH/PAS, and RPB/KOH/PAS-positive. ABH isoantigens of these tumors were not always deleted. Ultrastructurally, the neoplastic cells resembled goblet cells. Their plasma membrane had numerous microvilli with prominent glycocalyx. Proliferation and attenuation of tight junctions were noted. The gap junctions were few and small. Two types of desmosomes were found. The ultrastructural features of the neoplastic cells were attributed in part to the malignant transformation and in part to the direction of their differentiation. We have not observed any distinctive morphologic, histochemical, immunologic or ultrastructural features that might be diagnostic for these adenocarcinomas.

[The expression and significance of IgE in anaphylactic shock guinea-pigs].

PubMed

Gong, Zhi-qiang; Xiao, Feng; Feng, Qiong; Xu, Xiao-ming; Zheng, Jian

2006-02-01

To seek the pathomorphological targets for forensic expertise in anaphylactic shock. The expression of IgE in hearts, lungs, livers, spleens, kidneys, gastrics, intestinals, tracheas and tonsils of anaphylactic shock guinea-pigs was observed at 0, 6, 12 h and 24 h respectively by tissue chip S-P immuno-histochemical method. Positive expression of IgE presented in lungs and tracheas in the test group with the peak at 0 hour and it declined as time advanced, and also there were significant differences at different times (P<0.05). The immuno-histochemical method of detecting the expression of IgE in lungs, tracheas and spleens can be supposed to be the pathomorphological targets for forensic expertise in anaphylactic shock. The weakening of the positive expression of IgE in lungs and tracheas as the time advanced suggested that in this kind of case the autopsy should be arried out as early as possible.

Nuclei of plants as a sink for flavanols.

PubMed

Feucht, W; Polster, J

2001-01-01

Onion cepa (L.) and Tsuga canadensis (L.) Carr. were investigated histochemically on the association of flavanols to nuclei. The young roots of Onion cepa are totally devoid of flavanol structures. Therefore, the excised roots tips were directly incubated into different solutions of flavanols. After 3 h of incubation a flavanol binding on the nuclei was recognizable, as seen by a yellowish-brown tanning reaction. Still to ensure the presence of flavanols on the nuclei, subsequent staining with the p-dimethylaminocinnamaldehyde reagent (DMACA) resulted in an intense blue colouration. Tsuga canadensis has significant amounts of vacuolar flavanol deposits in all parts of the tree as indicated by the DMACA reagent. It is obvious that also the nuclei were associated strongly with flavanols which can be demonstrated particularly elegant in the cells of the seed wings by histochemical methods. However, the mode of flavanol release from the original deposits is not yet clear.

Histochemical discrimination of fibers in regenerating rat infraorbital nerve

NASA Technical Reports Server (NTRS)

Wilke, R. A.; Riley, D. A.; Sanger, J. R.

1992-01-01

In rat dorsal root ganglia, histochemical staining of carbonic anhydrase (CA) and cholinesterase (CE) yields a reciprocal pattern of activity: Sensory processes are CA positive and CE negative, whereas motor processes are CA negative and CE positive. In rat infraorbital nerve (a sensory peripheral nerve), we saw extensive CA staining of nearly 100% of the myelinated axons. Although CE reactivity in myelinated axons was extremely rare, we did observe CE staining of unmyelinated autonomic fibers. Four weeks after transection of infraorbital nerves, CA-stained longitudinal sections of the proximal stump demonstrated 3 distinct morphological zones. A fraction of the viable axons retained CA activity to within 2 mm of the distal extent of the stump, and the stain is capable of resolving growth sprouts being regenerated from these fibers. Staining of unmyelinated autonomic fibers in serial sections shows that CE activity was not retained as far distally as is the CA sensory staining.

Two new mutations in the 3' coding region of the glycogen debranching enzyme in a glycogen storage disease type IIIa Ashkenazi Jewish patient.

PubMed

Parvari, R; Shen, J; Hershkovitz, E; Chen, Y T; Moses, S W

1998-04-01

Glycogen storage disease type III (GSD III) is an autosomal recessive disease caused by the deficiency of glycogen debranching enzyme (AGL). We report the finding of two new mutations in a GSD IIIa Ashkenazi Jewish patient. Both mutations are insertion of an adenine into a stretch of 8 adenines towards the 3' end of the coding region, one at position 3904 (3904insA) in exon 30, the second at position 4214 (4214insA) in exon 32. The mutations cause frameshifts and premature terminations of the glycogen debranching enzyme, the first causing a frameshift at amino acid 1304, the second causing a frameshift at amino acid 1408 of the total of 1532. These mutations demonstrate the importance of the 125 amino acids at the carboxy-terminus of the debrancher enzyme for its activity and support the suggestion that the putative glycogen binding domain is located in the carboxy-terminus of the AGL. The mutations cause distinctive single-strand conformation polymorphism (SSCP) patterns enabling easy detection.

Unexplained high thyroid stimulating hormone: a "BIG" problem.

PubMed

Mendoza, Heidi; Connacher, Alan; Srivastava, Rajeev

2009-01-01

Macro-hormones and macro-enzymes are high molecular weight conjugates of hormones or enzymes, respectively, often with immunoglobulins. These are referred to as macromolecular complexes, and may cause artefactually elevated biochemical tests results. Macro enzymes of the most commonly measured serum enzymes have been identified and are recognised as a source of elevated measurements that may cause diagnostic confusion; macro-creatine kinase and macro-amylase are the two better known macro-enzymes in clinical practice. Literature on macro-hormones is largely restricted to macro-prolactin. We present a case of a clinically euthyroid patient, who had persistently elevated thyroid stimulating hormone (TSH) but free thyroxine within the reference limits. She underwent repeated thyroid investigations and thyroid hormone interference studies, until macro-TSH was identified as the most likely cause of unexplained elevated TSH. Following the identification and characterisation of this biochemical abnormality, she is no longer subject to repeated blood tests for assessment of thyroid function; the patient currently remains clinically euthyroid.

Changes in Activities of Respiratory Enzymes in Lungs of Guinea-pigs Exposed to Silica Dust: II. Comparison of the Effects of Quartz Dust and Lampblack on the Succinate Oxidase System

PubMed Central

Breyer, Maria G.; Kilroe-Smith, T. A.; Prinsloo, H.

1964-01-01

Kilroe-Smith and Breyer (1963) reported that in the early stages of silicosis in guinea-pigs exposed to the inhalation of quartz dust, before the formation of collagen, there were increases in the specific activities of the complete succinate oxidase system and succinate dehydrogenase. The effects on these enzymes of quartz dust have now been compared with the effects of the fibrogenically `inert' lampblack. Lampblack causes a slight increase in the specific activities of these enzymes but the effects are small compared to those caused by quartz. Lampblack also causes a much smaller increase in lung weight than quartz, thus the enzyme increases are roughly parallel to the rise in lung weight. It appears that the effects observed on the enzymes are part of the general pattern associated with the early stages of the development of silicosis. PMID:14106132

Histochemistry in biology and medicine: a message from the citing journals.

PubMed

Pellicciari, Carlo

2015-12-23

Especially in recent years, biomedical research has taken advantage of the progress in several disciplines, among which microscopy and histochemistry. To assess the influence of histochemistry in the biomedical field, the articles published during the period 2011-2015 have been selected from different databases and grouped by subject categories: as expected, biological and biomedical studies where histochemistry has been used as a major experimental approach include a wide of basic and applied researches on both humans and other animal or plant organisms. To better understand the impact of histochemical publications onto the different biological and medical disciplines, it was useful to look at the journals where the articles published in a multidisciplinary journal of histochemistry have been cited: it was observed that, in the five-years period considered, 20% only of the citations were in histochemical periodicals, the remaining ones being in journals of Cell & Tissue biology,  general and experimental Medicine, Oncology, Biochemistry & Molecular biology, Neurobiology, Anatomy & Morphology, Pharmacology & Toxicology, Reproductive biology, Veterinary sciences, Physiology, Endocrinology, Tissue engineering & Biomaterials,  as well as in multidisciplinary journals.It is easy to foresee that also in the future the histochemical journals will be an attended forum for basic and applied scientists in the biomedical field. It will be crucial that these journals be open to an audience as varied as possible, publishing articles on the application of refined techniques to very different experimental models: this will stimulate non-histochemist scientists to approach histochemistry whose application horizon could expand to novel and possibly exclusive subjects.

[Histochemical and histoenzymatic study of experimental endarteritis in rabbits. I. Femoral endarteritis].

PubMed

Abou-Haila, A; Hadjiisky, P; Roland, J; Orcel, L

1978-04-01

The parietal reaction after placing a cuff of polyethylene around the femoral artery has been studied in 18 (2,5 - 3 months old) male rabbits by using histologic, histochemical (4 macromolecular substances) and histoenzymatic techniques (16 enzymatic activities). Studies were performed on the 1st, 3rd, 5th, 15th and 21st day, and every 15 days during the 3 months of the experiment. This process induced rapidly in each animal a parietal reaction with adventitial oedema (1st day), hypoxia of the media (1st -5th day), cytoenzymatic activation followed by a cellular transformation and proliferation of the intima-media, that forms a diffuse intimal thickening (adaptation). Most cells of the thickening were, by their enzymatic activities, quite comparable to immature smooth muscle cells, which probably emigrated from the media: intense LDH, NADH2 - TR; moderate G6P-DH, SDH, NADPH2 - TR, alpha-GP-DH, ATP/ase; weak ICHD, beta-HB-DH. Moreover, some reactions (accentuation of beta-Glu/ase, UDGP-DH, Glu-DH, 5'N/ase) besides suggested the active participation of the cells in the production of extracellular conjonctive constituents, because the histochemical studies revealed the presence of metachromatic glycosamino-glycanes and positive APS substances during the edification of the diffuse intimal thickening. In advanced thickening, an histoenzymatic duality was observed, that might prove the double origin of the thickening cells: some superficial cells had the enzymatic characteristics of endothelial cells (increased activity of aerobic oxydoreductases). At every stage of the study, the thickening cells differed from the atherocytes by a lack of lipids in their cytoplasm.

Histochemistry in Biology and Medicine: A Message From the Citing Journals

PubMed Central

2015-01-01

Especially in recent years, biomedical research has taken advantage of the progress in several disciplines, among which microscopy and histochemistry. To assess the influence of histochemistry in the biomedical field, the articles published during the period 2011-2015 have been selected from different databases and grouped by subject categories. As expected, biological and biomedical studies where histochemistry has been used as a major experimental approach include a wide range of basic and applied researches on both humans and other animal or plant organisms. To better understand the impact of histochemical publications onto the different biological and medical disciplines, it was useful to look at the journals where the articles published in a multidisciplinary journal of histochemistry have been cited: it was observed that, in the five-years period considered, 20% only of the citations were in histochemical periodicals, the remaining ones being in journals of Cell & Tissue biology, general and experimental Medicine, Oncology, Biochemistry & Molecular biology, Neurobiology, Anatomy & Morphology, Pharmacology & Toxicology, Reproductive biology, Veterinary sciences, Physiology, Endocrinology, Tissue engineering & Biomaterials, as well as in multidisciplinary journals. It is easy to foresee that also in the future the histochemical journals will be an attended forum for basic and applied scientists in the biomedical field. It will be crucial that these journals be open to an audience as varied as possible, publishing articles on the application of refined techniques to very different experimental models: this will stimulate non-histochemist scientists to approach histochemistry whose application horizon could expand to novel and possibly exclusive subjects. PMID:26708189

Acetylcholinesterase activity in soleus muscle intrafusal and extrafusal fibres in tail suspended rats.

PubMed

Tang, Bin; Fan, Xiao-li; Wu, Su-di

2002-10-01

Objective. To explore the mechanisms involved in muscle atrophy and conversion of the fiber types induced by simulated weightlessness. Method. Weightlessness was simulated by tail suspension of female rats. Intrafusal and extrafusal fibers of soleus muscles in the rat were examined histochemically for their activity of acetylcholinesterase (AChE) and succinic dehydrogenase (SDH) in 7 d, 14 d, 21 d tail-suspended groups and control groups. Result. Staining for succinic dehydrogenase showed that simulated weightlessness caused obvious atrophy and change in fiber type composition in soleus muscle, with decrease of the proportion of type I fiber and increase of type II fiber. Acetylcholinesterase activities of intrafusal and extrafusal fibers were both decreased significantly after 21 d tail suspension. Conclusion. Simulated weightlessness could induce decrease of AChE activity in neuromuscular junctions, which might be linked with decrease in motor neuron activity.

Cutaneous protothecosis.

PubMed

Hillesheim, Paul B; Bahrami, Soon

2011-07-01

Prototheca species are an achlorophyllic algae that cause infections primarily in immunocompromised individuals. At least one-half of infectious cases are cutaneous. Because protothecosis is seldom suspected clinically, patients may be subjected to various treatment modalities for extended periods without satisfactory results. Cutaneous protothecosis shares similar clinical and pathologic findings with deep tissue fungal mycoses. The typical presentation occurs most commonly on the face and extremities as erythematous plaques, nodules, or superficial ulcers. Prototheca spp are spherical, unicellular, nonbudding organisms that are sometimes noted on routine hematoxylin-eosin staining but are best visualized with periodic acid-Schiff and Gomori methenamine-silver histochemical stains. Although protothecosis can be diagnosed on biopsy, culture of the organism on a medium such as Sabouraud dextrose agar is required for definitive diagnosis. Treatment may require a combination of surgical excision and antifungal agents. Therefore, cutaneous protothecosis should be considered in a lesion that appears suspicious for the more-common fungal infections.

The anti-human immunodeficiency virus agent 3'-fluorothymidine induces DNA damage and apoptosis in human lymphoblastoid cells.

PubMed Central

Sundseth, R; Joyner, S S; Moore, J T; Dornsife, R E; Dev, I K

1996-01-01

Patients infected with the human immunodeficiency virus experienced severe hematopoietic toxicity after treatment with the deoxynucleoside analog 3'-fluorothymidine (FLT). Using several methods for the analysis of genome integrity, including histochemical staining of the 3' ends of DNA and both conventional and pulsed-field agarose gel electrophoresis, we demonstrated that FLT caused extensive DNA fragmentation in CEM cells that was not observed when these cells were treated with other, less toxic thymidine analogs. In addition, a distinctive pattern of small DNA fragments that is characteristic of cells in the process of programmed cell death was observed in the genomic DNA of CEM cells treated with FLT. We conclude that FLT induces DNA fragmentation and apoptosis in a human cell line of hematopoietic origin, and we offer this observation as a possible explanation for the severe toxicity of FLT observed in vivo. PMID:8834875

Quantitation and histochemical localization of galectin-1 and galectin-1-reactive glycoconjugates in fetal development of bovine organs.

PubMed

Kaltner, H; Lips, K S; Reuter, G; Lippert, S; Sinowatz, F; Gabius, H J

1997-10-01

The display of cellular oligosaccharide chains is known to undergo marked developmental changes, as monitored histochemically with plant lectins. In conjunction with endogenous lectins respective ligand structures may have a functional role during fetal development. The assumption of a recognitive, functionally productive interplay prompts the study of the expression of a tissue lectin and of lectin-reactive glycoconjugates concomitantly. Focusing on common beta-galactosides as constituents of oligosaccharide chains and the predominant member of the family of galectins in mammals, namely galectin-1, the question therefore is addressed as to whether expression of lectin and lectin-reactive glycoconjugates exhibits alterations, assessed in three morphologically defined fetal stages and in adult bovine organs. Using a sandwich ELISA, the level of the rather ubiquitous galectin-1 is mostly increased in adult organs relative to respective fetal stages, except for the case of kidney. This developmental course is seen rather seldom, when the amounts of lectin-reactive glycoproteins or glycolipids are quantitated in solid-phase assays after tissue homogenization. Western blotting, combined with probing by labeled galectin-1, discloses primarily quantitative changes in the reactivity of individual glycoproteins. Performing the same assays on extract aliquots with a plant agglutinin, namely the galactoside-binding mistletoe lectin, whose fine specificity is different from galectin-1, its reduced extent of binding in solid-phase assays and the disparate profile of lectin-reactive glycoproteins reveal a non-uniform developmental alteration within the group of structural variants of beta-galactosides. Although sample preparation can affect ligand preservation and/or presentation and thus restricts the comparability of biochemical and histochemical results, especially for soluble reactants, the histochemical studies on frozen and paraffin-embedded sections of bovine heart, kidney and liver demonstrate that the localization of the galectin and of lectin-reactive epitopes can show a similar distribution, as seen in liver and heart, with organ-typical quantitative changes of a rather similar staining profile (heart, kidney) or notable changes in the spatial distribution (liver) in the course of development. This report emphasizes the potential value of combined monitoring of the lectin and its potential in vivo ligands to contribute to eventually unravel organ-related function(s) of a tissue lectin.

Modifications Caused by Enzyme-retting and Their Effect on Composite Performance

USDA-ARS?s Scientific Manuscript database

Bethune seed flax was collected from Canada with seed removed using a stripper header and straw pulled and left in field for several weeks. Unretted straw was decorticated providing a coarse fiber bundle feedstock for enzyme treatments. Enzyme treatments using a bacterial pectinolytic enzyme with ...

Use of the spectrophotometric color method for the determination of the age of skin lesions on the pig carcass and its relationship with gene expression and histological and histochemical parameters.

PubMed

Vitali, M; Conte, S; Lessard, M; Deschêne, K; Benoit-Biancamano, M O; Celeste, C; Martelli, G; Sardi, L; Guay, F; Faucitano, L

2017-09-01

The presence of lesions on the pig carcass is an indicator of poor animal welfare and has economic impact as it downgrades the carcass value. The assessment of the age of lesions on the carcass may help identify risk factors and ultimately prevent their occurrence. The aim of this study was to assess the age of lesions on pig carcasses through spectrophotometric color evaluation and to relate the results with gene expression and histological and histochemical parameters. A total of 96 barrows were mixed 4 times over 3 d before slaughter and 80 lesions were selected after skin lesion observations to define 4 age categories: < 7 h (T1), 7-25 h (T2), 25-30 h (T3), and 49-54 h (T4). A nonlesioned skin area was used as a control. At slaughter, 3 biopsies per lesion and control skin were taken immediately after bleeding for analyses of gene expression (, , , , , , , , , ), skin histological characteristics (inflammation, erosion or ulceration, and necrosis), and enzyme activity (alkaline phosphatase and adenosine triphosphatase). The number of lesions was counted on each carcass, and the color was assessed visually by a pictorial chart and instrumentally through a spectrophotometer. Delta values (Δ) were calculated as the difference between the value of the lesion and the value of the control for all measures, except for the histological analysis. Results indicated that visual color observation was not sufficiently accurate to discriminate lesions by time of infliction ( > 0.10), while the spectrophotometer ΔL* and Δa* values variation allowed the identification of < 7 h or > 25 h old lesions ( < 0.05). Similarly, the expression of , , , , and genes was higher ( < 0.05) in < 7 h old lesions, while gene expression was higher ( < 0.05) in < 25 h old lesions. As for the histological analysis, the severity of inflammation was correlated with the age of the lesion (lower score in < 7 h old lesions and higher score in > 25 h old lesions; < 0.05). To conclude, the spectrophotometric color assessment of the carcass lesions at slaughter appears to be a reliable method to discriminate between fresh and older lesions on the carcass at the abattoir.

[Phenotypic and genotypic spectra of patients with glucose-6-phosphate dehydrogenase deficiency gene known pathogenic variants: a single-center study].

PubMed

Chen, X; Yang, L; Wang, H J; Wu, B B; Lu, Y L; Dong, X R; Zhou, W H

2018-05-02

Objective: To analyze the hotspots of known pathogenic disease-causing variants of glucose-6-phosphate dehydrogenase (G6PD) and the phenotype spectrum of neonatal patients with known pathogenic disease-causing variants of G6PD. Methods: The known pathogenic disease-causing variants of G6PD were collected from Human Gene Mutation Database. Screening was performed for these variants among the 7 966 cases (2 357 neonatal, 5 609 non-neonatal) in the database of sequencing at Molecular Diagnosis Center, Children's Hospital of Fudan University. All these samples were from patients suspected with genetic disorder. The database contained Whole Exon Sequencing data and Clinical Exon Sequencing data. We screened out the patients with known pathogenic disease-causing variants of G6PD, analyzed the hotspot of G6PD and the phenotype spectrum of neonatal patients with known pathogenic disease-causing variants of G6PD. Results: (1) Among the next generation sequencing data of the 7 966 samples, 86 samples (1.1%) were detected as positive for the known pathogenic disease-causing variants of G6PD (positive samples set). In the positive sample set, 51 patients (33 males, 18 females) were newborn babies. Forty-three patients (26 males, 17 females) had the enzyme activity data of G6PD. (2) Among the 86 samples, Arg463His, Arg459Leu, Leu342Phe, Val291Met were the leading 4 disease-causing variants found in 72 samples (84%). (3) Male neonatal patients with the same variants had the statistically significant differences in enzyme activity: among 13 patients with Arg463His, enzyme activity of 9 patients was ranked as grade Ⅲ, 1 case ranked as Ⅳ, 3 cases had no activity data;among 10 patients with Arg459Leu, enzyme activity of 4 patients was ranked as Ⅱ, 4 cases ranked as Ⅲ, 2 cases had no activity data;among 2 patients with His32Arg, enzyme activity of one patient was ranked as Ⅱ, another was Ⅲ. Male neonatal patients with the same mutation and enzyme activity also had the statistically significant differences in phenotype spectrum: among 9 patients with Arg463His and level Ⅲ enzyme activity, 6 presented hyperbilirubinemia, 2 met the criteria for exchange transfusion therapy, 2 showed hemolysis;among 4 patients with Arg459Leu and level Ⅱ enzyme activity, 3 presented hyperbilirubinemia;among 4 patients with Arg459Leu and level Ⅲ enzyme activity, 2 presented hyperbilirubinemia, 1 met the standard of exchange transfusion therapy;among 3 patients with Val291Met and level Ⅲ enzyme activity, 1 presented hyperbilirubinemia. Conclusions: Arg463His, Arg459Leu, Leu342Phe, Val291Met were the hotspots variants for the G6PD. Patients with the same G6PD variants and sex present different phenotype, patients with the same G6PD variants, sex and enzyme activity also present different phenotype .

Modification of enzymes by use of high-pressure homogenization.

PubMed

Dos Santos Aguilar, Jessika Gonçalves; Cristianini, Marcelo; Sato, Helia Harumi

2018-07-01

High-pressure is an emerging and relatively new technology that can modify various molecules. High-pressure homogenization (HPH) has been used in several studies on protein modification, especially in enzymes used or found in food, from animal, plant or microbial resources. According to the literature, the enzymatic activity can be modulated under pressure causing inactivation, stabilization or activation of the enzymes, which, depending on the point of view could be very useful. Homogenization can generate changes in the structure of the enzyme modifying various chemical bonds (mainly weak bonds) causing different denaturation levels and, consequently, affecting the catalytic activity. This review aims to describe the various alterations due to HPH treatment in enzymes, to show the influence of high-pressure on proteins and to report the HPH effects on the enzymatic activity of different enzymes employed in the food industry and research. Copyright © 2018 Elsevier Ltd. All rights reserved.

Relevance of pharmacogenetic aspects of mercaptopurine metabolism in the treatment of interstitial lung disease.

PubMed

Bakker, Jaap A; Drent, Marjolein; Bierau, Jörgen

2007-09-01

Mercaptopurine therapy is increasingly important as immunosuppressive therapy in interstitial lung disease. We focus on human mercaptopurine metabolism and the defects in this metabolism causing adverse drug reactions. Defects in mercaptopurine metabolizing enzymes like thiopurine methyltransferase and inosine triphosphate pyrophosphohydrolase lead to severe adverse drug reactions, sometimes with fatal outcome. Other enzymes, still not thoroughly investigated, can give rise to toxic effects or decreased efficacy in mercaptopurine therapy when the activity of these enzymes is altered. Pharmacogenetic screening of potential patients for mercaptopurine therapy is important to avoid adverse drug reactions caused by inherited enzyme deficiencies in these metabolic pathways. Pretreatment screening for deficiencies of mercaptopurine metabolizing enzymes will significantly reduce the number of patients with an adverse drug reaction and concomitantly associated healthcare costs.

Ballistic Motion of Enzymes that Catalyze Highly Exothermic Reactions

NASA Astrophysics Data System (ADS)

Tsekouras, Konstantinos; Pressé, Steve

Recently we proposed that the experimentally observed enhanced diffusion of enzymes catalyzing highly exothermic reactions is a consequence of their mechanism for dissipating reaction energy. More specifically, we proposed that reaction energy spreads out from the reaction site in the form of an acoustic wave which causes the enzyme to asymmetrically deform into the solvent. The solvent reaction propels the enzyme. However, it has been noted that in water, high viscosity should reduce enzyme momentum to zero within a few ps, so any diffusion increase should not be observable. Here we provide a model explaining how small volumetric expansions of biomolecules inside water may cause fluid compression that in turn creates regions of low fluid density around the biomolecule. We then investigate the dynamics of the biomolecule in the presence of these perturbations.

Affinity alkylation of the active site of C/sub 21/ steroid side-chain cleavage cytochrome P-450 from neonatal porcine testis: a unique cysteine residue alkylated by 17-(bromoacetoxy)progesterone

DOE Office of Scientific and Technical Information (OSTI.GOV)

Onoda, M.; Haniu, M.; Yanagibashi, K.

1987-01-27

The affinity alkylating progesterone analogue 17-(bromoacetoxy)progesterone has been used to label the active site of a microsomal cytochrome P-450 enzyme from neonatal pig testis. The enzyme causes removal of the C/sub 20/ and C/sub 21/ side chains from the substrates progesterone and pregnenolone by catalyzing both 17-hydroxylase and C/sub 17,20/-lyase reactions, which produce the corresponding C/sub 1//sup 9/ steroidal precursors of testosterone. The progesterone analogue causes simultaneous inactivation of the two catalytic activities of the enzyme by a first-order kinetic process that obeys saturation kinetics. Progesterone and 17-hydroxyprogesterone each protect the enzyme against inactivation. The progesterone analogue is a competitivemore » inhibitor of the enzyme with K/sub i/ values of 8.4 ..mu..M and 7.8 ..mu..M for progesterone and 17-hydroxyprogesterone, respectively. The enzyme inactivation and kinetic data are consistent with a theory proposing that the analogue and the two substrates compete for the same active site. The radioactive analogue 17-((/sup 14/C)bromoacetoxy)progesterone causes inactivation of the enzyme with incorporation of 1.5-2.2 mol of the analogue per mole of inactivated enzyme. When this experiment is carried out in the presence of a substrate, then 0.9-1.2 mol of radioactive analogue is incorporated per mole of inactivated enzyme. The data suggest that the analogue can bind to two different sites, one of which is related to the catalytic site. Radiolabeled enzyme samples, from reactions of the /sup 14/C-labeled analogue with the enzyme alone or with enzyme in the presence of a substrate, were subjected to amino acid analysis and also in tryptic digestion and peptide mapping.« less

15-Hydroxyprostaglandin dehydrogenase is upregulated by hydroxychloroquine in rheumatoid arthritis fibroblast-like synoviocytes

PubMed Central

KIM, HAK-JAE; LEE, SORA; LEE, HAW-YONG; WON, HANSOL; CHANG, SUNG-HAE; NAH, SEONG-SU

2015-01-01

15-Hydroxyprostaglandin dehydrogenase (HPGD) is the key enzyme responsible for the metabolic inactivation of prostaglandin E2 (PGE2) catabolism. PGE2 is one of the predominant catabolic factors involved in rheumatoid arthritis (RA). However, the expression and regulation of HPGD in RA fibroblast-like synoviocyte (FLS) remain to be elucidated. Disease-modifying anti-rheumatic drugs (DMARDs) are the most important anti-arthritic drugs, which reduce the effect of joint injury. The aim of the present study was to assess the expression of HPGD in RA tissues and cells, and normal synovial tissues and cells. The effect of the most popular DMARDs, hydroxychloroquine, on the expression of HPGD in RA-FLS was also investigated. Western blotting and immuno-histochemical analysis demonstrated that the expression levels of HPGD in human synovium were lower in RA synovium compared with the normal and OA synovium. In RA-FLS, the expression of HPGD was increased following treatment with several DMARDs, including sulfasalazine, methotrexate, and hydroxychloroquine. Hydroxychloroquine (10 µM) treatment induced the phosphorylation of ERK, SAPK/JNK and p38. Hydroxychloroquine induced a decrease in the release of PGE2, which was restored by mitogen-activated protein (MAP) kinase pathway inhibitors. Hydroxychloroquine may therefore, affect the pathogenesis of RA through the MAP kinase pathway by regulating the expression of HPGD. PMID:26082314

In vivo marking of spontaneous or vaccine-induced fibrosarcomas in the domestic house cat, using an adenoviral vector containing a bifunctional fusion protein, GAL-TEK.

PubMed

Marini, F C; Cannon, J P; Belmont, J W; Shillitoe, E J; Lapeyre, J N

1995-09-01

We evaluated the ability of a replication-deficient, recombinant adenoviral vector to transfer the bifunctional gene GAL-TEK, which expresses a marking/therapeutic gene product, to naturally occurring cat fibrosarcomas in situ. GAL-TEK contains an in-frame fusion of the bacterial LacZ gene for histochemical marking of tumors with beta-galactosidase (beta-Gal) and the HSV tk gene for enzyme-prodrug activation of the prodrug ganciclovir (GCV) to induce selective tumor cell killing. GAL-TEK bifunctional marking and cell killing activities were tested in vitro after adenoviral vector infection of HT1080 human fibrosarcoma cells. The tk activity of GAL-TEK is shown to be almost as potent as HSV tk to catalyze conversion of GCV to GCV nucleotides and promote selective cell killing. Using 8 cats with recurring 2.5-cm2 fibrosarcomas that either arose spontaneously or were induced by vaccine, we determined experimentally the administration routes and times required for optimum GAL-TEK gene transfer by beta-Gal histological staining and reverse transcriptase polymerase chain reaction to the multiple compartments of the growing fibrosarcomas consonant with minimizing collateral infection of neighboring tissues and other unwanted side effects.

The distribution of cholinesterases in the cat carotid body

PubMed Central

Biscoe, T. J.; Silver, Ann

1966-01-01

1. The distribution of acetyl- and butyrylcholinesterase in the carotid body of the cat has been examined histochemically. Studies were made on normal carotid bodies and on carotid bodies from cats in which certain nerves had been cut some time previously. The nerves sectioned were the sinus nerve, the post-ganglionic sympathetic branch of the superior cervical ganglion or the preganglionic cervical sympathetic trunk. 2. It was confirmed that more butyrylcholinesterase than acetylcholinesterase is present. Both enzymes are found in three sites: (i) as strands, (ii) as plexuses, (iii) inside a few cells. 3. The distribution is unaffected by cutting the sinus nerve or preganglionic cervical sympathetic nerves. Disorganization and depletion of the cholinesterases in the strands and plexuses occurs when the post-ganglionic branch of the superior cervical ganglion is cut. The cholinesterase in cells is unaffected. 4. In carotid bodies in which vessels were filled with red blood cells or in which the vascular bed was injected with carmine-gelatine, it was seen that strands and plexuses are associated with blood vessels, and with blood vessels and cells respectively. 5. It is suggested that a cholinergic pathway controlling carotid body blood vessels runs in the post-ganglionic cervical sympathetic. ImagesabcdefPlate 2abcdef PMID:5942823

Mitochondrial modulators in experimental Huntington's disease: reversal of mitochondrial dysfunctions and cognitive deficits.

PubMed

Mehrotra, Arpit; Kanwal, Abhinav; Banerjee, Sanjay Kumar; Sandhir, Rajat

2015-06-01

Huntington's disease (HD) is a chronic neurodegenerative condition involving impaired mitochondrial functions. The present study evaluates the therapeutic potential of combined administration of mitochondrial modulators: alpha-lipoic acid and acetyl-l-carnitine on mitochondrial dysfunctions in 3-NP-induced HD. Our results reveal 3-NP administration resulted in compromise of mitochondrial functions in terms of: (1) impaired activity of mitochondrial respiratory chain enzymes, altered cytochrome levels, reduced histochemical staining of complex-II and IV, reduced in-gel activity of complex-I to V, and reduced mRNA expression of respiratory chain complexes; (2) enhanced mitochondrial oxidative stress indicated by increased malondialdehyde, protein carbonyls, reactive oxygen species and nitrite levels, along with decreased Mn-superoxide dismutase and catalase activity; (3) mitochondrial structural changes measured by mitochondrial swelling, reduced mitochondrial membrane potential and ultra-structure changes; (4) increased cytosolic cytochrome c levels, caspase-3 and -9 activity along with altered expression of apoptotic proteins (AIF, Bim, Bad, and Bax); and (5) impaired cognitive functions assessed using Morris water maze and Y-maze. Combination of mitochondrial modulators (alpha-lipoic acid + acetyl-l-carnitine) on the other hand ameliorated 3-NP-induced mitochondrial dysfunctions, oxidative stress, histologic alterations, and behavioral deficits, suggesting their therapeutic efficacy in the management of HD. Copyright © 2015 Elsevier Inc. All rights reserved.

Low Temperature Induces the Accumulation of Phenylalanine Ammonia-Lyase and Chalcone Synthase mRNAs of Arabidopsis thaliana in a Light-Dependent Manner.

PubMed Central

Leyva, A.; Jarillo, J. A.; Salinas, J.; Martinez-Zapater, J. M.

1995-01-01

Anthocyanins, which accumulate in leaves and stems in response to low temperature and changes in light intensity, are synthesized through the phenylpropanoid pathway that is controlled by key enzymes that include phenylalanine ammonia-lyase (PAL) and chalcone synthase (CHS). In this work we demonstrate that PAL and CHS mRNAs accumulate in leaves of Arabidopsis thaliana (L.) Heynh. upon exposure to low temperature in a light-dependent manner. The regulation of the PAL1 gene expression by low temperature and light was examined by analyzing the expression of the [beta]-glucuronidase (uidA) reporter gene in transgenic Arabidopsis plants containing the uidA gene of Escherichia coli under the control of the PAL1 promoter. The results indicate that the accumulation of PAL1 mRNA is transcriptionally regulated. Histochemical staining for [beta]-glucuronidase activity showed that the PAL1 promoter is preferentially activated in photosynthetically active cells, paralleling anthocyanin accumulation. Moreover, we show that light may also be implicated in the regulation of the CHS gene in response to bacterial infiltration. Finally, using two transparent testa Arabidopsis mutants that are unable to accumulate anthocyanins, we demonstrate that these pigments are not required for successful development of freezing tolerance in this species. PMID:12228452

Preventive effects of Chlorella on skeletal muscle atrophy in muscle-specific mitochondrial aldehyde dehydrogenase 2 activity-deficient mice.

PubMed

Nakashima, Yuya; Ohsawa, Ikuroh; Nishimaki, Kiyomi; Kumamoto, Shoichiro; Maruyama, Isao; Suzuki, Yoshihiko; Ohta, Shigeo

2014-10-11

Oxidative stress is involved in age-related muscle atrophy, such as sarcopenia. Since Chlorella, a unicellular green alga, contains various antioxidant substances, we used a mouse model of enhanced oxidative stress to investigate whether Chlorella could prevent muscle atrophy. Aldehyde dehydrogenase 2 (ALDH2) is an anti-oxidative enzyme that detoxifies reactive aldehydes derived from lipid peroxides such as 4-hydroxy-2-nonenal (4-HNE). We therefore used transgenic mice expressing a dominant-negative form of ALDH2 (ALDH2*2 Tg mice) to selectively decrease ALDH2 activity in the muscles. To evaluate the effect of Chlorella, the mice were fed a Chlorella-supplemented diet (CSD) for 6 months. ALDH2*2 Tg mice exhibited small body size, muscle atrophy, decreased fat content, osteopenia, and kyphosis, accompanied by increased muscular 4-HNE levels. The CSD helped in recovery of body weight, enhanced oxidative stress, and increased levels of a muscle impairment marker, creatine phosphokinase (CPK) induced by ALDH2*2. Furthermore, histological and histochemical analyses revealed that the consumption of the CSD improved skeletal muscle atrophy and the activity of the mitochondrial cytochrome c oxidase. This study suggests that long-term consumption of Chlorella has the potential to prevent age-related muscle atrophy.

The bouquet of grapevine (Vitis vinifera L. cv. Cabernet Sauvignon) flowers arises from the biosynthesis of sesquiterpene volatiles in pollen grains

PubMed Central

Martin, Diane M.; Toub, Omid; Chiang, Angela; Lo, Bernard C.; Ohse, Sebastian; Lund, Steven T.; Bohlmann, Jörg

2009-01-01

Terpenoid volatiles are important information molecules that enable pollinators to locate flowers and may protect reproductive tissues against pathogens or herbivores. Inflorescences of grapevine (Vitis vinifera L.) are composed of tiny green flowers that produce an abundance of sesquiterpenoid volatiles. We demonstrate that male flower parts of grapevines are responsible for sesquiterpenoid floral scent formation. We describe temporal and spatial patterns of biosynthesis and release of floral volatiles throughout the blooming of V. vinifera L. cv. Cabernet Sauvignon. The biosynthesis of sesquiterpene volatiles, which are emitted with a light-dependent diurnal pattern early in the morning at prebloom and bloom, is localized to anthers and, more specifically, within the developing pollen grains. Valencene synthase (VvValCS) enzyme activity, which produces the major sesquiterpene volatiles of grapevine flowers, is present in anthers. VvValCS transcripts are most abundant in flowers at prebloom stages. Western blot analysis identified VvValCS protein in anthers, and in situ immunolabeling located VvValCS protein in pollen grains during bloom. Histochemical staining, as well as immunolabeling analysis by fluorescent microscopy and transmission electron microscopy, indicated that VvValCS localizes close to lipid bodies within the maturing microspore. PMID:19359488

NADP-dependent enzymes are involved in response to salt and hypoosmotic stress in cucumber plants.

PubMed

Hýsková, Veronika; Plisková, Veronika; Červený, Václav; Ryšlavá, Helena

2017-07-01

Salt stress is one of the most damaging plant stressors, whereas hypoosmotic stress is not considered to be a dangerous type of stress in plants and has been less extensively studied. This study was performed to compare the metabolism of cucumber plants grown in soil with plants transferred to distilled water and to a 100 mM NaCl solution. Even though hypoosmotic stress caused by distilled water did not cause such significant changes in the relative water content, Na+/K+ ratio and Rubisco content as those caused by salt stress, it was accompanied by more pronounced changes in the specific activities of NADP-dependent enzymes. After 3 days, the specific activities of NADP-isocitrate dehydrogenase, glucose-6-phosphate dehydrogenase, NADP-malic enzyme and non-phosphorylating glyceraldehyde-3-phosphate dehydrogenase in leaves were highest under hypoosmotic stress, and lowest in plants grown in soil. In roots, salt stress caused a decrease in the specific activities of major NADP-enzymes. However, at the beginning of salt stress, NADP-galactose-1-dehydrogenase and ribose-1-dehydrogenase were involved in a plant defense response in both roots and leaves. Therefore, the enhanced demands of NADPH in stress can be replenished by a wide range of NADP-dependent enzymes.

Superoxide poisons mononuclear iron enzymes by causing mismetallation

PubMed Central

Gu, Mianzhi; Imlay, James A.

2013-01-01

Summary Superoxide (O2−) is a primary agent of intracellular oxidative stress. Genetic studies in many organisms have confirmed that excess O2− disrupts metabolism, but to date only a small family of [4Fe-4S] dehydratases have been identified as direct targets. This investigation reveals that in Escherichia coli O2− also poisons a broader cohort of non-redox enzymes that employ ferrous iron atoms as catalytic cofactors. These enzymes were inactivated by O2− both in vitro and in vivo. Although the enzymes are known targets of hydrogen peroxide, the outcome with O2− differs substantially. When purified enzymes were damaged by O2− in vitro, activity could be completely restored by iron addition, indicating that the O2− treatment generated an apoprotein without damaging the protein polypeptide. Superoxide stress inside cells caused the progressive mismetallation of these enzymes with zinc, which confers little activity. When O2− stress was terminated, cells gradually restored activity by extracting zinc from the proteins. The overloading of cells with zinc caused mismetallation even without O2− stress. These results support a model in which O2− repeatedly excises iron from these enzymes, allowing zinc to compete with iron for remetallation of their apoprotein forms. This action substantially expands the physiological imprint of O2− stress. PMID:23678969

Use of bilidase for the treatment of experimental hypertrophic postburn cicatrices.

PubMed

Loladze, M; Alibegashvili, M; Turmanidze, Ts; Iashvili, B; Kutivadze, D; Chanishvili, T

2005-01-01

The efficiency of bilidase (hyaluronidase preparation) was studied in guinea pigs with experimental postburn cicatrices. Bilidase promoted normalization of the structure and histochemical picture of new cicatricial tissue. The drug can be used for the treatment of hypertrophic postburn cicatrices.

Problems in aerial application : histochemistry of Weil stain on liver.

DOT National Transportation Integrated Search

1966-11-01

By use of histochemical blocking and extraction procedures, it was possible to show that the cytoplasmic compound to which the Weil stain attaches is a lipid containing a secondary amine. It is suggested that the compound could be further characteriz...

Porcine Intestinal Mast Cells. Evaluation of Different Fixatives for Histochemical Staining Techniques Considering Tissue Shrinkage

PubMed Central

Rieger, J.; Twardziok, S.; Huenigen, H.; Hirschberg, R.M.; Plendl, J.

2013-01-01

Staining of mast cells (MCs), including porcine ones, is critically dependent upon the fixation and staining technique. In the pig, mucosal and submucosal MCs do not stain or stain only faintly after formalin fixation. Some fixation methods are particularly recommended for MC staining, for example the fixation with Carnoy or lead salts. Zinc salt fixation (ZSF) has been reported to work excellently for the preservation of fixation-sensitive antigens. The aim of this study was to establish a reliable histological method for counting of MCs in the porcine intestinum. For this purpose, different tissue fixation and staining methods that also allow potential subsequent immunohistochemical investigations were evaluated in the porcine mucosa, as well as submucosa of small and large intestine. Tissues were fixed in Carnoy, lead acetate, lead nitrate, Zamboni and ZSF and stained subsequently with either polychromatic methylene blue, alcian blue or toluidine blue. For the first time our study reveals that ZSF, a heavy metal fixative, preserves metachromatic staining of porcine MCs. Zamboni fixation was not suitable for histochemical visualization of MCs in the pig intestine. All other tested fixatives were suitable. Alcian blue and toluidine blue co-stained intestinal goblet cells which made a prima facie identification of MCs difficult. The polychromatic methylene blue proved to be the optimal staining. In order to compare MC counting results of the different fixation methods, tissue shrinkage was taken into account. As even the same fixation caused shrinkagedifferences between tissue from small and large intestine, different factors for each single fixation and intestinal localization had to be calculated. Tissue shrinkage varied between 19% and 57%, the highest tissue shrinkage was found after fixation with ZSF in the large intestine, the lowest one in the small intestine after lead acetate fixation. Our study emphasizes that MC counting results from data using different fixation techniques can only be compared if the respective studyimmanent shrinkage factor has been determined and quantification results are adjusted accordingly. PMID:24085270

The histochemical profile of the rat extensor digitorum longus muscle differentiates after birth and dedifferentiates in senescence.

PubMed

Lehnert, M; Laurer, H; Maier, B; Frank, J; Marzi, I; Steudel, W-I; Mautes, A

2007-01-01

Age dependent motor unit dedifferentiation is a key component of impaired muscle function in advanced age. Here, we tested the hypothesis that rat muscle histochemical profile during the lifespan of an individual has an age-specific pattern since comprehensive longitudinal studies of muscle differentiation after birth and dedifferentiation in advanced age are scarce. Our results show that extensor digitorum longus muscle (EDL) is comprised only of two fiber types after birth, type slow-oxidative (SO) and type SDH-intermediate (SDH-INT), the latter being indicative for the presence of polyneuronal innervation. In contrast to the constantly growing cross-sectional area of the muscle fibers, a dramatic decrease in SDH-INT proportion occurs between day 14 and 21 after birth resulting in a complete loss of fiber type SDH-INT at the age of 90 days (p<0.05). At the age of 270 days, the fiber type composition of rat EDL dedifferentiates as shown by the reappearance of the SDH-INT type with a further increase at the age of 540 days (p<0.05). These changes in histochemical fiber type spectra are brought about by fiber type conversion within the fast twich fibers. The findings of the present study provide further evidence that fiber type conversion is a basic mechanism leading to motor unit differentiation and dedifferentiation during ontogenesis. Fiber type conversion shows a distinct time specific pattern and is also characteristic for motor unit regeneration after peripheral nerve repair. Factors that influence fiber type conversion and thereby motor unit organization may provide a future therapeutic option to enhance the regenerative capacity of motor units.

Differential diagnosis of vacuolar muscle biopsies: use of p62, LC3 and LAMP2 immunohistochemistry.

PubMed

Vittonatto, Elisa; Boschi, Silvia; CHIADò-Piat, Loredana; Ponzalino, Valentina; Bortolani, Sara; Brusa, Chiara; Rainero, Innocenzo; Ricci, Federica; Vercelli, Liliana; Mongini, Tiziana

2017-12-01

Intrafibral vacuoles are the morphological hallmark in a wide variety of human skeletal muscle disorders with different etiology. In most cases, differential diagnosis is feasible with a routine histochemical work up of muscle biopsy. Ultrastructural analysis is an important confirmatory tool, but it is not widely available. Immunohistochemical stainings for p62, LAMP2 and LC3 are commonly available as tissutal marker for autophagy. We compared the immunohistochemical patterns for autophagic markers p62, LC3 and LAMP2 with routine histochemical markers in 39 biopsies from patients with definite diagnoses of glycogen storage disease type 2 (LOPD or Pompe disease, PD), sporadic inclusion body myositis (sIBM), oculo-pharyngeal muscular dystrophy (OPMD) and necrotizing myopathy (NM). Moreover, we also analyzed muscles of 10 normal controls. In PD group, LC3 and LAMP2 showed an higher percentage of positive fibers, whereas p62 was limited to a minority of fibers, thus paralleling the results of histochemical stainings; in NM group, LAMP2 and LC-3 were diffusely and unspecifically expressed in necrotic fibers, with p62 significantly expressed only in two cases. OPMD biopsies did not reveal any significant positivity. The most interesting results were observed in sIBM group, where p62 was expressed in all cases, even in fibers without other markers positivity. There results, although limited to a small number of cases, suggest that the contemporary use of p62, LAMP2 and LC-3 staining may have an adjunctive role in characterizing muscle fiber vacuoles, revealing autophagic pathway activation and providing further clues for the understanding of pathogenetic mechanisms.s.

Validation of musculoskeletal ultrasound to assess and quantify muscle glycogen content. A novel approach.

PubMed

Hill, John C; Millán, Iñigo San

2014-09-01

Glycogen storage is essential for exercise performance. The ability to assess muscle glycogen levels should be an important advantage for performance. However, skeletal muscle glycogen assessment has only been available and validated through muscle biopsy. We have developed a new methodology using high-frequency ultrasound to assess skeletal muscle glycogen content in a rapid, portable, and noninvasive way using MuscleSound (MuscleSound, LCC, Denver, CO) technology. To validate the utilization of high-frequency musculoskeletal ultrasound for muscle glycogen assessment and correlate it with histochemical glycogen quantification through muscle biopsy. Twenty-two male competitive cyclists (categories: Pro, 1-4; average height, 183.7 ± 4.9 cm; average weight, 76.8 ± 7.8 kg) performed a steady-state test on a cyclergometer for 90 minutes at a moderate to high exercise intensity, eliciting a carbohydrate oxidation of 2-3 g·min⁻¹ and a blood lactate concentration of 2 to 3 mM. Pre- and post-exercise glycogen content from rectus femoris muscle was measured using histochemical analysis through muscle biopsy and through high-frequency ultrasound scans using MuscleSound technology. Correlations between muscle biopsy glycogen histochemical quantification (mmol·kg⁻¹) and high-frequency ultrasound methodology through MuscleSound technology were r = 0.93 (P < 0.0001) pre-exercise and r = 0.94 (P < 0.0001) post-exercise. The correlation between muscle biopsy glycogen quantification and high-frequency ultrasound methodology for the change in glycogen from pre- and post-exercise was r = 0.81 (P < 0.0001). These results demonstrate that skeletal muscle glycogen can be measured quickly and noninvasively through high-frequency ultrasound using MuscleSound technology.

Morphological and histochemical characterization of the mucosa of the digestive tract in Engraulis anchoita.

PubMed

Díaz, A O; García, A M; Devincenti, C V; Goldemberg, A L

2003-12-01

The histomorphological aspects as well as the histochemical content and distribution of glycoproteins (GPs) in the mucosa of the digestive tract of the anchovy Engraulis anchoita were studied. The buccopharyngeal cavity is lined by a squamous stratified epithelium with mucous superficial cells; the oesophagus shows two zones, cranial with a squamous stratified epithelium with mucous superficial cells and caudal with a columnar secretory epithelium. Finally, the stomach presents both the cranial and pyloric portion lined with a simple columnar epithelium. Tubular branched glands, formed by a single type of glandular cell, located along the stomach, are more numerous in the cranial portion. The GPs were identified with (1) oxidizable vicinal diols; (2) sialic acids and some of their chain variants, C7 or C9; (3) sialic acid residues with O-acyl substitution at C7 or C8; (4) carboxyl groups and (5) sulphate groups. Histochemical tests showed that the buccopharyngeal cavity presented the largest amount of the different types of mucosubstances. Epithelial secretory cells were found in the oesophagus, which synthesized a large quantity of sialosulphoglycoproteins likely to be related to a protective role. The surface epithelium of the stomach synthesizes and secretes acid and neutral GPs, probably related to the movement of fluids and to the absorption of easily digested substrates, respectively. Although great differences exist between different species, in E. anchoita as in other fish species, the wall of the digestive tract is composed of the four layers classically described for vertebrates. The GPs secreted by the epithelial cells are suggested to be important for the protection and inhibition of microorganisms. In addition, they are involved in enzymatic digestion of food, absorptive functions and lubrication of the alimentary tract.

Histochemical and morphometric characteristics of the normal human vastus medialis longus and vastus medialis obliquus muscles.

PubMed Central

Travnik, L; Pernus, F; Erzen, I

1995-01-01

The histochemical and morphometric characteristics of the vastus medialis longus and vastus medialis obliquus muscles were studied and compared with data on vastus lateralis. Cross-sections of autopsied muscles from 9 healthy men, aged 18-44 y, who had died suddenly were analysed. Data were obtained on proportions, cross-sectional diameter, and on atrophy and hypertrophy factors, of type 1, 2a, 2b, and 2c fibres. The analysis showed that the anatomical differences and the different functional demands placed on vastus medialis longus and vastus medialis obliquus are also expressed in different proportions and sizes of fibre types in the two muscles. The proportion of type 1 fibres was significantly higher (P < 0.01), and the proportion of 2b fibres was significantly lower (P < 0.01) in vastus medialis longus than in vastus medialis obliquus. The diameters of type 1 and type 2a fibres were significantly smaller (P < 0.01) in vastus medialis longus than in vastus medialis obliquus, although the differences were small. Within muscles a nonrandom arrangement of fibre types existed with the deeper portions of the muscles having more type 1 fibres than the more superficial portions. The histochemical and morphometric characteristics of vastus lateralis and vastus medialis obliquus show great similarity, reflecting the common function of both muscles which is taking part in transverse knee stability. Estimates of the limits of normality of the proportion, diameter, atrophy and hypertrophy factors of type 1, 2a, 2b, and 2c fibres might be useful in obtaining information on how different physiological and pathological conditions influence the proportion and size of different fibre types.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7592003

Potential Involvement of Draxin in the Axonal Projection of Cranial Nerves, Especially Cranial Nerve X, in the Chick Hindbrain.

PubMed

Zhang, Sanbing; Cui, Huixian; Wang, Lei; Kang, Lin; Huang, Guannan; Du, Juan; Li, Sha; Tanaka, Hideaki; Su, Yuhong

2016-07-01

The appropriate projection of axons within the nervous system is a crucial component of the establishment of neural circuitry. Draxin is a repulsive axon guidance protein. Draxin has important functions in the guidance of three commissures in the central nervous system and in the migration of neural crest cells and dI3 interneurons in the chick spinal cord. Here, we report that the distribution of the draxin protein and the location of 23C10-positive areas have a strong temporal and spatial correlation. The overexpression of draxin, especially transmembrane draxin, caused 23C10-positive axon bundles to misproject in the dorsal hindbrain. In addition, the overexpression of transmembrane draxin caused abnormal formation of the ganglion crest of the IX and X cranial nerves, misprojection of some anti-human natural killer-1 (HNK-1)-stained structures in the dorsal roof of the hindbrain, and a simultaneous reduction in the efferent nerves of some motoneuron axons inside the hindbrain. Our data reveal that draxin might be involved in the fascicular projection of cranial nerves in the hindbrain. © 2016 The Histochemical Society.

In-situ chemical analyses of trans-polyisoprene by histochemical staining and Fourier transform infrared microspectroscopy in a rubber-producing plant, Eucommia ulmoides Oliver.

PubMed

Bamba, Takeshi; Fukusaki, Ei-Ichiro; Nakazawa, Yoshihisa; Kobayashi, Akio

2002-10-01

The localization of polyisoprene in young stem tissues of Eucommia ulmoides Oliver was investigated by histochemical staining and Fourier transform infrared (FT-IR) microspectroscopy. The fibrous structures were stained with Oil Red O. FT-IR microspectroscopic analysis proved that the fibrous structures were trans-polyisoprene. Granular structures stained with the dye, and characteristic absorptions at 2,960 cm(-1) and 1,430 cm(-1) in FT-IR suggested that trans-polyisoprene accumulated in the vicinity of the cambium layer. We have thus successfully shown for the first time the localization of trans-polyisoprene in plant tissues, and our histological investigation allowed us to presume the main sites of biosynthesis and accumulation of trans-rubber. Furthermore, a new technical approach, the preparation of sections using an electronic freezing unit and the in situ analysis of polyisoprene using FT-IR microspectroscopy, is demonstrated to be a promising method for determining the accumulation of polyisoprene as well as other metabolites.

Characterization of atherosclerosis by histochemical and immunohistochemical methods in African grey parrots (Psittacus erithacus) and Amazon parrots (Amazona spp.).

PubMed

Fricke, Cornelia; Schmidt, Volker; Cramer, Kerstin; Krautwald-Junghanns, Maria-Elisabeth; Dorrestein, Gerry M

2009-09-01

The aim of the study was to characterize atherosclerotic changes in African grey parrots (Psittacus erithacus) and Amazon parrots (Amazona spp.) by histochemical and immunohistochemical methods. Samples of the aorta ascendens and trunci brachiocephalici from 62 African grey parrots and 35 Amazon parrots were stained by hematoxylin and eosin and Elastica van Gieson for grading of atherosclerosis in these birds. Four different stages were differentiated. The incidence of atherosclerosis in the examined parrots was 91.9% in African grey parrots and 91.4% in Amazon parrots. To evaluate the pathogenesis in birds, immunohistochemical methods were performed to demonstrate lymphocytes, macrophages, smooth muscle cells, and chondroitin sulfate. According to the missing lymphocytes and macrophages and the absence of invasion and proliferation of smooth muscle cells in each atherosclerotic stage, "response-to-injury hypothesis" seems inapplicable in parrots. Additionally, we found alterations of vitally important organs (heart, lungs) significantly correlated with atherosclerosis of the aorta ascendens.

The salivary glands of Ameiva ameiva (Teiidae, Lacertilia). A morphological, morphometric and histochemical study.

PubMed

Lopes, R A; Costa, J R; Piccolo, A M; Petenusci, S O

1982-01-01

The authors studied morphological, morphometric, and histochemically the mucosubstances and proteins in the salivary glands of the lizard Ameiva. Based on the results, the authors concluded: 1. The labial salivary gland is formed by small mucous and mucoserous glands; the sublingual gland by mucoserous cells. 2. Mucous cells show neutral and sulphated mucosubstances and sialic acid. Mucoserous cells of the labial gland show neutral mucosubstance, sialic acid, hyaluronic acid and protein radicals. Mucoserous cells of the sublingual gland show neutral mucosubstance, sialic acid and protein radicals. 3. The average values for acinar area were: 1,198.11 microns 2 for mucoserous acini and 2,105.95 microns 2 for mucous acini of the labial salivary gland. The average values for nucleus volume were: 47.41 microns 3 for mucoserous cells and 38.97 microns 4 for mucous cells. 4. The average values for acinar area and nuclear volume of the mucoserous cells of the subingual gland were respectively: 1,474.62 microns 2 and 67.77 microns 3.

Recent studies on the developing human hepatocellular carcinoma.

PubMed

Gerber, M A

1986-01-01

From our knowledge of characteristic phenotypic changes of the preneoplastic lesions during the stepwise evolution of hepatocellular carcinoma (HCC) in experimental models, we are now beginning to define the structural, histochemical, biochemical, antigenic and molecular properties of early HCC and of the putative preneoplastic changes in human liver. Histological, ultrastructural, morphometric and immunohistochemical studies suggest that adenomatous nodules of regenerating and hyperplastic hepatocytes are more likely to represent direct precursors of HCC than dysplastic hepatocytes. Histochemical and immunomorphological investigations show appreciable functional and phenotypic heterogeneity of human HCC as previously recognized in experimental hepatocarcinogenesis. Studies of altered expression of oncogenes in the regenerating liver and HCC are beginning to define the molecular mechanisms in cell growth and malignant transformation. Although integration of Hepadna viral DNA sequences frequently occurs during persistent infection in man and animals, the exact mechanism of viral oncogenesis remains to be elucidated. It is likely that the development of monoclonal antibodies to surface antigens on transformed hepatocytes will be useful for exploring lineage relationships between the cell populations involved in hepatocarcinogenesis.

New Insights into Therapeutic Strategies for the Treatment of Peritoneal Fibrosis: Learning from Histochemical Analyses of Animal Models

PubMed Central

Kitamura, Mineaki; Nishino, Tomoya; Obata, Yoko; Ozono, Yoshiyuki; Koji, Takehiko; Kohno, Shigeru

2014-01-01

Encapsulating peritoneal sclerosis (EPS) is a fatal complication that can occur in patients undergoing long-term peritoneal dialysis. It is characterized by bowel obstruction and marked sclerotic thickening of the peritoneal membrane. Although the mechanisms underlying the development of EPS are complex, angiogenesis, inflammation, and peritoneal fibrosis are known to be essential factors. Now, several animal models that exhibit EPS have pathophysiology similar to that of human EPS and have been proposed for use in research to provide insights into it. Recent histochemical methods also help us to understand the pathophysiology of EPS. Advances in basic research based on the findings in those animal models have enabled the development of several strategies for the prevention and treatment of EPS. We describe here interventional studies in some animal models for peritoneal fibrosis, one of the histological disorders findings characteristic to EPS, and we highlight the need for a sophisticated animal model that closely resembles human conditions. PMID:25392567

Metabolic enzymes dysregulation in heart failure: the prospective therapy.

PubMed

Parihar, Priyanka; Parihar, Mordhwaj Singh

2017-01-01

The heart failure accounts for the highest mortality rate all over the world. The development of preventive therapeutic approaches is still in their infancy. Owing to the extremely high energy demand of the heart, the bioenergetics pathways need to respond efficiently based on substrate availability. The metabolic regulation of such heart bioenergetics is mediated by various rate limiting enzymes involved in energy metabolism. Although all the pertinent mechanisms are not clearly understood, the progressive decline in the activity of metabolic enzymes leading to diminished ATP production is known to cause progression of the heart failure. Therefore, metabolic therapy that can maintain the appropriate activities of metabolic enzymes can be a promising approach for the prevention and treatment of the heart failure. The flavonoids that constitute various human dietary ingredients also effectively offer a variety of health benefits. The flavonoids target a variety of metabolic enzymes and facilitate effective management of the equilibrium between production and utilization of energy in the heart. This review discusses the broad impact of metabolic enzymes in the heart functions and explains how the dysregulated enzyme activity causes the heart failure. In addition, the prospects of targeting dysregulated metabolic enzymes by developing flavonoid-based metabolic approaches are discussed.

Effects of commercial pectolytic and cellulolytic enzyme preparations on the apple cell wall.

PubMed

Dongowski, G; Sembries, S

2001-09-01

The action of three different commercial enzyme combinations on apple cell wall material has been examined in a model system under conditions of mash and pomace treatment by using an alcohol-insoluble substance prepared from apples. A part of the total dietary fiber, for example, galacturonan (pectin), appeared in the soluble fraction after enzymatic mash treatment. The soluble fraction increased intensely during pomace treatment. Furthermore, enzyme actions caused a change in the water-binding capacity of residues as well as changes in the monosaccharide composition and in the molecular weight distribution of saccharides in filtrates (soluble parts). The extent of decomposition of cell wall material and the increase of soluble oligomeric and/or polymeric dietary fiber components are caused by both the composition (pectinases, cellulases, and hemicellulases) and the activities of the enzyme preparations. The model experiments allow an insight into the reactions occurring during enzyme action on the plant cell wall, for example, during apple juice production using pectolytic and cellulolytic enzyme preparations.

The response of single human cells to zero-gravity

NASA Technical Reports Server (NTRS)

Montgomery, P. O., Jr.; Cook, J. E.; Reynolds, R. C.; Paul, J. S.; Hayflick, L.; Stock, D.; Shulz, W. W.; Kimzey, S. L.; Thirolf, R. G.; Rogers, T.

1977-01-01

Microscopic and histochemical evaluations of human embrionic lung cells after exposure to zero-gravity are reported. Growth curves, DNA microspectrophotometry, phase microscopy, and ultrastructural studies of fixed cells revealed no effects on the cultures. Minor unexplained differences have been found in biochemical constituents of the samples.

Histochemical differences along the intestine of Corydoras paleatus (Siluriformes: Callichthyidae).

PubMed

Plaul, Silvia E; Barbeito, Claudio G; Díaz, Alcira O

2016-03-01

The Neotropical catfish Corydoras paleatus is a facultative air-breather and the caudal half of the intestine is involved in gas exchange. In South America, air-breathing fishes are found in tropical or sub-tropical freshwaters where the probability of hypoxia is high. The aim of this study was to characterize by traditional histochemical and lectinhistochemical methods the pattern of carbohydrate in the intestinal mucosa. Intestine samples were taken from 25 healthy adult specimens collected in Buenos Aires (Argentina). Samples were fixed by immersion in 10 % buffered formalin and routinely processed and embedded in paraffin wax. Subsequently, these sections were incubated in the biotinylated lectins battery. Labeled Streptavidin-Biotin (LSAB) system was used for detection, diaminobenzidine as chromogen and haematoxylin as a contrast. To locate and distinguish glycoconjugates (GCs) of the globet cells, we used the following histochemical methods: PAS; PAS*S; KOH/ PA*S; PA/Bh/KOH/PAS; KOH/PA*/Bh/PAS; Alcian Blue and Toluidine Blue at different pHs. Microscopically, the general structure of vertebrate intestine was observed and showed all the cell types characteristic of the intestinal epithelium. The cranial sector of catfish intestine is a site of digestion and absorption and its structure is similar to other fish groups. In contrast, enterocytes of the caudal portion are low cuboidal cells; and between these, globet cells and capillaries are observed, these latter may reach the mucosal lumen. Underlying the epithelium, observed a well-developed lamina propria-submucosa made of connective tissue; this layer was highly vascularized and did not exhibit glands. According to histochemistry, the diverse GCs elaborated and secreted in the intestine are associated with specific functions in relation to their physiological significance, with special reference to their role in lubrication, buffering effect and prevention of proteolytic damage to the epithelium together with other biological processes, such as osmoregulation and ion exchange. The lectinhistochemical analysis of the intestinal mucosa reveals the presence of terminal residues of glucose, mannose and galactose. In conclusion, this study has shown that GCs synthesized in the intestine of C. paleatus exhibit a high level of histochemical complexity and that the lectin binding pattern of the intestinal mucosa is characteristic of each species and the variations are related with the multiple functions performed by the mucus in the digestive tract. The information generated here may be a relevant biological tool for comparing and analyzing the possible glycosidic changes in the intestinal mucus under different conditions, such as changes in diet or different pathological stages.

Evaluation of liver enzyme levels and identification of asymptomatic liver disease patients in primary care.

PubMed

Cacciola, Irene; Scoglio, Riccardo; Alibrandi, Angela; Squadrito, Giovanni; Raimondo, Giovanni

2017-03-01

The evaluation of serum liver enzyme levels is the most used surrogate marker of liver injury in clinical practice. The prevalence and association of abnormal enzyme values with hepatitis B virus (HBV) and hepatitis C virus (HCV) infections, and with other major causes of liver damage (obesity, diabetes, dyslipidemia, and alcohol abuse) were evaluated in individuals attending the surgeries of 14 general practitioners (GPs) working in Messina. Alanine-amino-transferase, aspartate-amino-transferase, and gamma-glutamyl-transpeptidase measurements were measured in 7816 individuals consecutively attending the GP surgeries between January 1, 2011 and June 30, 2012. Five-thousand-eight-hundred-six subjects (74.3 %) had the tests performed, and 1189 of them (20.5 %) showed increased liver enzyme levels. Sixty-nine of these 1189 individuals (5.8 %) were HCV positive and 12 HBV positive (1 %), 755 (63.5 %) were overweight or obese, 288 (24.2 %) had diabetes, and 351 (29.5 %) had dyslipidemia; 262 (22 %) drank >2 alcoholic units/day. Overall, 57 % of individuals with abnormal liver enzymes had multiple possible causes of liver disease, 28 % one cause, and 15 % no apparent cause. In conclusion, this study shows that 1/5 of individuals attending GP surgeries have altered liver biochemistry and that overweight and metabolic disorders have become the major causes of liver damage even in South Italy, where HBV and HCV were endemic in the past century. Notably, many HCV and HBV patients are still unaware of their infected status, and GPs are essential for their timely identification.

Polygalacturonase Causes Lygus-like Damage on Plants: Cloning and Identification of Western Tarnished Plant Bug (Lygus hesperus) Polygalacturonases Secreted During Feeding

USDA-ARS?s Scientific Manuscript database

Abstract: Polygalacturonase (PG), an enzyme that degrades pectin within the plant tissue cell wall, has been postulated as the chemical cause of damage to plants by the mirid Lygus hesperus. Micro-injection of two pure recombinant Aspergillus niger PG II protein forms, the wild type enzymically acti...

Enzymes, detergents and skin: facts and fantasies.

PubMed

Basketter, D A; English, J S C; Wakelin, S H; White, I R

2008-06-01

In their raw state, enzymes of bacterial/fungal origin cause allergic reactions in the lung. Proteolytic enzymes also cause irritation to skin, eyes and the respiratory tract. For 40 years, encapsulated enzymes have been used worldwide in detergent products, especially laundry formulations, and have increasing importance due to biodegradability and functionality at low temperatures, offering environmental benefits. Uniquely to the U.K., for years it has been suggested that the inclusion of enzymes in such products leads to adverse skin reactions, including erythema, pruritus and exacerbation of eczema. In this review, we look at the facts, asking whether there is evidence that the hazards identified for enzymes translate into any risk for consumer health. By considering the actual exposures in consumer use and exaggerated product usage, it is concluded that the irritating and allergenic hazards of enzyme raw materials do not translate into a risk of skin reactions, either irritant or allergic. Investigations of numerous individuals with skin complaints attributed to laundry products demonstrate convincingly that enzymes were not responsible. Indeed, enzyme-containing laundry products have an extensive history of safe use. Thus, the supposed adverse effects of enzymes on skin seem to be a consequence of a mythology. The important practical lesson is that when primary or secondary care practitioners are presented with a skin complaint, it should not be dismissed as a result of using an enzyme-containing laundry product as the diagnosis will certainly lie elsewhere. Education for healthcare professionals could usefully be enhanced to take this on board.

INHIBITION OF THE DEVELOPMENT OF HEPATIC MICROSOMAL DETOXIFICATION ENZYMES BY X-IRRADIATION.

DTIC Science & Technology

of young, male rats, on the activity of these enzymes in the livers of adult animals, and on induced enzyme synthesis by phenobarbital . Exposure of 23...caused by phenobarbital administration. The results of these studies indicate that radiation specifically inhibits the synthesis of increased microsomal

Acute carbon tetrachloride induced lysosomal membrane damage and the membrane protecting effect of a new dihydroquinoline-type antioxidant.

PubMed

Toncsev, H; Pollák, Z; Kiss, A; Sréter, L; Fehér, J

1982-01-01

The authors examined the damage of lysosomal membrane caused by acute CCl4 intoxication by in vitro methods. They measured the acid phosphatase as well as beta-glucuronidase enzyme levels and determined the rate of release of these two enzymes. The in vivo changes in enzyme activity were extrapolated from the in vitro results. The CCl4 causes a significant increase in the permeability and rigidity of the lysosomal membrane. By oral and/or intraperitoneal administration of MTDQ the state of permeability can be improved or even corrected. On the basis of their results, the authors conclude that the lysosomal damage caused by CCl4 is mediated by peroxidation of lipids and the lysosomal membrane can be stabilised by MTDQ.

Phenylketonuria

MedlinePlus

... inherited disorder that causes an amino acid called phenylalanine to build up in the body. PKU is ... helps create the enzyme needed to break down phenylalanine. Without the enzyme necessary to process phenylalanine, a ...

Trafficking of Metastatic Breast Cancer Cells in Bone

DTIC Science & Technology

2006-08-01

Goldwater Award. She graduated in 2004 and is currently a graduate student at the University of Pennsylvania. • Elizabeth Chislock, a senior...carcinoma cells. J Histochem Cytochem 2000;48:643^51. 12. Amhlaoibh RN,Hoegh-Andersen P, Bru « nnerN, et al. Measurement of tumor load and distribution

Muscle fiber characteristics of broiler breast fillets with the wooden breast condition.

USDA-ARS?s Scientific Manuscript database

Broiler breast fillets exhibiting the wooden breast condition are described as having a rigid feel and abnormal texture attributes; however, changes at the muscle fiber level in wooden breast fillets are not well understood. The objective of this study was to compare the histochemical muscle fiber ...

Histochemical Seeing: Scientific Visualization and Art Education

ERIC Educational Resources Information Center

Knochel, Aaron

2013-01-01

What are the capacities of visual arts curricula to engage learning within narrow frameworks of overly "scientistic" standards (Lather, 2007)? With growing emphasis in schools under STEM initiatives and evidence-based standards, the possible cross-pollination of effects that art education may have on a science-centric education may be a…

Activation of immobilized enzymes by acoustic wave resonance oscillation.

PubMed

Nishiyama, Hiroshi; Watanabe, Tomoya; Inoue, Yasunobu

2014-12-01

Acoustic wave resonance oscillation has been used successfully in the development of methods to activate immobilized enzyme catalysts. In this study, resonance oscillation effects were demonstrated for enzyme reactions on galactose oxidase (GAD), D-amino acid oxidase (DAAO), and L-amino acid oxidase (LAAO), all of which were immobilized covalently on a ferroelectric lead zirconate titanate (PZT) device that could generate thickness-extensional resonance oscillations (TERO) of acoustic waves. For galactose oxidation on immobilized GAD in a microreactor, TERO generation immediately increased enzyme activity 2- to 3-fold. Eliminating TERO caused a slight decrease in the activity, with ∼90% of the enhanced activity retained while the reaction proceeded. Contact of the enhanced enzyme with a galactose-free solution caused almost complete reversion of the activity to the original low level before TERO generation, indicating that, not only TERO-induced GAD activation, but also preservation of the increased activity, required a galactose substrate. Similar activity changes with TERO were observed for enzyme reactions on DAAO and LAAO. Kinetic analysis demonstrated that TERO helped strengthen the interactions of the immobilized enzyme with the reactant substrate and promoted formation of an activation complex. Copyright © 2014 Elsevier Inc. All rights reserved.

High pressure, temperature and time-dependent effects on enzymatic and microbial properties of fresh sugarcane juice.

PubMed

Chauhan, O P; Ravi, N; Roopa, N; Kumar, Sumeet; Raju, P S

2017-11-01

Efficacy of variable high pressure, temperature and time on the browning causing enzymes and microbial activities, which are major spoilage factors during preservation of sugarcane juice, was studied. The juice was processed at 200-600 MPa pressure for 2-8 min at 40 and 60 °C and their effect on polyphenol oxidase and peroxidase as well as microbiological quality in terms of total plate count, yeast and molds and total coliforms was studied. Application of high pressures were found to cause significant decrease in enzymatic and microbial activities. The effects were found to be significantly more pronounced at 60 °C as compared to 40 °C. Process time also caused significant ( p  < 0.05) negative effect on microbial and enzyme activities. The sugarcane juice treated at 600 MPa for 6 min at 60 °C was found sufficient to inactivate the microbial counts completely. Whereas, enzymes were found to be completely inactivated in the samples processed at 600 MPa for 8 min at 60 °C. A pressure of 600 MPa at 60 °C for 8 min could be applied during commercial preservation of sugarcane juice for getting complete inactivation of browning causing enzymes and spoilage causing microorganisms.

Regulation of Ubiquitin Enzymes in the TGF-β Pathway.

PubMed

Iyengar, Prasanna Vasudevan

2017-04-20

The transforming growth factor-β (TGF-β) pathway has a tumor suppressor role in normal and premalignant cells but promotes oncogenesis in advanced cancer cells. Components of the pathway are tightly controlled by ubiquitin modifying enzymes and aberrations in these enzymes are frequently observed to dysregulate the pathway causing diseases such as bone disorders, cancer and metastasis. These enzymes and their counterparts are increasingly being tested as druggable targets, and thus a deeper understanding of the enzymes is required. This review summarizes the roles of specific ubiquitin modifying enzymes in the TGF-β pathway and how they are regulated.

Equilibrium softening of an enzyme explored with the DNA spring

NASA Astrophysics Data System (ADS)

Tseng, Chiao-Yu; Zocchi, Giovanni

2014-04-01

We explore enzyme mechanics using a system of two mechanically coupled biomolecules. Measurements of the mechanical modulation of enzymatic activity in a Luciferase—DNA chimera are presented. These are molecules where the enzyme is deformed by the action of a DNA spring. The response of the enzyme for different states of stress is examined. It is found that small changes in the stress cause large changes in activity. This nonlinear behavior is qualitatively interpreted as arising from a soft regime of the enzyme beyond linear elasticity. This soft regime may enable large conformational motion in enzymes.

A male sexually dimorphic trait provides antimicrobials to eggs in blenny fish.

PubMed

Giacomello, Eva; Marchini, Daniela; Rasotto, Maria B

2006-09-22

Predation and microbial infections are the major causes of natural mortality for early life stages of oviparous species. The parental traits reducing the effects of predation are rather well described, whereas antimicrobial mechanisms enhancing offspring survival are largely unexplored. In this paper, we report that a male sexually dimorphic trait, the anal glands, of the redlip blenny (Ophioblennius atlanticus atlanticus) and the peacock blenny (Salaria pavo), two fish species with paternal egg care, produce a mucus enriched with antimicrobial substances. Histological and histochemical analyses showed that the anal glands of these species are characterized by the massive presence of mucus-secreting cells. Anal gland extracts, from both the hydrophilic and the hydrophobic protein fraction, exhibited a lysozyme-like activity. Field observations demonstrated that redlip blenny males, while performing egg care, rub the anal region over the nest internal surface, probably facilitating the transfer of mucus to eggs. These results strongly indicate that this sexually dimorphic trait is involved in egg defence against microbial infections.

[Effect of gamma-aminobutyric acid on peripheral mechanisms regulating autonomic functions].

PubMed

Godovalova, L A

1976-01-01

Experiments with cats ascertained the potentiating action of GABA (100,300,500 mg/kg) on the pressor reactions of the small intestine vessels, the systemic arterial pressure, depressing (100 mg/kg) and facilitating (500 mg/kg) effect upon the reactions of inhibition of the small intestine motor activity evoked by the efferent stimulation of the celiac nerve. Adrenolytics (dihydroergotoxin, inderal) abolished the facilitating effects of GABA. The latter (0.01 solution) inhibited spontaneous contractions of isolated small intestine lengths. As proved histochemically GABA (500 mg/kg) reduces the catecholamines content in the suprarenals, in the solar plexus ganglia and in vessles "in vivo". It also increases the catecholamines content in the small intestine wall in experiments in vivo and reduces in vitro tests. The potentiating action of GABA on the vegetative reactions in efferent stimulation of the ciliac nerve occurs, apparently, due to an increased ejection of catecholamines by suprarenals and lowered the content of catecholamines in the solar plexus ganglia, which causes facilitated conduction of excitation in the ganglia.

[Population characteristics of mucous tissue basocytes in the Mongolian gerbil's jejunum following the 12-day orbital flight onboard space platform "Foton-M3"].

PubMed

Atyakshin, D A; Bykov, E G

2013-01-01

Optical (light) microscopy and histochemical techniques were used for the first-ever studies of the population characteristics of tissue basocytes in the jejunum mucous membrane in three groups of gerbils Meriones unguiculatus: flown over 12 days aboard space platform Foton-M3, subjected to spaceflight factors simulation (SFS) in dedicated system Kontur-L (2) and maintained in standard vivarium conditions (control). Space flight was shown to induce quantitative and qualitative changes in the population of jejunum mucus labrocytes. Reduction of the basocytes population, alterations in age composition and ratio of the morphofunctional cell types in microgravity were indicative of cytoplasmic aggregation intensity, paths of biosynthesis products release into the intersticium, and their tinctorial properties. Also, heparin maturation and liberalization into the extracellular space in support of the jejunum mucus adaptive functions progressed with greater intensity. SFS did not affect size of the basocytes population significantly although it did cause qualitative rearrangements in the population structure.

The effects of antibodies on cells

PubMed Central

Dumonde, D. C.; Bitensky, Lucille; Cunningham, G. J.; Chayen, J.

1965-01-01

Biochemical and histochemical methods were used to study the interaction of antibodies and complement with mouse Ehrlich ascites tumour cells. In the presence of complement, both iso- and hetero-antibodies caused cell lysis with penetration of antibodies into the damaged cells, as detected by immunofluorescence; the cells were then unable to support aerobic glycolysis though they retained their ability to consume oxygen in the presence of succinate. Under these conditions there was unmasking of phospholipid particularly at the cell surface, together with lysosomal changes resulting in diffuse staining for lysosomal acid—phosphatase. In the absence of complement, antibodies did not appear to penetrate the cells which respired normally and were not lysed. However, in these cells there was intense lysosomal activation accompanied by unmasking of cytoplasmic phospholipid; it appeared that an immune reaction confined to the cell surface was able to induce changes in the cytoplasm without acutely impairing the viability of the cell. ImagesFIGS. 1-8FIGS. 9-14 PMID:14245309

The nerve supply of the lumbar intervertebral disc.

PubMed

Edgar, M A

2007-09-01

The anatomical studies, basic to our understanding of lumbar spine innervation through the sinu-vertebral nerves, are reviewed. Research in the 1980s suggested that pain sensation was conducted in part via the sympathetic system. These sensory pathways have now been clarified using sophisticated experimental and histochemical techniques confirming a dual pattern. One route enters the adjacent dorsal root segmentally, whereas the other supply is non-segmental ascending through the paravertebral sympathetic chain with re-entry through the thoracolumbar white rami communicantes. Sensory nerve endings in the degenerative lumbar disc penetrate deep into the disrupted nucleus pulposus, insensitive in the normal lumbar spine. Complex as well as free nerve endings would appear to contribute to pain transmission. The nature and mechanism of discogenic pain is still speculative but there is growing evidence to support a 'visceral pain' hypothesis, unique in the muscloskeletal system. This mechanism is open to 'peripheral sensitisation' and possibly 'central sensitisation' as a potential cause of chronic back pain.

Porphyrins and Porphyria Diagnosis

MedlinePlus

... diagnosis The porphyrias are caused by deficiencies of enzymes of the heme biosynthetic pathway. This pathway, like ... heme. Heme is essential for life, and each enzyme is also essential, because it is responsible for ...

Change in enzyme production by gradually drying culture substrate during solid-state fermentation.

PubMed

Ito, Kazunari; Gomi, Katsuya; Kariyama, Masahiro; Miyake, Tsuyoshi

2015-06-01

The influence of drying the culture substrate during solid-state fermentation on enzyme production was investigated using a non-airflow box. The drying caused a significant increase in enzyme production, while the mycelium content decreased slightly. This suggests that changes in the water content in the substrate during culture affect enzyme production in fungi. Copyright © 2014 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

[Mechanisms of endogenous drug resistance acquisition by spontaneous chromosomal gene mutation].

PubMed

Fukuda, H; Hiramatsu, K

1997-05-01

Endogenous resistance in bacteria is caused by a change or loss of function and generally genetically recessive. However, this type of resistance acquisition are now prevalent in clinical setting. Chromosomal genes that afford endogenous resistance are the genes correlated with the target of the drug, the drug inactivating enzymes, and permeability of the molecules including the antibacterial agents. Endogenous alteration of the drug target are mediated by the spontaneous mutation of their structural gene. This mutation provides much lower affinity of the drugs for the target. Gene expression of the inactivating enzymes, such as class C beta-lactamase, is generally regulated by regulatory genes. Spontaneous mutations in the regulatory genes cause constitutive enzyme production and provides the resistant to the agent which is usually stable for such enzymes. Spontaneous mutation in the structural gene gives the enzyme extra-spectrum substrate specificity, like ESBL (Extra-Spectrum-beta-Lactamase). Expression of structural genes encoding the permeability systems are also regulated by some regulatory genes. The spontaneous mutation of the regulatory genes reduce an amount of porin protein. This mutation causes much lower influx of the drug in the cell. Spontaneous mutation in promoter region of the structural gene of efflux protein was observed. This mutation raised the gene transcription and overproduced efflux protein. This protein progresses the drug efflux from the cell.

Effect of Blockade of Nitric Oxide Synthesis on the Renin Secretory Response to Frusemide in Conscious Rabbits

NASA Technical Reports Server (NTRS)

Reid, Ian A.; Chou, Lance

1995-01-01

The enzyme nitric oxide synthase is present in the macula densa and may participate in the control of renin secretion by the adjacent juxtagiomerular cells. In the present study, we investigated the effect of inhibiting nitric oxide synthase on the renin secretory response to frusemide, which stimulates renin secretion by blocking Na(+)-K(+)-2Cl(-) co-transport in the macula densa. Injection of frusemide in 12 conscious rabbits elicited a transient increase in mean arterial pressure from 84 +/- 2 to 92 +/-3 mm Hg at 5 min (P less than 0.01) and a sustained increase in heart rate from 246 +/- 6 to 281 +/- 10 beats/min at 45 min (P less than 0.01). Plasma renin activity increased from 8.0 +/- 1.2 to 14.3 +/- 1.8, 12.4 +/- 1.6 and 11.6 +/- 1.5 pmol/2h ml at 15, 30 and 45min respectively (P less than 0.01). There were no changes in plasma sodium and potassium concentrations or osmoiality. Inhibition of nitric oxide synthase with N(sup G)-nitro-L- arginine methyl ester increased mean arterial pressure by 9 mm Hg, decreased heart rate and plasma renin activity, and markedly suppressed the renin response to frusemide (from 4.6 +/- 0.7 to 7.6 +/- 1.7, 4.7 +/- 1.0 and 4.6 +/- 0.7pmol/2h ml at 15, 30 and 45 min respectively). By contrast, infusion of an equipressor dose of phenylephrine did not suppress the renin response to frusemide. Histochemical studies with the NADPH diaphorase technique confirmed the presence of nitric oxide synthase in the macula densa, and suggested that enzyme activity is increased by treatment with frusemide. These results are consistent with a role for the L- arginine-nitric oxide pathway in the modulation of renin secretion by the macula densa.

Expression profile of senescence-associated beta-galactosidase and activation of telomerase in human ovarian surface epithelial cells undergoing immortalization.

PubMed

Litaker, J R; Pan, J; Cheung, Y; Zhang, D K; Liu, Y; Wong, S C; Wan, T S; Tsao, S W

1998-11-01

Senescence is a specific physiological stage of cells characterized by long population doubling time. It accounts for the inability of normal somatic cells to undergo indefinite cell division. As the number of population doublings increase, cell cycle regulatory mechanisms come into play and signal cells to exit the cell cycle and become senescent. Senescence has been implicated in the aging process and may function as a tumor suppressor mechanism in human cells. The ability to measure the degree of cellular senescence is important in understanding the biological processes regulating cell aging and immortalization. Senescent cells exhibit an enzyme termed senescence-associated histochemical staining. Cells immortalized by viral oncogenes often enter a stage of crisis at the early phase of immortalization. The cells at crisis have a long population doubling time. Cells at the crisis stage resemble senescent cells and the expression of SA- beta-Gal may be used to monitor the process of immortalization. In this study the expression profile of SA-beta-Gal was examined in human ovarian surface epithelial cells (HOSE 6-3) undergoing immortalization by the human papilloma viral oncogene E6 and E7 (HPV E6 and E7). Our results showed a low percentage (12.0%) of HOSE 6-3 cells expressing SA-beta-Gal activity at the pre-crisis stage. The percentage of HOSE 6-3 cells expressing SA-beta-Gal activity was highest (39.2%) at the crisis stage. When HOSE 6-3 cells achieved immortalized status there was a sharp decrease in cells (1. 3%) expressing SA-beta-Gal activity. In addition, an inverse relationship between the expression of SA-beta-Gal activity and telomerase activity was noted in cells undergoing immortalization. The results confirm that the SA-beta-Gal enzyme is a good marker for monitoring the population of cells undergoing senescence at different stages of immortalization and that telomerase activation is a characteristic feature of post-crisis cells.

Ouabain-sensitive Rb+ uptake in mouse eggs and preimplantation conceptuses

DOE Office of Scientific and Technical Information (OSTI.GOV)

Van Winkle, L.J.; Campione, A.L.

1991-07-01

The results of histochemical and immunocytochemical studies have been used elsewhere to support the hypothesis that Na+/K(+)-ATPase expression is initiated or increases dramatically in preimplantation mouse conceptuses just before they begin to cavitate. Moreover, localization of the enzyme in the inner membrane of the mural trophoblast is thought to be involved directly in formation and maintenance of the blastocyst cavity. Presumably, Na+/K(+)-ATPase extrudes the cation, Na+, and therefore water into the cavity. The cation transporting activity of the enzyme can be determined by measuring ouabain-sensitive Rb+ uptake by cells. Therefore, we measured Rb+ uptake in mouse eggs and preimplantation conceptusesmore » at various stages of development. 86Rb+ uptake by conceptuses increased linearly with time for at least 60 min in medium containing 0.7 mM total Rb+ plus K+ in the absence or presence of 1.0 mM ouabain, and ouabain inhibited more than 70% of 86Rb+ uptake. The ouabain concentration at 1/2 of maximum inhibition of the ouabain-sensitive component of 86Rb+ uptake was about 10-20 microM in eggs and conceptuses at all stages of preimplantation development. Moreover, ouabain-sensitive Rb+ uptake had a twofold higher Vmax value in blastocysts than in eggs or conceptuses at earlier stages of development (i.e., approximately 173 vs 70-100 fmole.conceptus-1.min-1), although the total cell surface area also was probably about two times greater in blastocysts than in eggs or other conceptuses. Ouabain-sensitive Rb+ transport in eggs and conceptuses may have occurred via a single ouabain-sensitive Rb+ transporter with a Hill coefficient of 1.5-1.8 (Hill plots). When it was assumed that the Hill coefficient had a value of 2.0, however, eggs and conceptuses appeared to contain at least two forms of Na+/K(+)-ATPase activity.« less

Interaction between wheat alpha-amylase/trypsin bi-functional inhibitor and mammalian digestive enzymes: Kinetic, equilibrium and structural characterization of binding.

PubMed

Cuccioloni, Massimiliano; Mozzicafreddo, Matteo; Ali, Ishtiaq; Bonfili, Laura; Cecarini, Valentina; Eleuteri, Anna Maria; Angeletti, Mauro

2016-12-15

Alpha-amylase/trypsin bi-functional inhibitors (ATIs) are non-gluten protein components of wheat and other cereals that can hypersensitise the human gastrointestinal tract, eventually causing enteropathies in predisposed individuals. These inhibitory proteins can act both directly by targeting specific pro-inflammatory receptors, and indirectly by impairing the activity of digestive enzymes, the latter event causing the accumulation of undigested peptides with potential immunogenic properties. Herein, according to a concerted approach based on in vitro and in silico methods we characterized kinetics, equilibrium parameters and modes of binding of the complexes formed between wheat ATI and two representative mammalian digestive enzymes, namely trypsin and alpha-amylase. Interestingly, we demonstrated ATI to target both enzymes with independent binding sites and with moderately high affinity. Copyright © 2016 Elsevier Ltd. All rights reserved.

Development of extracellular matrix in chick paravertebral sympathetic ganglia.

PubMed

Luckenbill-Edds, L

1986-08-01

Alcian blue staining coupled with enzyme digestion or critical electrolyte staining revealed differences in the development of extracellular matrix (ECM) within sympathetic ganglia compared with the surrounding capsule. On day 5 of chick development (Hamburger-Hamilton stage 26) only hyaluronic acid (HA) could be detected in the ECM surrounding condensing primary ganglia. By day 7 (st 30) the ganglionic capsule contained HA, as well as sulfated glycosaminoglycans (GAGs), and this pattern continued into the adult stage. During the later stages of embryonic life (st 41-45) satellite cells appear, showing fine structural characteristics that point to their role in the secretion of intraganglionic ECM. Only during these stages could ECM be detected histochemically within ganglia, the same stages (days 15-19) when routine electron microscopic methods reveal collagen fibrils embedded in a granular ground substance. Thus, the intraganglionic environment appears as a separate compartment free of detectable amounts of GAG until late embryonic stages when ECM is secreted around satellite cells. This developmental pattern could represent a role of ECM in the histological stabilization of ganglia during the late stages of differentiation, since the appearance of intraganglionic ECM is correlated with the appearance of small dense-cored vesicles characteristic of adult neurons. The developmental pattern of ECM in differentiating sympathetic ganglia is compared with that of other tissues that undergo condensation and morphogenesis.

Ontogeny and distribution of alkaline and acid phosphatases in the digestive system of California halibut larvae (Paralichthys californicus).

PubMed

Zacarias-Soto, Magali; Barón-Sevilla, Benjamín; Lazo, Juan P

2013-10-01

Studies aimed to assess the digestive physiology of marine fish larvae under culture conditions are important to further understand the functional characteristics and digestive capacities of the developing larvae. Most studies to date concentrate on intestinal lumen digestion and little attention to the absorption process. Thus, the objectives of this study were to histochemically detect and quantify some of the enzymes responsible for absorption and intracellular digestion of nutrients in the anterior and posterior intestine of California halibut larvae. Alkaline and acid phosphatases were detected from the first days post-hatch (dph). Alkaline phosphatase maintained a high level of activity during the first 20 dph in both intestinal regions. Thereafter, a clear intestinal regionalization of the activity was observed with the highest levels occurring in the anterior intestine. Acid phosphatase activity gradually increased in both intestinal regions during development, and a regionalization of the activity was not observed until late in development, once the ocular migration began. Highest levels were observed in the anterior intestine at the end of metamorphosis concomitant with the stomach development. The results from this study show some morphological and physiological changes are occurring during larval development and a clear regionalization of the absorption process as the larvae develops. These ontological changes must be considered in the elaboration of diets according to the digestive capacity of the larvae.

Long-term liver-specific functions of hepatocytes in electrospun chitosan nanofiber scaffolds coated with fibronectin.

PubMed

Rajendran, Divya; Hussain, Ali; Yip, Derek; Parekh, Amit; Shrirao, Anil; Cho, Cheul H

2017-08-01

In this study, a new 3D liver model was developed using biomimetic nanofiber scaffolds and co-culture system consisting of hepatocytes and fibroblasts for the maintenance of long-term liver functions. The chitosan nanofiber scaffolds were fabricated by the electrospinning technique. To enhance cellular adhesion and spreading, the surfaces of the chitosan scaffolds were coated with fibronectin (FN) by adsorption and evaluated for various cell types. Cellular phenotype, protein expression, and liver-specific functions were extensively characterized by immunofluorescent and histochemical stainings, albumin enzyme-linked immunosorbent assay and Cytochrome p450 detoxification assays, and scanning electron microscopy. The electrospun chitosan scaffolds exhibited a highly porous and randomly oriented nanofibrous structure. The FN coating on the surface of the chitosan nanofibers significantly enhanced cell attachment and spreading, as expected, as surface modification with this cell adhesion molecule on the chitosan surface is important for focal adhesion formation and integrin binding. Comparison of hepatocyte mono-cultures and co-cultures in 3D culture systems indicated that the hepatocytes in co-cultures formed colonies and maintained their morphologies and functions for prolonged periods of time. The 3D liver tissue model developed in this study will provide useful tools toward the development of engineered liver tissues for drug screening and tissue engineering applications. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 2119-2128, 2017. © 2017 Wiley Periodicals, Inc.

Targeted transgenic overexpression of mitochondrial thymidine kinase (TK2) alters mitochondrial DNA (mtDNA) and mitochondrial polypeptide abundance: transgenic TK2, mtDNA, and antiretrovirals.

PubMed

Hosseini, Seyed H; Kohler, James J; Haase, Chad P; Tioleco, Nina; Stuart, Tami; Keebaugh, Erin; Ludaway, Tomika; Russ, Rodney; Green, Elgin; Long, Robert; Wang, Liya; Eriksson, Staffan; Lewis, William

2007-03-01

Mitochondrial toxicity limits nucleoside reverse transcriptase inhibitors (NRTIs) for acquired immune deficiency syndrome. NRTI triphosphates, the active moieties, inhibit human immunodeficiency virus reverse transcriptase and eukaryotic mitochondrial DNA polymerase pol-gamma. NRTI phosphorylation seems to correlate with mitochondrial toxicity, but experimental evidence is lacking. Transgenic mice (TGs) with cardiac overexpression of thymidine kinase isoforms (mitochondrial TK2 and cytoplasmic TK1) were used to study NRTI mitochondrial toxicity. Echocardiography and nuclear magnetic resonance imaging defined cardiac performance and structure. TK gene copy and enzyme activity, mitochondrial (mt) DNA and polypeptide abundance, succinate dehydrogenase and cytochrome oxidase histochemistry, and electron microscopy correlated with transgenesis, mitochondrial structure, and biogenesis. Antiretroviral combinations simulated therapy. Untreated hTK1 or TK2 TGs exhibited normal left ventricle mass. In TK2 TGs, cardiac TK2 gene copy doubled, activity increased 300-fold, and mtDNA abundance doubled. Abundance of the 17-kd subunit of complex I, succinate dehydrogenase histochemical activity, and cristae density increased. NRTIs increased left ventricle mass 20% in TK2 TGs. TK activity increased 3 logs in hTK1 TGs, but no cardiac phenotype resulted. NRTIs abrogated functional effects of transgenically increased TK2 activity but had no effect on TK2 mtDNA abundance. Thus, NRTI mitochondrial phosphorylation by TK2 is integral to clinical NRTI mitochondrial toxicity.

Targeted Transgenic Overexpression of Mitochondrial Thymidine Kinase (TK2) Alters Mitochondrial DNA (mtDNA) and Mitochondrial Polypeptide Abundance

PubMed Central

Hosseini, Seyed H.; Kohler, James J.; Haase, Chad P.; Tioleco, Nina; Stuart, Tami; Keebaugh, Erin; Ludaway, Tomika; Russ, Rodney; Green, Elgin; Long, Robert; Wang, Liya; Eriksson, Staffan; Lewis, William

2007-01-01

Mitochondrial toxicity limits nucleoside reverse transcriptase inhibitors (NRTIs) for acquired immune deficiency syndrome. NRTI triphosphates, the active moieties, inhibit human immunodeficiency virus reverse transcriptase and eukaryotic mitochondrial DNA polymerase pol-γ. NRTI phosphorylation seems to correlate with mitochondrial toxicity, but experimental evidence is lacking. Transgenic mice (TGs) with cardiac overexpression of thymidine kinase isoforms (mitochondrial TK2 and cytoplasmic TK1) were used to study NRTI mitochondrial toxicity. Echocardiography and nuclear magnetic resonance imaging defined cardiac performance and structure. TK gene copy and enzyme activity, mitochondrial (mt) DNA and polypeptide abundance, succinate dehydrogenase and cytochrome oxidase histochemistry, and electron microscopy correlated with transgenesis, mitochondrial structure, and biogenesis. Antiretroviral combinations simulated therapy. Untreated hTK1 or TK2 TGs exhibited normal left ventricle mass. In TK2 TGs, cardiac TK2 gene copy doubled, activity increased 300-fold, and mtDNA abundance doubled. Abundance of the 17-kd subunit of complex I, succinate dehydrogenase histochemical activity, and cristae density increased. NRTIs increased left ventricle mass 20% in TK2 TGs. TK activity increased 3 logs in hTK1 TGs, but no cardiac phenotype resulted. NRTIs abrogated functional effects of transgenically increased TK2 activity but had no effect on TK2 mtDNA abundance. Thus, NRTI mitochondrial phosphorylation by TK2 is integral to clinical NRTI mitochondrial toxicity. PMID:17322372

Qualitative and quantitative observations of bone tissue reactions to anodised implants.

PubMed

Sul, Young-Taeg; Johansson, Carina B; Röser, Kerstin; Albrektsson, Tomas

2002-04-01

Research projects focusing on biomaterials related factors; the bulk implant material, the macro-design of the implant and the microsurface roughness are routinely being conducted at our laboratories. In this study, we have investigated the bone tissue reactions to turned commercially pure (c.p.) titanium implants with various thicknesses of the oxide films after 6 weeks of insertion in rabbit bone. The control c.p. titanium implants had an oxide thickness of 17-200 nm while the test implants revealed an oxide thickness between 600 and 1000 nm. Routine histological investigations of the tissue reactions around the implants and enzyme histochemical detections of alkaline and acid phosphatase activities demonstrated similar findings around both the control and test implants. In general, the histomorphometrical parameters (bone to implant contact and newly formed bone) revealed significant quantitative differences between the control and test implants. The test implants demonstrated a greater bone response histomorphometrically than control implants and the osteoconductivity was more pronounced around the test implant surfaces. The parameters that differed between the implant surfaces, i.e. the oxide thickness, the pore size distribution, the porosity and the crystallinity of the surface oxides may represent factors that have an influence on the histomorphometrical results indicated by a stronger bone tissue response to the test implant surfaces, with an oxide thickness of more than 600 nm.

Ultrastructural analysis of testicular tissue and sperm by transmission and scanning electron microscopy.

PubMed

Chemes, Hector E

2013-01-01

Transmission electron microscopy (TEM) studies have provided the basis for an in-depth understanding of the cell biology and normal functioning of the testis and male gametes and have opened the way to characterize the functional role played by specific organelles in spermatogenesis and sperm function. The development of the scanning electron microscope (SEM) extended these boundaries to the recognition of cell and organ surface features and the architectural array of cells and tissues. The merging of immunocytochemical and histochemical approaches with electron microscopy has completed a series of technical improvements that integrate structural and functional features to provide a broad understanding of cell biology in health and disease. With these advances the detailed study of the intricate structural and molecular organization as well as the chemical composition of cellular organelles is now possible. Immunocytochemistry is used to identify proteins or other components and localize them in specific cells or organelles with high specificity and sensitivity, and histochemistry can be used to understand their function (i.e., enzyme activity). When these techniques are used in conjunction with electron microscopy their resolving power is further increased to subcellular levels. In the present chapter we will describe in detail various ultrastructural techniques that are now available for basic or translational research in reproductive biology and reproductive medicine. These include TEM, ultrastructural immunocytochemistry, ultrastructural histochemistry, and SEM.

Synaptic plasticity and gravity: Ultrastructural, biochemical and physico-chemical fundamentals

NASA Astrophysics Data System (ADS)

Rahmann, H.; Slenzka, K.; Körtje, K. H.; Hilbig, R.

On the basis of quantitative disturbances of the swimming behaviour of aquatic vertebrates (``loop-swimming'' in fish and frog larvae) following long-term hyper-g-exposure the question was raised whether or not and to what extent changes in the gravitational vector might influence the CNS at the cellular level. Therefore, by means of histological, histochemical and biochemical analyses the effect of 2-4 x g for 9 days on the gross morphology of the fish brain, and on different neuronal enzymes was investigated. In order to enable a more precise analysis in future-μg-experiments of any gravity-related effects on the neuronal synapses within the gravity-perceptive integration centers differentiated electron-microscopical and electronspectroscopical techniques have been developed to accomplish an ultrastructural localization of calcium, a high-affinity Ca2+-ATPase, creatine kinase and cytochrome oxidase. In hyper-g animals vs. 1-g controls, a reduction of total brain volume (15 %), a decrease in creatine kinase activity (20 %), a local increase in cytochrome oxidase activity, but no differences in Ca2+/Mg2+-ATPase activities were observed. Ultrastructural peculiarities of synaptic contact formation in gravity-related integration centers (Nucleus magnocellularis) were found. These results are discussed on the basis of a direct effect of hyper-gravity not only on the gravity-sensitive neuronal integration centers but possibly also on the physico-chemical properties of the lipid bilayer of neuronal membranes in general.

Hepatopancreatic intoxication of lambda cyhalothrin insecticide on albino rats

PubMed Central

Elhalwagy, Manal EA; Abd-Alrahman, Sherif H; Nahas, AA; Ziada, Reem M; Mohamady, Aziza H

2015-01-01

Background: Despite the known adverse effects of lambda cyhalothrin insecticide, little is known about its hepatopancreatic intoxication effects. The present study was carried out to elucidate sub-chronic effect of Karat 2.5% EC formulation of lambda cyhalothrin on male albino rats. Methods: To explore the effects of exposure to lambda cyhalothrin on rats and its mechanism, low (1/40 of LD50, 5 mg/kg/day) and high dose (1/4 of LD50, 50 mg/kg/day) lambda cyhalothrin were applied to rats via drinking water for 3 months. Blood samples were collected monthly, and the animals were dissected for liver and pancreas’s examination at the end of the experiment. Lambda cyhalothrin administration was associated with the elevation in lipid peroxidation marker, malondialdehyde (MDA), reduction in SH-protein a major marker for antioxidant, as well as basel paraoxonase (PON) in both treated groups throughout the experimental periods. Results: In addition, significant elevations in liver enzymes alanin amino transferase, (ALT), and aspartate amino transferase (AST), as well as plasma acetylcholinesterase (AChE) and glucose level. While, significant reduction in insulin level through the experimental periods. Results of histopathological and histochemical studies showed that lambda cyhalothrin exposure induces liver and pancreatic tissues damage and depletion in glycogen content was pronounced in liver of both treated groups. Conclusions: In conclusion subchronic intoxication with lambda cyhalothrin formulation induced remarkable changes in the examined parameters. PMID:26221269

Hepatopancreatic intoxication of lambda cyhalothrin insecticide on albino rats.

PubMed

Elhalwagy, Manal Ea; Abd-Alrahman, Sherif H; Nahas, A A; Ziada, Reem M; Mohamady, Aziza H

2015-01-01

Despite the known adverse effects of lambda cyhalothrin insecticide, little is known about its hepatopancreatic intoxication effects. The present study was carried out to elucidate sub-chronic effect of Karat 2.5% EC formulation of lambda cyhalothrin on male albino rats. To explore the effects of exposure to lambda cyhalothrin on rats and its mechanism, low (1/40 of LD50, 5 mg/kg/day) and high dose (1/4 of LD50, 50 mg/kg/day) lambda cyhalothrin were applied to rats via drinking water for 3 months. Blood samples were collected monthly, and the animals were dissected for liver and pancreas's examination at the end of the experiment. Lambda cyhalothrin administration was associated with the elevation in lipid peroxidation marker, malondialdehyde (MDA), reduction in SH-protein a major marker for antioxidant, as well as basel paraoxonase (PON) in both treated groups throughout the experimental periods. In addition, significant elevations in liver enzymes alanin amino transferase, (ALT), and aspartate amino transferase (AST), as well as plasma acetylcholinesterase (AChE) and glucose level. While, significant reduction in insulin level through the experimental periods. Results of histopathological and histochemical studies showed that lambda cyhalothrin exposure induces liver and pancreatic tissues damage and depletion in glycogen content was pronounced in liver of both treated groups. In conclusion subchronic intoxication with lambda cyhalothrin formulation induced remarkable changes in the examined parameters.

Postharvest internal browning of pineapple fruit originates at the phloem.

PubMed

Luengwilai, Kietsuda; Beckles, Diane M; Siriphanich, Jingtair

2016-09-01

A typical symptom of postharvest chilling injury (PCI) in pineapple fruit (Ananas comosus (L.) Merr.) is internal browning (IB) near the fruit core. Since vascular bundles (VBs) are localized to this region, it was hypothesized that the VBs might be the site of IB. To test this, the anatomy and histochemistry of VBs during chilling stress in four pineapple cultivars with different levels of sensitivity to PCI were examined. Fruit were stored at 10°C for up to three weeks to stimulate translucency symptoms (TS; the initiation of IB). After three weeks of chilling exposure, the cultivars 'MD2' showed 0%, 'Pattavia' and 'Savee' showed 10-16%, and 'Trad Sri Thong' showed 100% TS and IB symptom. Scanning electron microscopy and in situ histochemical staining techniques that detect enzymes and substrates commonly associated with IB initiation were used in parallel. The TS of pineapple fruit coincided with the collapse of the phloem tissue. The VBs in the tissue where IB was initiated (i.e., the flesh adjacent to the core or F/C) had the highest activity of polyphenol oxidase, hydrogen peroxide, and phenolic compounds. The IB-resistant 'MD2' genotype had fewer VBs, but a greater proportion of sclerenchyma fibers (P<0.05) than did the susceptible 'Trad Sri Thong'. Based on these data, the first report of pineapple IB occurrence in the phloem was proposed. Copyright © 2016 Elsevier GmbH. All rights reserved.

Use of Propolis Hydroalcoholic Extract to Treat Colitis Experimentally Induced in Rats by 2,4,6-Trinitrobenzenesulfonic Acid

PubMed Central

Gonçalves, Cely Cristina Martins; Hernandes, Luzmarina; Bersani-Amado, Ciomar Aparecida; Franco, Selma Lucy; Silva, Joaquim Felipe de Souza

2013-01-01

This study focused on the therapeutic effect of a propolis SLNC 106PI extract on experimental colitis. Wistar adult rats received 0.8 mL rectal dose of one of the following solutions: saline (group S), 20 mg TNBS in 50% ethanol (group TNBS), 20 mg TNBS in 50% ethanol and propolis extract in saline (group TNBS-P), propolis extract in saline (group SP), and 20 mg TNBS in 50% ethanol and 50 mg/kg mesalazine (group TNBS-M). The animals were euthanized 7 or 14 days after the colitis induction. Samples of the distal colon were harvested for the analysis of myeloperoxidase (MPO) enzyme activity and for morphometric analysis in paraffin-embedded histological sections with hematoxylin-eosin or histochemical staining. The animals treated with TNBS exhibited the typical clinical signs of colitis. Increased MPO activity confirmed the presence of inflammation. TNBS induced the development of megacolon, ulceration, transmural inflammatory infiltrate, and thickened bowel walls. Treatment with propolis moderately reduced the inflammatory response, decreased the number of cysts and abscesses, inhibited epithelial proliferation, and increased the number of goblet cells. The anti-inflammatory activity of the propolis SLNC 106 extract was confirmed by the reductions in both the inflammatory infiltrate and the number of cysts and abscesses in the colon mucosa. PMID:24101941

Histochemical study of lectin binding sites in fourth and fifth instar gypsy moth larval midgut epithelium

Treesearch

Algimantas P. Valaitis

2011-01-01

There is evidence that the gypsy moth, Lymantria dispar, midgut epithelial brush border membrane has membrane-bound glycoconjugates, such as BTR-270 and aminopeptidase N (APN), which function as high affinity binding sites (receptors) for the insecticidal proteins produced by Bacillus thuringiensis (Bt). As gypsy...

CHX14 is a plasma membrane K-efflux transporter that regulates K+ redistribution in "Arabidopsis thaliana"

USDA-ARS?s Scientific Manuscript database

Potassium (K(+)) is essential for plant growth and development, yet the molecular identity of many K(+) transporters remains elusive. Here we characterized cation/H(+) exchanger (CHX) 14 as a plasma membrane K(+) transporter. "CHX14" expression was induced by elevated K(+) and histochemical analysis...

Pancreatic Neuroendocrine Tumor with Atypical Radiologic Presentation.

PubMed

Singh, Ramandeep; Calhoun, Sean; Shin, Minchul; Katz, Robert

2008-01-01

An atypical radiographic presentation of a rare non-functional pancreatic neuroendocrine tumor as seen on US, CT and MRI is described. Radiographic-pathologic correlation via gross autopsy specimens and immuno-histochemical staining demonstrates the pancreas to be markedly enlarged with extensive calcifications and numerous tiny cysts secondary to diffuse neoplastic infiltration without a focal mass.

Colour in digital pathology: a review.

PubMed

Clarke, Emily L; Treanor, Darren

2017-01-01

Colour is central to the practice of pathology because of the use of coloured histochemical and immunohistochemical stains to visualize tissue features. Our reliance upon histochemical stains and light microscopy has evolved alongside a wide variation in slide colour, with little investigation into the implications of colour variation. However, the introduction of the digital microscope and whole-slide imaging has highlighted the need for further understanding and control of colour. This is because the digitization process itself introduces further colour variation which may affect diagnosis, and image analysis algorithms often use colour or intensity measures to detect or measure tissue features. The US Food and Drug Administration have released recent guidance stating the need to develop a method of controlling colour reproduction throughout the digitization process in whole-slide imaging for primary diagnostic use. This comprehensive review introduces applied basic colour physics and colour interpretation by the human visual system, before discussing the importance of colour in pathology. The process of colour calibration and its application to pathology are also included, as well as a summary of the current guidelines and recommendations regarding colour in digital pathology. © 2016 John Wiley & Sons Ltd.

A histochemical study of the distribution of lectin binding sites in the developing branchial area of the trout Salmo trutta.

PubMed Central

Rojo, M C; Blánquez, M J; González, M E

1996-01-01

A histochemical study of the branchial area of brown trout embryos from 35 to 71 d of incubation is reported. A battery of 6 different horseradish peroxidase-labelled lectins, the PAS reaction and Alcian blue staining were used to study the distribution of carbohydrate residues in glycoconjugates along the pharyngeal and branchial epithelia. Con A and WGA reacted at every site of the branchial region thus showing the ubiquitous presence of alpha-D-mannose and N-acetyl-D-glucosamine. WGA, DBA and SBA were good markers for the hatching gland cells (HGCs) and mucous cells. Other lectins, such as PNA and UEA I, reacted only for a short time at some sites during the considered period of incubation. From 35 d until posthatching stages, a manifest strong reaction was noted both in the dorsal epithelium of branchial arches and the HGCs as shown by SBA reactivity. This may be significant with regard to the controversial origin of HGCs, which is thought to be endodermal. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 PMID:8982837

Structural determination and histochemical localization of ghrelin in the red-eared slider turtle, Trachemys scripta elegans.

PubMed

Kaiya, Hiroyuki; Sakata, Ichiro; Kojima, Masayasu; Hosoda, Hiroshi; Sakai, Takafumi; Kangawa, Kenji

2004-08-01

We purified ghrelin peptide and determined the cDNA sequence encoding the precursor protein from the stomach of the red-eared slider turtle, Trachemys scripta elegans. The Trachemys ghrelin is comprised of 25-amino acids and has the sequence GSSFLSPEYQNTQQRKDPKKHTKLN. The third serine residue was modified by n-octanoic (C8:0), decanoic (C10:0) or unsaturated decanoic acid (C10:1). The carboxyl-terminal end of the peptide was not amidated, as seen in the ghrelins of other land vertebrates. Quantitative real-time PCR analysis revealed high levels of gene expression in the stomach and moderate levels in the large intestine and pancreas. Histochemical studies of turtle stomach revealed that ghrelin-immunopositive (ghrelin-ip) cells, which were small and round, were observed in the mucosal layer of the stomach but not in the myenteric plexus, and ghrelin-mRNA-expressing (ghrelin-ex) cells detected by in situ hybridization were scattered in a similar distribution as ghrelin-ip cells. These results indicate that ghrelin is present in reptiles.

Cytological and histochemical gradients on two Copaifera langsdorffii Desf. (Fabaceae)--Cecidomyiidae gall systems.

PubMed

de Oliveira, Denis Coelho; Carneiro, Renê Gonçalves da Silva; Magalhães, Thiago Alves; Isaias, Rosy Mary dos Santos

2011-10-01

Previous ultrastructural and histochemical analysis proposed patterns in the accumulation of substances in galls of Diptera: Cecidomyiidae in some plant species of the temperate region. Similar analyses were done to verify the conservativeness of these patterns in the Neotropical region, where a great number of species of Cecidomyiidae is responsible for a wide diversity of morphotypes. Two gall morphotypes induced by Cecidomyiidae in a unique host plant, Copaifera langsdorffii, were studied. The gradients of carbohydrates and the activity of invertases and acid phosphatases were similar, but the cytological gradients and distribution of proteins evidenced that the sites of the induction as well as the amount of neoformed tissues may be peculiar to each gall system. The production of lipids just in the secretory cavities either in the non-galled or galled tissues indicated a potentiality of the host plant which could not be manipulated by the galling insects. Further, the absence of nucleus in the nutritive tissue, an exclusive feature of the horn-shaped galls, indicates cell death attributed to the feeding habit of the galling herbivore.

Preventing False Negatives for Histochemical Detection of Phenolics and Lignins in PEG-Embedded Plant Tissues

PubMed Central

Ferreira, Bruno G.; Falcioni, Renan; Guedes, Lubia M.; Avritzer, Sofia C.; Antunes, Werner C.; Souza, Luiz A.; Isaias, Rosy M.S.

2016-01-01

Polyethylene glycol (PEG) is a low-cost and advantageous embedding medium, which maintains the majority of cell contents unaltered during the embedding process. Some hard or complex plant materials are better embedded in PEG than in other usual embedding media. However, the histochemical tests for phenolics and lignins in PEG-embedded plant tissues commonly result in false negatives. We hypothesize that these false negatives should be prevented by the use of distinct fixatives, which should avoid the bonds between PEG and phenols. Novel protocols for phenolics and flavanols detection are efficiently tested, with fixation of the samples in ferrous sulfate and formalin or in caffeine and sodium benzoate, respectively. The differentiation of lignin types is possible in safranin-stained sections observed under fluorescence. The Maule’s test faultlessly distinguishes syringyl-rich from guaiacyl- and hydroxyphenyl-rich lignins in PEG-embedded material under light microscopy. Current hypothesis is corroborated, that is, the adequate fixation solves the false-negative results, and the new proposed protocols fill up some gaps on the detection of phenolics and lignins. PMID:28117630

Automated image analysis for quantification of reactive oxygen species in plant leaves.

PubMed

Sekulska-Nalewajko, Joanna; Gocławski, Jarosław; Chojak-Koźniewska, Joanna; Kuźniak, Elżbieta

2016-10-15

The paper presents an image processing method for the quantitative assessment of ROS accumulation areas in leaves stained with DAB or NBT for H 2 O 2 and O 2 - detection, respectively. Three types of images determined by the combination of staining method and background color are considered. The method is based on the principle of supervised machine learning with manually labeled image patterns used for training. The method's algorithm is developed as a JavaScript macro in the public domain Fiji (ImageJ) environment. It allows to select the stained regions of ROS-mediated histochemical reactions, subsequently fractionated according to the weak, medium and intense staining intensity and thus ROS accumulation. It also evaluates total leaf blade area. The precision of ROS accumulation area detection is validated by the Dice Similarity Coefficient in the case of manual patterns. The proposed framework reduces the computation complexity, once prepared, requires less image processing expertise than the competitive methods and represents a routine quantitative imaging assay for a general histochemical image classification. Copyright © 2016 Elsevier Inc. All rights reserved.

The influence of hollyhock extract administration on testicular function in rats.

PubMed

Papiez, Monika A

2004-11-01

It has been reported, recently that an aqueous extract from hollyhock flowers (Althaea rosea Cav. varietas nigra) induces weak metabolic changes in rat testes. In the present study, the in vivoinfluence of a methanolic extract was investigated on the metabolism and morphology of the rat testis. To this end, histochemical, morphometric and radioimmunological methods were used. The rats drank the extract at a dose of 100 mg/day for 7 weeks. The histochemical activities of glucose-6-phosphate dehydrogenase (G6PDH) and Delta(5)beta-hydroxysteroid dehydrogenase (Delta(5)betaHSD) increased significantly statistically in the Leydig cells of the experimental rats in comparison with controls. There were no significant changes in either the diameter of seminiferous tubules or the height of seminiferous epithelium after hollyhock administration. Further, only a small amount of hyperplasia of the interstitial tissue was observed. The morphological and histoenzymatic changes in the Leydig cells indicate that the methanolic hollyhock extract has a direct but small influence on rat testes. The insignificant changes in testicular testosterone and estradiol content suggest that the extract does not disturb steroidogenesis.

Evaluation of Dogs with Border Collie Collapse, Including Response to Two Standardized Strenuous Exercise Protocols.

PubMed

Taylor, Susan; Shmon, Cindy; Su, Lillian; Epp, Tasha; Minor, Katie; Mickelson, James; Patterson, Edward; Shelton, G Diane

2016-01-01

Clinical and metabolic variables were evaluated in 13 dogs with border collie collapse (BCC) before, during, and following completion of standardized strenuous exercise protocols. Six dogs participated in a ball-retrieving protocol, and seven dogs participated in a sheep-herding protocol. Findings were compared with 16 normal border collies participating in the same exercise protocols (11 retrieving, five herding). Twelve dogs with BCC developed abnormal mentation and/or an abnormal gait during evaluation. All dogs had post-exercise elevations in rectal temperature, pulse rate, arterial blood pH, PaO2, and lactate, and decreased PaCO2 and bicarbonate, as expected with strenuous exercise, but there were no significant differences between BCC dogs and normal dogs. Electrocardiography demonstrated sinus tachycardia in all dogs following exercise. Needle electromyography was normal, and evaluation of muscle biopsy cryosections using a standard panel of histochemical stains and reactions did not reveal a reason for collapse in 10 dogs with BCC in which these tests were performed. Genetic testing excluded the dynamin-1 related exercise-induced collapse mutation and the V547A malignant hyperthermia mutation as the cause of BCC. Common reasons for exercise intolerance were eliminated. Although a genetic basis is suspected, the cause of collapse in BCC was not determined.

Collagenous mucosal inflammatory diseases of the gastrointestinal tract.

PubMed

Freeman, Hugh J

2005-07-01

Collagenous mucosal inflammatory diseases involve the columnar-lined gastric and intestinal mucosa and have become recognized increasingly as a significant cause of symptomatic morbidity, particularly in middle-aged and elderly women, especially with watery diarrhea. Still, mechanisms involved in the pathogenesis of this diarrhea remain poorly understood and require further elucidation. The prognosis and long-term outcome of these disorders has been documented only to a limited extent. Recent clinical and pathologic studies have indicated that collagenous mucosal inflammatory disease is a more extensive pathologic process that concomitantly may involve several sites in the gastric and intestinal mucosa. The dominant pathologic lesion is a distinct subepithelial hyaline-like deposit that has histochemical and ultrastructural features of collagen overlying a microscopically defined inflammatory process. An intimate relationship with other autoimmune connective tissue disorders is evident, particularly celiac disease. This is intriguing because these collagenous disorders have not been shown to be gluten dependent. Collagenous mucosal inflammatory disorders may represent a relatively unique but generalized inflammatory response to a multitude of causes, including celiac disease, along with a diverse group of pharmacologic agents. Some recent reports have documented treatment success but histopathologic reversal has been more difficult to substantiate owing to the focal, sometimes extensive nature, of this pathologic process.

Comparison of the larynx-associated lymphoid tissue in the false vocal cords and subglottis in pediatric autopsies.

PubMed

Rossi e Silva, Renata Calciolari; Olegário, Janainna Grazielle Pacheco; Abate, Débora Tavares de Rezende e Silva; Salge, Ana Karina Marques; Peres, Luiz César; Corrêa, Rosana Rosa Miranda; Castro, Eumenia Costa da Cunha; Teixeira, Vicente de Paula Antunes

2012-12-15

The aim this work was to compare the distribution of cellular phenotypes of the LF in the FVC to the ones in the subglottic region in pediatric autopsy, relating this distribution to age and different causes of death. We analyzed 60 larynges of newborns and children autopsied in the period from 1993 to 2003. The fragments were prepared in order to perform histochemical and immunohistochemical techniques. The morphological analysis showed cases that presented LF only in FVC (35%), LF only in the subglottic region (20%), lack of LF in FVC (30%) and lymphoid aggregates, which did not characterize an LF (15%). The cases of LF in the subglottic region were significantly younger compared to the ones that presented LF in the FVC (p=0.017). The LF in the subglottic region was bigger than the LF in the FVC (p=0.020). There was no significant difference between the cause of death and cellular phenotype for both FVC and the subglottic region. In conclusion, the cells that make up the LF in the FVC in newborns and children younger than one year have functional characteristics similar to LF cells in the subglottic region, suggesting that there are similarities with LALT. Copyright © 2012 Elsevier GmbH. All rights reserved.

Dementia in a retired world boxing champion: case report and literature review.

PubMed

Nowak, L A; Smith, G G; Reyes, P F

2009-01-01

Dementia in retired boxers, also referred to as "dementia pugilistica" (DP), is usually attributed to repeated concussive and subconcussive blows to the head. We report the case of a former world boxing champion whose progressive cognitive decline could be ascribed to DP, cerebral infarcts and Wernicke-Korsakoff syndrome. This case demonstrates that dementia in retired boxers may be caused and/or exacerbated by etiologic factors other than DP. We correlated the clinical features with the histochemical and immunohistochemical changes observed on autopsy brain material from a retired boxer, reviewed the literature on boxing-related dementia, and compared our findings with previous reports on DP. Neuropathologic examination revealed numerous neurofibrillary tangles (NFTs), rare neuritic plaques (NPs), multiple cerebral infarcts, fenestrated septum pellucidum, atrophic and gliotic mamillary bodies, and pale substantia nigra and locus ceruleus. Our neuropathologic data confirmed the notion that dementia in retired boxers could be due to several factors such as DP, multiple cerebral infarcts and Wernicke-Korsakoff syndrome. Our findings illustrate the need to comprehensively examine former boxers with dementia as well as carefully evaluate the neuropathologic changes that may cause or contribute to the patient's cognitive and behavioral symptoms. Such an approach is crucial in order to provide prompt and more definitive therapies.

Human immunodeficiency virus-infected macrophages produce soluble factors that cause histological and neurochemical alterations in cultured human brains.

PubMed Central

Pulliam, L; Herndier, B G; Tang, N M; McGrath, M S

1991-01-01

We wanted to establish an in vitro human model for AIDS-associated dementia and pursue the hypothesis that this disease process may be a result of soluble factors produced by HIV-infected macrophages. Human brain aggregates were prepared from nine different brain specimens, and were treated with supernatants from in vitro HIV-infected macrophages (SI), uninfected macrophages (SU), infected T cells, or macrophage-conditioned media from four AIDS patients. Seven of nine treated brains exposed to SI showed peripheral rarefaction after 1 wk of incubation that by ultrastructural analysis showed cytoplasmic vacuolation. Aggregates from two of three brain cultures treated with SI for 3 wk became smaller, an approximately 50% decrease in size. The degree of apparent toxicity in brains exposed to patient-derived macrophage supernatants paralleled the proportion of macrophages found to be expressing HIV p24. Ultrastructural abnormalities were not observed in brains treated with supernatants from HIV-infected T cells, uninfected macrophages, or LPS-activated macrophages. Levels of five neurotransmitter amino acids were decreased in comparison to the structural amino acid leucine. These findings suggest that HIV-infected macrophages, infected both in vitro as well as derived from AIDS patients' peripheral blood, produce factors that cause reproducible histochemical, ultrastructural, and functional abnormalities in human brain aggregates. Images PMID:1671392

Elevation of Glucose 6-Phosphate Dehydrogenase Activity Induced by Amplified Insulin Response in Low Glutathione Levels in Rat Liver

PubMed Central

Taniguchi, Misako; Mori, Nobuko; Iramina, Chizuru

2016-01-01

Weanling male Wistar rats were fed on a 10% soybean protein isolate (SPI) diet for 3 weeks with or without supplementing 0.3% sulfur-containing amino acids (SAA; methionine or cystine) to examine relationship between glutathione (GSH) levels and activities of NADPH-producing enzymes, glucose 6-phosphate dehydrogenase (G6PD) and malic enzyme (ME), in the liver. Of rats on the 10% SPI diet, GSH levels were lower and the enzyme activities were higher than of those fed on an SAA-supplemented diet. Despite the lower GSH level, γ-glutamylcysteine synthetase (γ-GCS) activity was higher in the 10% SPI group than other groups. Examination of mRNAs of G6PD and ME suggested that the GSH-suppressing effect on enzyme induction occurred prior to and/or at transcriptional levels. Gel electrophoresis of G6PD indicated that low GSH status caused a decrease in reduced form and an increase in oxidized form of the enzyme, suggesting an accelerated turnover rate of the enzyme. In primary cultured hepatocytes, insulin response to induce G6PD activity was augmented in low GSH levels manipulated in the presence of buthionine sulfoximine. These findings indicated that elevation of the G6PD activity in low GSH levels was caused by amplified insulin response for expression of the enzyme and accelerated turnover rate of the enzyme molecule. PMID:27597985

Sialomucins are characteristically O-acylated in poorly differentiated and colloid prostatic adenocarcinomas.

PubMed

Sáez, C; Japón, M A; Conde, A F; Poveda, M A; Luna-Moré, S; Segura, D I

1998-12-01

Mucinous glycoproteins are secreted by prostatic adenocarcinomas and might play important roles in tumor invasion and metastasis. Their histochemical properties on routine biopsy specimens have not been fully characterized. We present a histochemical study of mucin in 21 prostatic adenocarcinomas, with particular focus on the demonstration of different types of sialomucins. We applied the following histochemical techniques to routinely processed, formalin-fixed, paraffin-embedded tissue sections: Alcian blue (pH 2.5) and periodic acid-Schiff to reveal both acidic and neutral mucins; high iron diamine and Alcian blue (pH 2.5) to show sulfated and acidic nonsulfated mucosubstances simultaneously; periodic acid borohydride, potassium hydroxide, and periodic acid-Schiff to demonstrate O-acylated sialic acids; periodic acid thionine-Schiff, potassium hydroxide, and periodic acid-Schiff to differentiate pre-existing glycols from those revealed after saponification procedures; and periodic acid borohydride and periodic acid-Schiff to show C9-O-acylated sialic acid. These techniques are useful tools for demonstrating neutral and acidic (sialo- and sulfo-) mucins and di(C8,C9- or C7,C9-)-O-acylated, tri(C7,C8,C9-)-O-acylated and mono(C9)-O-acylated sialomucins. Most prostatic adenocarcinomas showed acidic mucins, with sialomucins predominating over sulfomucins. Well-differentiated and moderately differentiated noncolloid tumors had non-O-acylated sialomucins. Poorly differentiated tumors contained mono-O-acylated (C9) sialomucins, and colloid-type tumors secreted mono-, di-, and tri-O-acylated sialoglycoproteins. Acidic mucins, mainly sialomucins, constitute the major secretory component in prostatic adenocarcinomas, and our results show that the O-acylation of these sialoglycoproteins inversely correlates with tumor differentiation. Well-differentiated and moderately differentiated tumors are not O-acylated, whereas the poorly differentiated ones characteristically have O-acylated sialomucins in C9. Adenocarcinomas of the colloid type, thought to bear a poor prognosis, are the most heavily O-acylated.

Postnatal development of autonomic and sensory innervation of thoracic hairy skin in the rat. A histochemical, immunocytochemical, and radioenzymatic study.

PubMed

Schotzinger, R J; Landis, S C

1990-05-01

Histochemical, immunocytochemical, and radioenzymatic techniques were used to examine the neurotransmitter-related properties of the innervation of thoracic hairy skin in rats during adulthood and postnatal development. In the adult, catecholamine-containing fibers were associated with blood vessels and piloerector muscles, and ran in nerve bundles throughout the dermis. The distribution of tyrosine hydroxylase (TH)-immunoreactive (IR) fibers was identical. Neuronal fibers displaying neuropeptide Y (NPY) immunoreactivity were seen in association with blood vessels. Double-labeling studies suggested that most, if not all, NPY-IR fibers were also TH-IR and likewise most, if not all, vessel-associated TH-IR fibers were also NPY-IR. Calcitonin gene-related peptide (CGRP)-IR fibers were observed near and penetrating into the epidermis, in close association with hair follicles and blood vessels, and in nerve bundles. A similar distribution of substance P (SP)-IR fibers was evident. In adult animals treated as neonates with the sympathetic neurotoxin 6-hydroxydopamine, a virtual absence of TH-IR and NPY-IR fibers was observed, whereas the distribution of CGRP-IR and SP-IR fibers appeared unaltered. During postnatal development, a generalized increase in the number, fluorescence intensity, and varicose morphology of neuronal fibers displaying catecholamine fluorescence, NPY-IR, CGRP-IR, and SP-IR was observed. By postnatal day 21, the distribution of the above fibers had reached essentially adult levels, although the density of epidermal-associated CGRP-IR and SP-IR fibers was significantly greater than in the adult. The following were not evident in thoracic hairy skin at any timepoint examined: choline acetyltransferase activity, acetylcholinesterase histochemical staining or immunoreactivity, fibers displaying immunoreactivity to vasoactive intestinal peptide, cholecystokinin, or leucine-enkephalin. The present study demonstrates that the thoracic hairy skin in developing and adult rats receives an abundant sympathetic catecholaminergic and sensory innervation, but not a cholinergic innervation.

Diffusion and clearance of superparamagnetic iron oxide nanoparticles infused into the rat striatum studied by MRI and histochemical techniques

NASA Astrophysics Data System (ADS)

Wang, F. H.; Kim, D. K.; Yoshitake, T.; Johansson, S. M.; Bjelke, B.; Muhammed, M.; Kehr, J.

2011-01-01

The purpose of the present study was to investigate, by MRI and histochemical techniques, the diffusion and clearance abilities of superparamagnetic iron oxide nanoparticles (SPION) coated with dextran (Dextran-SPION) and gold (Au-SPION) following their local infusions into the rat brain. In separate groups of anesthetized rats, the Dextran-SPION and Au-SPION were infused at concentrations of 0.01, 0.1, 1 and 5 µg Fe/0.5 µl and at the flow rate of 0.5 µl min - 1 into the left and right striata, respectively. Repetitive T2-weighted spin-echo MRI scans were performed at time intervals of 1, 6, 12, 24, 48, 72 h, and one, two and eight weeks after inoculation. Following infusion of Dextran-SPION (0.1 µg and 1 µg Fe), the maximal distribution volume was observed at about 12-24 h after inoculation and two weeks later the Fe signals were undetectable for the lower dose. On the other hand, Au-SPION remained tightly localized in the closest vicinity of the infusion site as revealed by unchanged MRI signal intensities and strong histochemical staining of Fe2 + and Fe3 + ions in the corresponding brain slices. Immunohistochemical staining of astrocytic and microglial reactions revealed that there were no marked differences in GFAP, VIM or OX-42 labeling observed between the nanoparticle types, however the astrocytic reaction was more pronounced in rats receiving nanoparticles compared to the control (aCSF-infused) rats. In conclusion, the present data demonstrate that the viral-sized Dextran-SPION were able to diffuse freely through the interstitial space of the brain being progressively cleared out from the infusion site within two weeks. Thus, Dextran-SPION could be beneficially used in MRI-guided diagnostic applications such as in experimental oncology or as labels and carriers for targeted drug delivery, whereas Au-SPION could be used for labeling and tracking the transplanted stem cells in experimental MRI.

Occupational asthma caused by cellulase and lipase in the detergent industry.

PubMed

Brant, A; Hole, A; Cannon, J; Helm, J; Swales, C; Welch, J; Taylor, A Newman; Cullinan, P

2004-09-01

Three employees from two different detergent companies were investigated for occupational asthma, using skin prick tests, serum specific IgE, and specific bronchial challenge. Two were challenged with lipase and one with cellulase. All three cases had immunological evidence of sensitisation to the detergent enzymes with which they worked. Bronchial challenge in each provoked a reproducible dual asthmatic response, which reproduced their work related symptoms. These are the first reported cases of occupational asthma attributable to cellulase and lipase in the detergent industry. Four of the most common enzymes used in this industry have now been reported to cause occupational asthma; continued vigilance and caution are needed when working with these or other enzymes.

Genetics Home Reference: NGLY1-congenital disorder of deglycosylation

MedlinePlus

... gene. The enzyme produced from this gene, called N -glycanase 1, helps cells get rid of abnormal ... that cause NGLY1 -CDDG impair production of the N -glycanase 1 enzyme, resulting in a severe reduction ...

Slow domain reconfiguration causes power-law kinetics in a two-state enzyme.

PubMed

Grossman-Haham, Iris; Rosenblum, Gabriel; Namani, Trishool; Hofmann, Hagen

2018-01-16

Protein dynamics are typically captured well by rate equations that predict exponential decays for two-state reactions. Here, we describe a remarkable exception. The electron-transfer enzyme quiescin sulfhydryl oxidase (QSOX), a natural fusion of two functionally distinct domains, switches between open- and closed-domain arrangements with apparent power-law kinetics. Using single-molecule FRET experiments on time scales from nanoseconds to milliseconds, we show that the unusual open-close kinetics results from slow sampling of an ensemble of disordered domain orientations. While substrate accelerates the kinetics, thus suggesting a substrate-induced switch to an alternative free energy landscape of the enzyme, the power-law behavior is also preserved upon electron load. Our results show that the slow sampling of open conformers is caused by a variety of interdomain interactions that imply a rugged free energy landscape, thus providing a generic mechanism for dynamic disorder in multidomain enzymes.

Network Architecture Predisposes an Enzyme to Either Pharmacologic or Genetic Targeting.

PubMed

Jensen, Karin J; Moyer, Christian B; Janes, Kevin A

2016-02-24

Chemical inhibition and genetic knockdown of enzymes are not equivalent in cells, but network-level mechanisms that cause discrepancies between knockdown and inhibitor perturbations are not understood. Here we report that enzymes regulated by negative feedback are robust to knockdown but susceptible to inhibition. Using the Raf-MEK-ERK kinase cascade as a model system, we find that ERK activation is resistant to genetic knockdown of MEK but susceptible to a comparable degree of chemical MEK inhibition. We demonstrate that negative feedback from ERK to Raf causes this knockdown-versus-inhibitor discrepancy in vivo. Exhaustive mathematical modeling of three-tiered enzyme cascades suggests that this result is general: negative autoregulation or feedback favors inhibitor potency, whereas positive autoregulation or feedback favors knockdown potency. Our findings provide a rationale for selecting pharmacologic versus genetic perturbations in vivo and point out the dangers of using knockdown approaches in search of drug targets.

[The eye and nutrition].

PubMed

Amemiya, T

1999-12-01

To examine the effect of vitamins and trace elements on ocular tissue. Rats or mice were fed diets deficient in the trace elements Zn, Cu, Mn, Se, Mg, and Cr or in vitamins A, B12, C, and E. In some rats Al and vitamin A were injected in excessive amounts. We studied the conjunctiva, cornea, retina, and optic nerve with a light microscope, transmission and scanning electron microscopes, an energy dispersive X-ray analyser, and an ion microscope. Histochemical, cytochemical, and immunohistochemical techniques were applied to the pathological specimens. Deficiencies of Zn, Cu, Mn, and vitamins A, C and E caused a loss of goblet cells in the conjunctiva and a prominent decrease of microvilli and microplicae in the conjunctiva and cornea. The elements in the goblet cells were changed in these conditions. In addition, epithelial cells showed poor fibrous development and abnormal distribution of chromatin in the nucleus. Zn, Cu, Mn, and vitamins A and E deficiencies caused photoreceptor cells to degenerate and disappear. Se deficiency reduced the horizontal and amacrine cells. Vitamin B12 deficiency reduced nerve fibers in the nerve fiber layer of the retina. Mg deficiency induced multifocal necrosis in the retinal pigment epithelium and apoptotic nuclear changes in the photoreceptor cells. Cr deficiency showed abnormal phagocytosis of the photoreceptor outer segment discs in the retinal pigment epithelium. Vitamin B12 was found to be related to the circadian rhythm in the retina. Deficiencies of Zn, Cu, Mn, and vitamins A, B12, and E induced degeneration and disappearance of myelin lamellae in the myelinated optic nerve fibers. In hypervitaminosis A, lipid droplets appeared in the retinal pigment epithelium and alcohol dehydrogenase disappeared in the retinal pigment epithelium and photoreceptor outer segments. Excessive Al was toxic to the retina, which showed disappearance of photoreceptor cells. Al deposits were seen in dendrites and neurons in the outer plexiform layer. Zn seemed to be necessary for corneal epithelial cell wound healing. Trace elements usually are contained in enzymes, which have many metabolic functions. They are related to synthesis and breakdown of many substances. Some trace elements such as Zn, Cu, Mn, and Se and vitamins including vitamins A, C, and E prevent peroxidation of lipids. Some vitamins have an affinity for specific tissues such as epithelial cells, nerve fibers, and neuronal cells and are needed for cell differentiation, development, and maintenance. Cu, Zn, Mn, Se, Mg, and Cr and vitamins A, B12, C, and E are necessary for maintenance of cellular structure and metabolism.

The eye and nutrition

PubMed

Amemiya

2000-05-01

Purpose: To examine the effect of vitamins and trace elements on ocular tissue.Materials and Methods: Rats or mice were fed diets deficient in the trace elements Zn, Cu, Mn, Se, Mg, and Cr or in vitamins A, B(12), C, and E. In some rats Al and vitamin A were injected in excessive amounts. We studied the conjunctiva, cornea, retina, and optic nerve with a light microscope, transmission and scanning electron microscopes, an energy dispersive X-ray analyzer, and an ion microscope. Histochemical, cytochemical, and immunohistochemical techniques were applied to the pathological specimens.Results: Deficiencies of Zn, Cu, Mn, and vitamins A, C and E caused a loss of goblet cells in the conjunctiva and a prominent decrease of microvilli and microplicae in the conjunctiva and cornea. The elements in the goblet cells were changed in these conditions. In addition, epithelial cells showed poor fibrous development and abnormal distribution of chromatin in the nucleus.Zn, Cu, Mn, and vitamins A and E deficiencies caused photoreceptor cells to degenerate and disappear. Se deficiency reduced the horizontal and amacrine cells. Vitamin B(12) deficiency reduced nerve fibers in the nerve fiber layer of the retina. Mg deficiency induced multifocal necrosis in the retinal pigment epithelium and apoptotic nuclear changes in the photoreceptor cells. Cr deficiency showed abnormal phagocytosis of the photoreceptor outer segment discs in the retinal pigment epithelium. Vitamin B(12) was found to be related to the circadian rhythm in the retina.Deficiencies of Zn, Cu, Mn, and vitamins A, B(12), and E induced degeneration and disappearance of myelin lamellae in the myelinated optic nerve fibers.In hypervitaminosis A, lipid droplets appeared in the retinal pigment epithelium and alcohol dehydrogenase disappeared in the retinal pigment epithelium and photoreceptor outer segments. Excessive Al was toxic to the retina, which showed disappearance of photoreceptor cells. Al deposits were seen in dendrites and neurons in the outer plexiform layer.Zn seemed to be necessary for corneal epithelial cell wound healing.Discussion: Trace elements usually are contained in enzymes, which have many metabolic functions. They are related to synthesis and breakdown of many substances. Some trace elements such as Zn, Cu, Mn, and Se and vitamins including vitamins A, C, and E prevent peroxidation of lipids. Some vitamins have an affinity for specific tissues such as epithelial cells, nerve fibers, and neuronal cells and are needed for cell differentiation, development, and maintenance.Conclusion: Cu, Zn, Mn, Se, Mg, and Cr and vitamins A, B(12), C, and E are necessary for maintenance of cellular structure and metabolism.

Fiber typing in aging muscle.

PubMed

Purves-Smith, Fennigje M; Sgarioto, Nicolas; Hepple, Russell T

2014-04-01

It is accepted widely that fast-twitch muscle fibers are preferentially impacted in aging muscle, yet we hypothesize that this is not valid when aging muscle atrophy becomes severe. In this review, we summarize the evidence of fiber type-specific effect in aging muscle and the potential confounding roles of fibers coexpressing multiple myosin heavy-chain isoforms and their histochemical identification.

HEPATOCYTE EXPRESION OF TUMOR ASSOCIATED ALDEHYDE DEHYDROGENASE (ALDH-3) AND P21 RAS FOLLOWING DIETHYLNITROSAMINE (DEN) INITIATION AND CHRONIC EXPOSURE TO DI(2-ETHYLHEXYL) PHTHALATE (DHEP)

EPA Science Inventory

Phthalate esters such as di(2-ethylhexyl)phthalate (DEHP)either promote or inhibit rat liver tumorigenesis depending on the carcinogenesis protocol. In this study, we examined the expression of two histochemical markers, the tumor associated isozyme of aldehyde dehydrogenase (ALD...

Transgene silencing of sucrose synthase in alfalfa (Medicago sativa L.) stem vascular tissue suggests a role for invertase in cell wall cellulose synthesis

USDA-ARS?s Scientific Manuscript database

Alfalfa (Medicago sativa L.) plants were transformed with two constructs: (1) a truncated phosphoenolpyruvate carboxylase promoter isolated from alfalfa nodules (PEPC-4) fused to GUS; and (2) PEPC-4 fused with sucrose synthase (SUS) isolated from alfalfa nodules. Histochemical staining for GUS in st...

A review on the effects of supercritical carbon dioxide on enzyme activity.

PubMed

Wimmer, Zdenek; Zarevúcka, Marie

2010-01-19

Different types of enzymes such as lipases, several phosphatases, dehydrogenases, oxidases, amylases and others are well suited for the reactions in SC-CO(2). The stability and the activity of enzymes exposed to carbon dioxide under high pressure depend on enzyme species, water content in the solution and on the pressure and temperature of the reaction system. The three-dimensional structure of enzymes may be significantly altered under extreme conditions, causing their denaturation and consequent loss of activity. If the conditions are less adverse, the protein structure may be largely retained. Minor structural changes may induce an alternative active protein state with altered enzyme activity, specificity and stability.

A Review on the Effects of Supercritical Carbon Dioxide on Enzyme Activity

PubMed Central

Wimmer, Zdeněk; Zarevúcka, Marie

2010-01-01

Different types of enzymes such as lipases, several phosphatases, dehydrogenases, oxidases, amylases and others are well suited for the reactions in SC-CO2. The stability and the activity of enzymes exposed to carbon dioxide under high pressure depend on enzyme species, water content in the solution and on the pressure and temperature of the reaction system. The three-dimensional structure of enzymes may be significantly altered under extreme conditions, causing their denaturation and consequent loss of activity. If the conditions are less adverse, the protein structure may be largely retained. Minor structural changes may induce an alternative active protein state with altered enzyme activity, specificity and stability. PMID:20162013

Biological abatement of enzyme inhibitors

USDA-ARS?s Scientific Manuscript database

Lignocellulose pretreatments release phenolic compounds that cause enzyme inhibition and deactivation. Bio-abatement, the biological removal of furfurals, acetic acid and phenolics, may utilize fungal fermentation to metabolize these compounds to CO2, water, cell mass, and heat. Our work with Coni...

Genetics Home Reference: pseudocholinesterase deficiency

MedlinePlus

... choline ester drugs. It is likely that the enzyme has other functions in the body, but these functions are not ... cause pseudocholinesterase deficiency result in an abnormal pseudocholinesterase enzyme that does not function properly. Other mutations prevent the production of the ...

Mitochondrial Neurogastrointestinal Encephalomyopathy Caused by Thymidine Phosphorylase Enzyme Deficiency: From Pathogenesis to Emerging Therapeutic Options

PubMed Central

Yadak, Rana; Sillevis Smitt, Peter; van Gisbergen, Marike W.; van Til, Niek P.; de Coo, Irenaeus F. M.

2017-01-01

Mitochondrial neurogastrointestinal encephalomyopathy (MNGIE) is a progressive metabolic disorder caused by thymidine phosphorylase (TP) enzyme deficiency. The lack of TP results in systemic accumulation of deoxyribonucleosides thymidine (dThd) and deoxyuridine (dUrd). In these patients, clinical features include mental regression, ophthalmoplegia, and fatal gastrointestinal complications. The accumulation of nucleosides also causes imbalances in mitochondrial DNA (mtDNA) deoxyribonucleoside triphosphates (dNTPs), which may play a direct or indirect role in the mtDNA depletion/deletion abnormalities, although the exact underlying mechanism remains unknown. The available therapeutic approaches include dialysis and enzyme replacement therapy, both can only transiently reverse the biochemical imbalance. Allogeneic hematopoietic stem cell transplantation is shown to be able to restore normal enzyme activity and improve clinical manifestations in MNGIE patients. However, transplant related complications and disease progression result in a high mortality rate. New therapeutic approaches, such as adeno-associated viral vector and hematopoietic stem cell gene therapy have been tested in Tymp-/-Upp1-/- mice, a murine model for MNGIE. This review provides background information on disease manifestations of MNGIE with a focus on current management and treatment options. It also outlines the pre-clinical approaches toward future treatment of the disease. PMID:28261062

A futile cycle, formed between two ATP-dependant gamma-glutamyl cycle enzymes, gamma-glutamyl cysteine synthetase and 5-oxoprolinase: the cause of cellular ATP depletion in nephrotic cystinosis?

PubMed

Kumar, Akhilesh; Bachhawat, Anand Kumar

2010-03-01

Cystinosis, an inherited disease caused by a defect in the lysosomal cystine transporter (CTNS), is characterized by renal proximal tubular dysfunction. Adenosine triphosphate (ATP) depletion appears to be a key event in the pathophysiology of the disease, even though the manner in which ATP depletion occurs is still a puzzle. We present a model that explains how a futile cycle that is generated between two ATP-utilizing enzymes of the gamma-glutamyl cycle leads to ATP depletion. The enzyme gamma-glutamyl cysteine synthetase (gamma-GCS), in the absence of cysteine, forms 5-oxoproline (instead of the normal substrate, gamma-glutamyl cysteine) and the 5-oxoproline is converted into glutamate by the ATP-dependant enzyme, 5-oxoprolinase. Thus, in cysteine-limiting conditions, glutamate is cycled back into glutamate via 5-oxoproline at the cost of two ATP molecules without production of glutathione and is the cause of the decreased levels of glutathione synthesis, as well as the ATP depletion observed in these cells. The model is also compatible with the differences seen in the human patients and the mouse model of cystinosis, where renal failure is not observed.

Effect of heavy metals ions on enzyme activity in the Mediterranean mussel, Donax trunculus

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mizrahi, L.; Achituv, Y.

Heavy metal ions strongly are bound by sulfhydryl groups of proteins. Sulfhydryl binding changes the structure and enzymatic activities of proteins and causes toxic effects evident at the whole organism level. Heavy metal ions like Cd, Cu, Hg, Zn, and Pb in sufficiently high concentrations might kill organisms or cause other adverse effects that changing aquatic community structures. Bivalves are known to be heavy metal accumulators. The aim of the present study was to examine the effects of different concentrations of each of five heavy metal ions on the activity of four enzymes in D. trunculus. As it is knownmore » that heavy metals inhibit the activity of a wide range of enzymes, the authors chose representative examples of dehydrogenases (lactate and malate dehydrogenases), respiratory enzyme (cytochrome oxidase) and digestive enzyme ({alpha}-amylase). The acute effects of different concentrations of selected metals were examined. These concentrations were higher than those found usually in the locality where the animals occur, but might be encountered during a given event of pollution.« less

DOE Office of Scientific and Technical Information (OSTI.GOV)

None

In order to stop bacteria from reproducing and causing a disease like tuberculosis, researchers must first block its enzymes' ability to bind with certain molecules. A research team from Brandeis University worked with the Advanced Protein Characterization Facility at Argonne National Laboratory to define 13 different bacterial structures and uncover the mechanism by which their enzymes form and break bonds with molecules. This animation depicts how an enzyme may be inhibited using this knowledge.

Life-history evolution and the microevolution of intermediary metabolism: activities of lipid-metabolizing enzymes in life-history morphs of a wing-dimorphic cricket.

PubMed

Zera, Anthony J; Zhao, Zhangwu

2003-03-01

Although a considerable amount of information is available on the ecology, genetics, and physiology of life-history traits, much more limited data are available on the biochemical and genetic correlates of life-history variation within species. Specific activities of five enzymes of lipid biosynthesis and two enzymes of amino acid catabolism were compared among lines selected for flight-capable (LW[f]) versus flightless (SW) morphs of the cricket Gryllus firmus. These morphs, which exist in natural populations, differ genetically in ovarian growth (100-400% higher in SW) and aspects of flight capability including the size of wings and flight muscles, and the concentration of triglyceride flight fuel (40% greater in LW[f]). Consistently higher activity of each enzyme in LW(f) versus SW-selected lines, and strong co-segregation between morph and enzyme activity, demonstrated genetically based co-variance between wing morph and enzyme activity. Developmental profiles of enzyme activities strongly paralleled profiles of triglyceride accumulation during adulthood and previous measures of in vivo lipid biosynthesis. These data strongly imply that genetically based elevation in activities of lipogenic enzymes, and enzymes controlling the conversion of amino acids into lipids, is an important cause underlying the elevated accumulation of triglyceride in the LW(f) morph, a key biochemical component of the trade-off between elevated early fecundity and flight capability. Global changes in lipid and amino-acid metabolism appear to have resulted from microevolutionary alteration of regulators of metabolism. Finally, strong genotype x environment (diet) interactions were observed for most enzyme activities. Future progress in understanding the functional causes of life-history evolution requires a more detailed synthesis of the fields of life-history evolution and metabolic biochemistry. Wing polymorphism is a powerful experimental model in such integrative studies.

Toward reducing immunogenicity of enzyme replacement therapy: altering the specificity of human β-glucuronidase to compensate for α-iduronidase deficiency.

PubMed

Chuang, Huai-Yao; Suen, Ching-Shu; Hwang, Ming-Jing; Roffler, Steve R

2015-11-01

Enzyme replacement therapy (ERT) is an effective treatment for many patients with lysosomal storage disorders caused by deficiency in enzymes involved in cell metabolism. However, immune responses that develop against the administered enzyme in some patients can hinder therapeutic efficacy and cause serious side effects. Here we investigated the feasibility of a general approach to decrease ERT immunogenicity by altering the specificity of a normal endogenous enzyme to compensate for a defective enzyme. We sought to identify human β-glucuronidase variants that display α-iduronidase activity, which is defective in mucopolysaccharidosis (MPS) type I patients. A human β-glucuronidase library was screened by the Enzyme Cleavable Surface-Tethered All-purpose Screen sYstem to isolate variants that exhibited 100-290-fold greater activity against the α-iduronidase substrate 4-methylumbelliferyl α-l-iduronide and 7900-24 500-fold enzymatic specificity shift when compared with wild-type β-glucuronidase. In vitro treatment of MPS I cells with the β-glucuronidase variants significantly restored lysosome appearance similar to treatment with α-iduronidase. Our study suggests that β-glucuronidase variants can be isolated to display α-iduronidase activity and produce significant phenotype improvement of MPS I cells. This strategy may represent a possible approach to produce enzymes that display therapeutic benefits with potentially less immunogenicity. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Activity loss by H46A mutation in Mycobacterium tuberculosis isocitrate lyase is due to decrease in structural plasticity and collective motions of the active site.

PubMed

Shukla, Rohit; Shukla, Harish; Tripathi, Timir

2018-01-01

Mycobacterium tuberculosis isocitrate lyase (MtbICL) is a crucial enzyme of the glyoxylate cycle and is a validated anti-tuberculosis drug target. Structurally distant, non-active site mutation (H46A) in MtbICL has been found to cause loss of enzyme activity. The aim of the present work was to explore the structural alterations induced by H46A mutation that caused the loss of enzyme activity. The structural and dynamic consequences of H46A mutation were studied using multiple computational methods such as docking, molecular dynamics simulation and residue interaction network analysis (RIN). Principal component analysis and cross correlation analysis revealed the difference in conformational flexibility and collective modes of motions between the wild-type and mutant enzyme, particularly in the active site region. RIN analysis revealed that the active site geometry was disturbed in the mutant enzyme. Thus, the dynamic perturbation of the active site led to enzyme transition from its active form to inactive form upon mutation. The computational analyses elucidated the mutant-specific conformational alterations, differential dominant motions, and anomalous residue level interactions that contributed to the abrogated function of mutant MtbICL. An understanding of interactions of mutant enzymes may help in modifying the existing drugs and designing improved drugs for successful control of tuberculosis. Copyright © 2017 Elsevier Ltd. All rights reserved.

The effect of β-N-methylamino-L-alanine (BMAA) on oxidative stress response enzymes of the macrophyte Ceratophyllum demersum.

PubMed

Esterhuizen-Londt, M; Pflugmacher, S; Downing, T G

2011-04-01

Cyanobacteria are known to produce bioactive secondary metabolites such as hepatotoxins, cytotoxins and neurotoxins. The newly recognized neurotoxin β-N-methylamino-L-alanine (BMAA) is a naturally occurring non-protein amino acid found in the majority of cyanobacterial genera tested. Evidence that exists for implication of BMAA in neurodegenerative disorders relies on bioaccumulation and biomagnification from symbiotic cyanobacteria. Uptake and accumulation of free BMAA by various non-symbiotic organisms, including aquatic macrophytes, has been documented but to date limited evidence of ecotoxicology exists. We therefore investigated the effect of BMAA on the oxidative stress responses of the macrophyte, Ceratophyllum demersum. Markers for oxidative stress in this study are the antioxidative enzymes superoxide dismutase, catalase, guaiacol peroxidase, glutathione peroxidase and glutathione reductase. We found that BMAA had an inhibitory effect on all the oxidative stress response enzymes tested in plants exposed to BMAA. However enzymes not related to oxidative stress response were not affected by BMAA in in vitro experiments. Binding studies in the presence of BMAA showed reduced enzyme specific activity over time compared to the control. This study shows that BMAA causes oxidative stress indirectly as it inhibits antioxidant enzymes required to combat reactive oxygen species that cause damage to cells. Further investigations are required to fully understand the inhibitory effect of BMAA on these enzymes. Copyright © 2011 Elsevier Ltd. All rights reserved.

Genetics Home Reference: Bietti crystalline dystrophy

MedlinePlus

... broken down and converted into energy, but the enzyme's specific function is not well understood. CYP4V2 gene mutations that cause Bietti crystalline dystrophy impair or eliminate the function of this enzyme and are believed to affect lipid breakdown. However, ...

Genetics Home Reference: TK2-related mitochondrial DNA depletion syndrome, myopathic form

MedlinePlus

... mtDNA. Specifically, this enzyme plays a role in recycling mtDNA building blocks (nucleotides) so that errors in ... kinase 2. A decrease in enzyme activity impairs recycling of mtDNA nucleotides, causing a shortage of nucleotides ...

Release of enzymes from lysosomes by irradiation and the relation of lipid peroxide formation to enzyme release

PubMed Central

Wills, E. D.; Wilkinson, A. E.

1966-01-01

1. Acid phosphatase, cathepsin and β-glucuronidase are released from rat-liver lysosomes by irradiation in vitro. Enzyme release is detectable after a dose of 1krad and increases with dose up to 100krads. 2. Maximum radiation effects were observed when the lysosomes were kept for 20hr. at 4° or 20° after irradiation. 3. An atmosphere of nitrogen considerably decreases enzyme release from lysosomes. 4. Enzyme release is enhanced by ascorbic acid and decreased by vitamin E. 5. Irradiation causes formation of lipid peroxides in lysosomes, and enzyme release increases with lipid peroxide formation. 6. It is suggested that lipid peroxide formation leads to rupture of the lysosome membrane and allows release of the contained hydrolytic enzymes. PMID:5964962

Lipid Storage Myopathy in Behçet's Disease: A Rare Cause of Elevated Serum Creatine Kinases Levels

PubMed Central

Yilmaz, Sedat; Cinar, Muhammet; Karslioglu, Yıldırım; Simsek, Ismail; Erdem, Hakan; Pay, Salih; Dinc, Ayhan

2012-01-01

Muscular involvement in Behçet's disease is rare and there are only a few case reports in the literature. The causes of elevated muscle enzymes in a patient with Behcet's disease are many, including myositis, drug-induced myopathy, metabolic myopathy, and the disease itself. We herein have defined an algorithmic approach to a patient with Behcet's disease and elevated muscle enzymes and report a case of coexisting of lipid storage myopathy. PMID:22937450

Evaluation of 2-Thioxo-2,3,5,6,7,8-hexahydropyrimido[4,5-d]pyrimidin-4(1H)-one analogues as GAA Activators

PubMed Central

Marugan, Juan J.; Zheng, Wei; Motabar, Omid; Southall, Noel; Goldin, Ehud; Sidransky, Ellen; Aungst, Ronald A.; Liu, Ke; Sadhukhan, Subir Kumar; Austin, Christopher P.

2010-01-01

Pompe disease is a lysosomal storage disease (LSD) caused by a deficiency in the lysosomal enzyme acid α-glucosidase. In several LSDs, enzyme inhibitors have been used as small molecule chaperones to facilitate and increase the translocation of mutant protein from the endoplasmic reticulum to the lysosome. Enzyme activators with chaperone activity would be even more desirable as they would not inhibit the enzyme after translocation and might potentiate the activity of the enzyme that is successfully translocated. Herein we report our initial findings of a new series of acid α-glucosidase activators. PMID:20206419

Evaluation of 2-thioxo-2,3,5,6,7,8-hexahydropyrimido[4,5-d]pyrimidin-4(1H)-one analogues as GAA activators.

PubMed

Marugan, Juan J; Zheng, Wei; Motabar, Omid; Southall, Noel; Goldin, Ehud; Sidransky, Ellen; Aungst, Ronald A; Liu, Ke; Sadhukhan, Subir Kumar; Austin, Christopher P

2010-05-01

Pompe disease is a lysosomal storage disease (LSD) caused by a deficiency in the lysosomal enzyme acid alpha-glucosidase. In several LSDs, enzyme inhibitors have been used as small molecule chaperones to facilitate and increase the translocation of mutant protein from the endoplasmic reticulum to the lysosome. Enzyme activators with chaperone activity would be even more desirable as they would not inhibit the enzyme after translocation and might potentiate the activity of the enzyme that is successfully translocated. Herein we report our initial findings of a new series of acid alpha-glucosidase activators.

Enzymatic Detachment of Staphylococcus epidermidis Biofilms

PubMed Central

Kaplan, Jeffrey B.; Ragunath, Chandran; Velliyagounder, Kabilan; Fine, Daniel H.; Ramasubbu, Narayanan

2004-01-01

The gram-positive bacterium Staphylococcus epidermidis is the most common cause of infections associated with catheters and other indwelling medical devices. S. epidermidis produces an extracellular slime that enables it to form adherent biofilms on plastic surfaces. We found that a biofilm-releasing enzyme produced by the gram-negative periodontal pathogen Actinobacillus actinomycetemcomitans rapidly and efficiently removed S. epidermidis biofilms from plastic surfaces. The enzyme worked by releasing extracellular slime from S. epidermidis cells. Precoating surfaces with the enzyme prevented S. epidermidis biofilm formation. Our findings demonstrate that biofilm-releasing enzymes can exhibit broad-spectrum activity and that these enzymes may be useful as antibiofilm agents. PMID:15215120

Differential effects of non-ionic detergents on microsomal and sarcolemmal adenylate cyclase in cardiac muscle

PubMed Central

Sulakhe, Prakash V.; Narayanan, Njanoor

1978-01-01

1. About 4 and 23% of the homogenate adenylate cyclase activity was recovered in the microsomal and sarcolemmal fractions isolated from guinea-pig heart ventricles. 2. Cardiac microsomal adenylate cyclase activity [basal as well as p[NH]ppG (guanyl-5′-yl imidodiphosphate)- and NaF-stimulated] was increased over 2-fold in the presence of Lubrol-PX (0.01–0.1%). 3. The sarcolemmal enzyme, however, showed concentration-dependent inhibition caused by the detergent under all assay conditions, except when p[NH]ppG was included in the assay. In the latter case, the detergent (0.01–0.02%) caused a modest increase (30–45%) in enzyme activity. 4. Another non-ionic detergent, Triton X-100, also stimulated the microsomal cyclase and inhibited the sarcolemmal enzyme. 5. With either membrane fraction, Lubrol-PX solubilized the enzyme when the detergent/membrane protein ratio was 2.5 (μmol of detergent/mg of protein). 6. The findings with homogenate and a washed particulate fraction resembled those obtained with sarcolemma, and those with isolated sarcoplasmic reticulum resembled those with microsomal preparations. 7. p[NH]ppG, and to some extent NaF, protected the detergent-induced inactivation of the enzyme observed at higher detergent concentrations (0.5% Lubrol-PX and 0.05–0.5% Triton X-100). 8. In the absence of detergents, p[NH]ppG increased the basal enzyme activity about 2-fold in microsomal fractions, but did not appreciably stimulate the sarcolemmal enzyme. Isoproterenol, on the other hand, increased the sarcolemmal enzyme activity (>2-fold) in the presence of p[NH]ppG and caused only moderate stimulation (31%) of the microsomal enzyme under these conditions. 9. These findings support the view that, although the bulk of adenylate cyclase resides in heart sarcolemma (plasma membrane), the microsomal activity cannot be accounted for solely by contamination of the microsomal fraction with sarcolemma, as has been suggested by others [Besch, Jones & Watanabe (1976) Circ. Res. 39, 586–595; Engelhard, Plut & Storm (1976) Biochim. Biophys. Acta 451, 48–61]. Further, the results of this study show that cardiac sarcoplasmic-reticulum membranes possess this enzyme. PMID:736892

Resolution of Hydronephrosis in a Patient With Mucopolysaccharidosis Type II With Enzyme Replacement Therapy.

PubMed

Nishiyama, Kei; Imai, Takashi; Ohkubo, Kazuhiro; Sanefuji, Masafumi; Takada, Hidetoshi

2017-03-01

Mucopolysaccharidosis type II (MPS II) is caused by deficiency of lysosomal enzyme iduronate-2-sulfatase. Insufficient activity of the enzyme results in accumulation of glycosaminoglycans leading to progressive multisystem pathologies. MPS II is less likely to be complicated by kidney and urinary tract problems. We report a boy with MPS II, who developed left hydronephrosis. His hydronephrosis improved after starting enzyme replacement therapy. It was suggested that MPS II was closely associated with the pathogenesis of hydronephrosis. Copyright © 2016 Elsevier Inc. All rights reserved.

Human β-glucuronidase: structure, function, and application in enzyme replacement therapy.

PubMed

Naz, Huma; Islam, Asimul; Waheed, Abdul; Sly, William S; Ahmad, Faizan; Hassan, Imtaiyaz

2013-10-01

Lysosomal storage diseases occur due to incomplete metabolic degradation of macromolecules by various hydrolytic enzymes in the lysosome. Despite structural differences, most of the lysosomal enzymes share many common features including a lysosomal targeting motif and phosphotransferase recognition sites. β-Glucuronidase (GUSB) is an important lysosomal enzyme involved in the degradation of glucuronate-containing glycosaminoglycan. The deficiency of GUSB causes mucopolysaccharidosis type VII (MPSVII), leading to lysosomal storage in the brain. GUSB is a well-studied protein for its expression, sequence, structure, and function. The purpose of this review is to summarize our current understanding of sequence, structure, function, and evolution of GUSB and its lysosomal enzyme targeting. Enzyme replacement therapy reported for this protein is also discussed.

Morphological and histochemical demonstration of hooks in the rostellar region of Cysticercus bovis.

PubMed

Zdárská, Z

1976-01-01

Hooklets and early developmental stages of hooks were discovered in the tegument among the microtriches in the rostellar region of 83- and 108-day-old Cysticercus bovis. The cortical layer of hooks consists of proteins with tyrosine and SS groups. The rostellar sac and bulb are distinctly developed in the parenchyma of the rostellar region.

Bronislawa and Mieczyslaw Konopacki - pioneers of the application of histochemistry to embryology in Poland.

PubMed

Bielanska-Osuchowska, Zofia

2008-01-01

In this paper, we review the life and work of Mieczyslaw Konopacki and his wife Bronislawa Konopacka. These Polish scientists introduced histochemical methods to embryological studies at the beginning of the XXth century; they were pioneers in this regard in Poland and among the first in the world to implement such methods in this way.

News and views in Histochemistry and Cell Biology.

PubMed

Asan, Esther; Drenckhahn, Detlev

2004-12-01

Advances in histochemical methodology and ingenious applications of novel and improved methods continue to confirm the standing of morphological means and approaches in research efforts, and contribute significantly to increasing our knowledge about structures and functions in all areas of the life sciences from cell biology to pathology. Reports published during recent months documenting this progress are summarized in the present review.

A quantitative study of the histochemical and morphometric characteristics of the human cricopharyngeus muscle.

PubMed Central

Brownlow, H; Whitmore, I; Willan, P L

1989-01-01

Samples of human cricopharyngeus muscles obtained at postmortem were assessed for fibre type composition and fibre size. Fibre type was determined using serial cryostat sections exposed to several histochemical reactions; narrow fibre diameter and fibre area were measured from photomicrographs using a digitiser interfaced to a microcomputer. Results were compared with values from samples of vastus lateralis obtained from the same subjects. Cricopharyngeus muscle, in comparison with vastus lateralis, contained significantly more oxidative fibres but fewer glycolytic fibres and significantly more Type I fibres but fewer Type IIB. Cricopharyngeal fibres were significantly smaller than the fibres in vastus lateralis and in neither muscle were fibre sizes normally distributed. In each muscle most Type I fibres were oxidative, and the ratio of oxidative: glycolytic fibres was similar for Type IIA and IIB fibres. The fibre type proportions and fibre sizes in cricopharyngeus did not vary significantly with age or between males and females. The composition of cricopharyngeus--mostly Type I oxidative fibres and few Type II glycolytic fibres--correlated well with the functions of sustained tonicity to prevent aerophagia and occasional forceful contraction during deglutition. Images Fig. 1 PMID:2621147

MORPHOLOGICAL AND HISTOCHEMICAL CHANGES IN THE SPLEEN AND LYMPH NODE OF WHITE MICE IN RESPONSE TO RADIATION AND ANTIGENIC EFFECTS (in Russian)

DOE Office of Scientific and Technical Information (OSTI.GOV)

Akopova, V.A.

1961-01-01

Research was carried out on 90 white ntice subjected to x irradiation (Series I), vaccination with a brucellosis vaccine (Series II), and to a combined action of these factors (Series III and IV). The spleen and lymph nodes were studied both morphologically and histochemically. When exposing the mice to a total x-ray dose of 335 r, visible destructive changes and the lowering of desoxyribonucleic and ribonucleic acid level were found to occur in the spleen and lymph nodes. In the case of a single vaccination of the mice with the brucellosis vaccine, a certain vaccinal hyperplasia was observed in themore » spleen and lymph nodes, the level of desoxyribonucleic and ribonucleic acid being normal. In vaccinating the white mice, following total irradiation, destructive changes along with a gradual lowering of desoxyribonucleic and ribonucleic acid level are observed in the spleen and lymph nodes. The total x irradiation following a singie vaccination with the brucellosis vaccine, brings about slight destructive changes along with a simultaneous reticular hyperplasia in the spleen and lymph nodes. The levels of desoxyribonucleic and ribonucleic acids are high at all times. (auth)« less

Microspore development of three coniferous species: affinity of nuclei for flavonoids.

PubMed

Feucht, Walter; Treutter, Dieter; Dithmar, Heike; Polster, Jürgen

2008-12-01

The nuclear localization of blue-staining flavanols was investigated histochemically throughout microsporogenesis in yellow cypress (Callitropsis nootkatensis (D. Don) Oerst., formerly Cupressus nootkatensis), juniper (Juniperus communis L.) and yew (Taxus baccata L.). During meiotic development, both the cytoplasm and nuclei of microspores of all species contained varying amounts of flavanols; however, the flavanols were largely confined to the nuclei in microspores just released from tetrads. Quantification by HPLC analysis indicated that, in all species, catechin and epicatechin were the dominant nuclear flavanols. At the early free microspore stage, the nuclear flavanols were barely detectable in all species, but they increased fivefold on incubation in the presence of 0.1 mM benzylaminopurine (BA) or zeatin. Histochemical studies revealed that, in addition to non-fluorescing flavanols, microspores contained yellow-fluorescing flavonoids, which yielded a distinct HPLC flavonoid profile for each species. In yellow cypress, the hydrolyzed flavonoids were identified as quercetin, apigenin, kaempferol and luteolin, whereas only quercetin and myricetin were found in microspores of juniper and in anthers of yew. Application of a UV-VIS titration technique revealed that the aglycone quercetin seems to interact more strongly with histone H3 than either glycoside rutin or kaempferol.

A Historical Perspective on the Identification of Cell Types in Pancreatic Islets of Langerhans by Staining and Histochemical Techniques

PubMed Central

2015-01-01

Before the middle of the previous century, cell types of the pancreatic islets of Langerhans were identified primarily on the basis of their color reactions with histological dyes. At that time, the chemical basis for the staining properties of islet cells in relation to the identity, chemistry and structure of their hormones was not fully understood. Nevertheless, the definitive islet cell types that secrete glucagon, insulin, and somatostatin (A, B, and D cells, respectively) could reliably be differentiated from each other with staining protocols that involved variations of one or more tinctorial techniques, such as the Mallory-Heidenhain azan trichrome, chromium hematoxylin and phloxine, aldehyde fuchsin, and silver impregnation methods, which were popularly used until supplanted by immunohistochemical techniques. Before antibody-based staining methods, the most bona fide histochemical techniques for the identification of islet B cells were based on the detection of sulfhydryl and disulfide groups of insulin. The application of the classical islet tinctorial staining methods for pathophysiological studies and physiological experiments was fundamental to our understanding of islet architecture and the physiological roles of A and B cells in glucose regulation and diabetes. PMID:26216133

Pollen morphology and viability in Bromeliaceae.

PubMed

Souza, Everton H; Souza, Fernanda V D; Rossi, Mônica L; Packer, Renan M; Cruz-Barros, Maria Amelia V; Martinelli, Adriana P

2017-01-01

Pollen morphology characterization is important in taxonomy, conservation and plant breeding, and pollen viability studies can support breeding programs. This study investigated pollen morphology and male fertility in 18 species of Bromeliaceae with ornamental potential. For morphological characterization, pollen grains were acetolyzed and characterization of exine was done using scanning and transmission electron microscopy. Pollen viability was investigated by in vitro germination and histochemical tests. Species belonging to Aechmea and Ananas genera presented medium size pollen, except for Ae. fasciata, with large pollen. Al. nahoumii, P. sagenarius and the Vriesea species analyzed showed large pollen, except for V. carinata, with very large pollen. Pollen of Aechmea, Ananas and P. sagenarius presented bilateral symmetry, diporate, exine varying from tectate to semitectate. Al. nahoumii and Vriesea species presented pollen with bilateral symmetry, monocolpate; exine was semitectate, reticulate and heterobrochate. Germination percentage and tube growth were greater in SM and BKM media. Histochemical tests showed pollen viability above 70% for all species, except for Ananas sp. (40%). Pollen morphology is important for the identification of species, especially in this family, which contains a large number of species. High rates of viability favor fertilization and seed production, essential for efficient hybrid production and conservation.

Influence of unilateral tooth loss in the temporomandibular joint and masseter muscle of rabbits.

PubMed

Im, Jae-Hyung; Kim, Su-Gwan; Oh, Ji-Su; Lim, Sung-Chul; Ha, Jung-Min

2012-07-01

The purpose of this study was to evaluate the influence of the masticatory system in patients with missing teeth. The influence of tooth loss on the masticatory system was analyzed with the use of bone scintigraphy ((99m)Tc-MDP) and histochemistry. Eight white rabbits (New Zealand, 12 weeks old) were used. The rabbits were divided into 2 groups: 6 weeks and 12 weeks. Teeth were extracted unilaterally in each rabbit under general anesthesia. Six and 12 weeks after extraction, scintigraphy was conducted, and the rabbits were killed and their masseter muscles removed for histochemical analysis. The results of bone metabolism (relative ratio) measured by bone scintigraphy were 48.27% at extraction sites and 51.73% at nonextraction sites at 6 weeks and 39.96% at extraction sites and 60.04% at nonextraction sites at 12 weeks. There was a significant difference at 12 weeks (P < .05). Tissue calcium contents and osteoclast counts showed different results between the extraction and nonextraction sites, but these differences did not reach statistical significance. The bone metabolism of temporomandibular joints and histochemical aspects of masticatory muscles may be associated with occlusal alterations following tooth loss. Copyright © 2012 Elsevier Inc. All rights reserved.

Lectin histochemistry of the interdigital gland in the Japanese serow (Capricornis crispus) in winter.

PubMed Central

Atoji, Y; Suzuki, Y; Sugimura, M

1988-01-01

The interdigital gland of the Japanese serow was examined by histological and lectin histochemical techniques. The gland is composed of a thin-walled pouch and a duct. Both regions contain sebaceous and apocrine glands, but the development of each component was significantly less marked than those of the skin in the region. In particular, only a small amount of sebaceous and apocrine glandular elements was found in the pouch, although they were more abundant in the duct. Histochemical staining of the sebaceous and apocrine glands showed similar reactions to six lectins except for UEA in the interdigital gland and digital surface skin. UEA reacted with the apocrine part of the interdigital gland, but not with the gland in the digital surface skin. In addition, tubules in the apocrine gland revealed eight different staining patterns with UEA. These stainings possibly represent a cyclic activity of glandular tubules and suggest that the apocrine portion of the interdigital gland has a different function from that of the body skin. Images Fig. 1 Fig. 3 Fig. 4 Fig. 5 Fig. 6 Fig. 7 Fig. 8 Fig. 9 Fig. 10 Fig. 11 Fig. 12 PMID:3254889

Neonatal myosin in bovine and pig tensor tympani muscle fibres.

PubMed Central

Scapolo, P A; Rowlerson, A; Mascarello, F; Veggetti, A

1991-01-01

In previous studies of middle ear muscles, the classification of fibre types by histochemical methods was particularly difficult in the bovine and porcine tensor tympani muscle, suggesting the presence of immature fibres. We therefore reexamined the tensor tympani from pigs and cattle of various ages immunohistochemically, using a panel of antimyosin antibodies, including one (anti-NE) specific for neonatal and embryonic myosins. Fibres positive to anti-NE were found in tensor tympani in both species in all ages examined; only a few of these fibres reacted exclusively with this antibody; some also contained slow myosin and the majority also contained adult fast (type IIA) myosin. Furthermore, although the remaining fibres included some of the classical types I and IIA, the majority of them showed a mismatch between their histochemical and immunohistochemical profiles. The morphological appearance of the muscle, the widespread presence of neonatal myosin (often together with another myosin in the same fibre) and the persistence of this composition from birth to adulthood, could be explained by an incomplete development of the muscle fibres, resulting in a 'muscle' much better suited to the role of a ligament. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 PMID:1810932

Mapping and reconstruction of domoic acid-induced neurodegeneration in the mouse brain.

PubMed

Colman, J R; Nowocin, K J; Switzer, R C; Trusk, T C; Ramsdell, J S

2005-01-01

Domoic acid, a potent neurotoxin and glutamate analog produced by certain species of the marine diatom Pseudonitzschia, is responsible for several human and wildlife intoxication events. The toxin characteristically damages the hippocampus in exposed humans, rodents, and marine mammals. Histochemical studies have identified this, and other regions of neurodegeneration, though none have sought to map all brain regions affected by domoic acid. In this study, mice exposed (i.p.) to 4 mg/kg domoic acid for 72 h exhibited behavioral and pathological signs of neurotoxicity. Brains were fixed by intracardial perfusion and processed for histochemical analysis. Serial coronal sections (50 microm) were stained using the degeneration-sensitive cupric silver staining method of DeOlmos. Degenerated axons, terminals, and cell bodies, which stained black, were identified and the areas of degeneration were mapped onto Paxinos mouse atlas brain plates using Adobe Illustrator CS. The plates were then combined to reconstruct a 3-dimensional image of domoic acid-induced neurodegeneration using Amira 3.1 software. Affected regions included the olfactory bulb, septal area, and limbic system. These findings are consistent with behavioral and pathological studies demonstrating the effects of domoic acid on cognitive function and neurodegeneration in rodents.

Ovary organization and oogenesis in the tardigrade Macrobiotus polonicus Pilato, Kaczmarek, Michalczyk & Lisi, 2003 (Eutardigrada, Macrobiotidae): ultrastructural and histochemical analysis.

PubMed

Poprawa, Izabela; Schlechte-Wełnicz, Weronika; Hyra, Marta

2015-05-01

The female reproductive system, the process of oogenesis, and the morphology of the egg capsule of Macrobiotus polonicus were analyzed using transmission and scanning electron microscopy and histochemical methods. The female reproductive system of Macrobiotus polonicus consists of a single ovary and a single oviduct that opens into the cloaca. The seminal receptacle filled with sperm cells is present. The ovary is divided into two parts: a germarium that is filled with oogonia and a vitellarium that is filled with branched clusters of the germ cells. Meroistic oogenesis occurs in the species that was examined. The yolk material is synthesized by the oocyte (autosynthesis) and by the trophocytes and is transported to the oocyte through cytoplasmic bridges. The process of the formation of the egg envelopes starts in the late vitellogenesis. The egg capsule is composed of two envelopes-the vitelline envelope and the three-layered chorion. The vitelline envelope is of the primary type while the chorion is of a secondary type. The surface of the chorion is covered with conical processes that terminate with a strongly indented terminal disc.

The functional consequences of mis-sense mutations affecting an intra-molecular salt bridge in arylsulphatase A.

PubMed Central

Schestag, Frank; Yaghootfam, Afshin; Habetha, Matthias; Poeppel, Peter; Dietz, Frank; Klein, Roger A; Zlotogora, Joel; Gieselmann, Volkmar

2002-01-01

Metachromatic leukodystrophy is a lysosomal storage disorder caused by the deficiency of arylsulphatase A. We describe the functional consequences of three mis-sense mutations in the arylsulphatase A gene (Asp-335-Val, Arg-370-Trp and Arg-370-Gln), affecting an apparent intramolecular Asp-335 to Arg-370 salt bridge, and interpret the effects and clinical consequences on the basis of the three-dimensional structure of arylsulphatase A. Asp-335-Val and Arg-370-Trp substitutions each cause a complete loss of enzyme activity and are associated with the most severe form of the human disease, whereas the Arg-370-Gln-substituted enzyme retains some residual activity, being found in a patient suffering from the milder juvenile form of the disease. Detailed analysis reveals that formation of the apparent salt bridge depends critically on the presence of aspartic acid and arginine residues at positions 335 and 370, respectively. Substitution by various other amino acids, including glutamic acid and lysine, affects enzyme function severely. Biosynthesis and immunoprecipitation studies indicate that the Asp-335-Val substitution affects folding of arylsulphatase A more severely than either the Arg-370-Trp or Arg-370-Gln substitutions. In vitro mutagenesis data show that clinical severity correlates with the space occupied by residue 370. The combination with structural data suggests that the bulky tryptophan residue broadens the cleft held together by the apparent salt bridge, whereas the smaller glutamine residue still allows the cleft to close, yielding a less severely affected enzyme. The position of residue 370 in the three-dimensional structure of the enzyme provides a plausible explanation for the differing severities in loss of enzyme function caused by the mutations and thus the clinical phenotype. PMID:12086582

Dipeptidyl peptidase-4 inhibitor induced angioedema - an overlooked and potentially lethal adverse drug reaction?

PubMed

Scott, Susanne Irene; Andersen, Michelle Fog; Aagaard, Lise; Buchwald, Christian Von; Rasmussen, Eva Rye

2017-02-14

Introduction Angioedema is a potentially fatal adverse drug reaction of some medications, as swellings of the upper airways can cause death by asphyxiation. Angiotensin converting enzyme-inhibitors are widely known to cause angioedema but less is known about the association between dipeptidyl peptidase-4 inhibitors (gliptins) and angioedema. Dipeptidyl peptidase-4 inhibitors are anti-diabetic drugs used to improve glycaemic control. They, as a class effect, inadvertently affect the degradation of the vasoactive kinins bradykinin and substance P, both of which can cause angioedema due to vasodilatation and increase in vascular permeability in the capillaries. Objective To assess the risk and pathomechanism of angioedema due to inhibition of dipeptidyl peptidase-4 inhibitors when used as monotherapy and in combination with angiotensin converting enzyme-inhibitors. Method PubMed, Embase, the Cochrane Library, PubMed Central, Web of Science, Google Scholar and clinicaltrials.gov were searched using different combinations of keywords "angioedema", "dipeptidyl peptidase 4", "dipeptidyl peptidase 4 inhibitors", "gliptins", "bradykinin", "substance P" and "angiotensin converting enzyme-inhibitors". Original research papers were preferably used as references and their bibliographies were used to further the search for original research results. Results Both angiotensin converting enzyme and dipeptidyl peptidase-4 are major enzymes in the degradation pathway of bradykinin and substance P, and when inhibited pharmacologically - especially at the same time - the theoretical risk of angioedema is increased due to accumulation of vasoactive kinins. Conclusion Treatment with dipeptidyl peptidase-4 inhibitors must be carefully considered and monitored especially during concurrent treatment with angiotensin converting enzyme-inhibitors or when treating patients with a known predisposition to angioedema. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

The influence of gemfibrozil on malondialdehyde level and paraoxonase 1 activity in wistar and fisher rats.

PubMed

Macan, Marija; Marija, Macan; Konjevoda, Paško; Paško, Konjevoda; Lovric, Jasna; Jasna, Lovrić; Koprivanac, Marijan; Marijan, Koprivanac; Kelava, Marta; Marta, Kelava; Vrkic, Nada; Nada, Vrkić; Bradamante, Vlasta; Vlasta, Bradamante

2011-06-01

There are diverse experimental data about the influence of gemfibrozil (GEM) on the production of hydrogen peroxide (H(2)O(2)) and antioxidant enzymes. We investigated the influence of GEM treatment on the production of malondialdehyde (MDA) level in tissues of normolipidaemic Wistar and Fisher rats which is an index of lipid peroxidation. Because serum paraoxonase 1 (PON1) is an important enzyme with specific protective function on metabolism of lipid peroxides, we examined the influence of GEM on PON1 activity in liver and serum. MDA level and enzyme activities were also determined 10 days after withdrawal of GEM treatment. The significantly increased levels of MDA in liver, kidney and heart of both rat strains were obtained after 3 weeks of GEM treatment. We propose two possibilities for the increase of MDA levels caused by GEM, induction of peroxisome proliferation and activities of enzymes that participated in occurrence of H(2)O(2) and possible reduction of enzyme activities including in H(2)O(2) metabolism. Ten days after withdrawal of GEM treatment, MDA levels in all tissue levels of both rat strains were less in comparison with GEM treatment. GEM caused a significant drop of PON1 activity in serum and liver of Fisher rats, and in liver of Wistar rats. We suggest that GEM, through induction of lipid peroxidation, caused the damage of hepatocytes with consequent reduction of PON1 synthesis. The increase in PON1 activity in serum and tissues of both rat strains 10 days after withdrawal of GEM treatment shows the fast recovery of enzyme synthesis. © 2011 The Authors. Basic & Clinical Pharmacology & Toxicology © 2011 Nordic Pharmacological Society.

Salmon caviar-induced anaphylactic shock.

PubMed

Flais, Michael J; Kim, Susan S; Harris, Kathleen E; Greenberger, Paul A

2004-01-01

Foods, particularly shellfish and nuts, are commonly implicated as causes of anaphylaxis. Salmon caviar, to our knowledge, is an exceedingly rare cause of anaphylactic shock. This study describes a patient who experienced anaphylactic shock on her initial ingestion of caviar. Skin testing and inhibition assays using the enzyme-linked immunosorbent assay were performed. On the percutaneous test, the patient had a 7 x 9 mm/15 x 55 mm wheal/ erythema reaction to caviar liquid. Caviar liquid caused 68, 73, and 73% inhibition of immunoglobulin E (IgE) binding at concentrations of 0.01, 0.1, and 1.0 mg/mL in an enzyme-linked immunosorbent assay, respectively. There was no evidence for IgE antibodies that could be demonstrated to bind to the caviar eggs. Despite using both metoprolol and lisinopril, the patient responded promptly to subcutaneous epinephrine. This report indicates that an IgE-mediated response occurred after caviar ingestion. Although she experienced anaphylactic shock, the patient recovered quickly after epinephrine administration despite routine use of a 3-adrenergic blocker and angiotensin-converting enzyme inhibitor.

The D153del Mutation in GNB3 Gene Causes Tissue Specific Signalling Patterns and an Abnormal Renal Morphology in Rge Chickens

PubMed Central

Tummala, Hemanth; Fleming, Stewart; Hocking, Paul M.; Wehner, Daniel; Naseem, Zahid; Ali, Manir; Inglehearn, Christopher F.; Zhelev, Nikolai; Lester, Douglas H.

2011-01-01

Background The GNB3 gene is expressed in cone but not rod photoreceptors of vertebrates, where it acts as the β transducin subunit in the colour visual transduction process. A naturally occurring mutation ‘D153del’ in the GNB3 gene causes the recessively inherited blinding phenotype retinopathy globe enlarged (rge) disease in chickens. GNB3 is however also expressed in most other vertebrate tissues suggesting that the D153del mutation may exert pathological effects that outlie from eye. Principal Findings Recombinant studies in COS-7 cells that were transfected with normal and mutant recombinant GNB3 constructs and subjected to cycloheximide chase showed that the mutant GNB3d protein had a much shorter half life compared to normal GNB3. GNB3 codes for the Gβ3 protein subunit that, together with different Gγ and Gα subunits, activates and regulates phosphorylation cascades in different tissues. As expected, the relative levels of cGMP and cAMP secondary messengers and their activated kinases such as MAPK, AKT and GRK2 were also found to be altered significantly in a tissue specific manner in rge chickens. Histochemical analysis on kidney tissue sections, from rge homozygous affected chickens, showed the chickens had enlargement of the glomerular capsule, causing glomerulomegaly and tubulointerstitial inflammation whereas other tissues (brain, heart, liver, pancreas) were unaffected. Significance These findings confirm that the D153del mutation in GNB3 gene targets GNB3 protein to early degradation. Lack of GNB3 signalling causes reduced phosphorylation activity of ERK2 and AKT leading to severe pathological phenotypes such as blindness and renal abnormalities in rge chickens. PMID:21887213

Experimental strategy to discover microbes with gluten-degrading enzyme activities

NASA Astrophysics Data System (ADS)

Helmerhorst, Eva J.; Wei, Guoxian

2014-06-01

Gluten proteins contained in the cereals barley, rye and wheat cause an inflammatory disorder called celiac disease in genetically predisposed individuals. Certain immunogenic gluten domains are resistant to degradation by mammalian digestive enzymes. Enzymes with the ability to target such domains are potentially of clinical use. Of particular interest are gluten-degrading enzymes that would be naturally present in the human body, e.g. associated with resident microbial species. This manuscript describes a selective gluten agar approach and four enzyme activity assays, including a gliadin zymogram assay, designed for the selection and discovery of novel gluten-degrading microorganisms from human biological samples. Resident and harmless bacteria and/or their derived enzymes could potentially find novel applications in the treatment of celiac disease, in the form of a probiotic agent or as a dietary enzyme supplement.

Experimental Strategy to Discover Microbes with Gluten-degrading Enzyme Activities.

PubMed

Helmerhorst, Eva J; Wei, Guoxian

2014-05-05

Gluten proteins contained in the cereals barley, rye and wheat cause an inflammatory disorder called celiac disease in genetically predisposed individuals. Certain immunogenic gluten domains are resistant to degradation by mammalian digestive enzymes. Enzymes with the ability to target such domains are potentially of clinical use. Of particular interest are gluten-degrading enzymes that would be naturally present in the human body, e.g. associated with resident microbial species. This manuscript describes a selective gluten agar approach and four enzyme activity assays, including a gliadin zymogram assay, designed for the selection and discovery of novel gluten-degrading microorganisms from human biological samples. Resident and harmless bacteria and/or their derived enzymes could potentially find novel applications in the treatment of celiac disease, in the form of a probiotic agent or as a dietary enzyme supplement.

Ruxolitinib

MedlinePlus

... by scar tissue and causes decreased blood cell production). Ruxolitinib is in a class of medications called kinase inhibitors. It works by blocking an enzyme that causes scar tissue to form in the bone marrow.

Neurotuberculosis in cattle in southern Brazil.

PubMed

Konradt, Guilherme; Bassuino, Daniele Mariath; Bianchi, Matheus Viezzer; Bandinelli, Marcele Bettim; Driemeier, David; Pavarini, Saulo Petinatti

2016-06-01

Tuberculosis in cattle is a chronic infectious-contagious disease characterized by the development of nodular lesions (granulomas) in mainly the lungs and regional lymph nodes. It is caused by Mycobacterium tuberculosis complex, an acid-fast bacillus (AFB). Tuberculosis in the central nervous system is a rare condition in cattle. Herein, we describe the clinical and pathological findings of six neurotuberculosis cases in cattle diagnosed in Southern Brazil. The average age of the cattle affected was 12 months, and they varied in breed and sex. The clinical history ranged from 5 to 30 days and was characterized by motor incoordination, opisthotonus, blindness, and progression to recumbency. The cattle were euthanized, and grossly, the leptomeninges at the basilar brain showed marked and diffuse expansion, with nodular yellowish lesions ranging in size. On microscopic examination, there were multifocal granulomas located mainly in the meninges, though sometimes extending to adjacent neuropil or existing as isolated granulomas in neuropil. AFBs were observed in the cytoplasm of epithelioid macrophages and multinucleated giant cells through Ziehl-Neelsen histochemical staining and identified as Mycobacterium sp. through immunohistochemistry.

Deciphering the role of the phenylpropanoid metabolism in the tolerance of Capsicum annuum L. to Verticillium dahliae Kleb.

PubMed

Novo, Marta; Silvar, Cristina; Merino, Fuencisla; Martínez-Cortés, Teresa; Lu, Fachuang; Ralph, John; Pomar, Federico

2017-05-01

Verticillium dahliae is an economically relevant soilborne pathogen that causes vascular wilt in several crops, including pepper (Capsicum annuum). Fungal infection is usually visualized as a vascular browning, likely due to the onset of phenylpropanoid metabolism, which also seems to play a crucial role in the tolerance of some pepper varieties. In the current work, the potential function of distinct phenylpropanoid derivatives (suberin, lignin and phenolic compounds) in the pepper tolerance response against V. dahliae, was investigated. Histochemical and biochemical analyses ruled out suberin as a key player in the pepper-fungus interaction. However, changes observed in lignin composition and higher deposition of bound phenolics in infected stems seemed to contribute to the reinforcement of cell walls and the impairment of V. dahliae colonization. Most importantly, this is the first time that the accumulation of the hydroxycinnamic acid amide N-feruloyltyramine was reported in pepper stems in response to a vascular fungus. Fungitoxic activity for that hydroxycinnamate-tyramine conjugate was demonstrated as well. Copyright © 2017 Elsevier B.V. All rights reserved.

An ultrastructural and immunogold localization study of proteoglycans associated with the osteocytes of fetal bone in osteogenesis imperfecta.

PubMed

Sarathchandra, P; Pope, F M; Ali, S Y

1996-06-01

Osteogenesis imperfecta (OI) is a rare, heterogeneous, inherited connective tissue disorder frequently caused by abnormalities of type I collagen. It is characterized by bone fragility, osteopenia, and progressive skeletal deformities. Electron microscopy of three OI type II fetal bone samples revealed numerous large osteocyte lacunae. In addition, there was a perilacunar osteoid-like band of collagen surrounding the osteocytes, which was unmineralized and morphologically unusual. Furthermore, large osteocyte lacunae contained fine particles and filamentous material similar to the expected ultrastructural appearance of proteoglycans. More detailed examination was carried out using histochemical and immunogold localization of proteoglycans at light and ultrastructural levels. These tests and the use of electron probe X-ray microanalysis confirmed that the material in the osteocyte lacunae was proteoglycan. In contrast, in the age- and site-matched normal fetal bone, all the osteocyte lacunae appeared negative for proteoglycan. Proteoglycans are regarded as inhibitors of calcification. Our observation of substantial amounts of proteoglycan in abnormally enlarged osteocytic lacunae of some OI fetal bone suggests association with the abnormal bone of this particular subtype of OI type II.

Whitespotted puffer Arothron hispidus, a new host for lymphocystis in Qingdao Aquarium of China.

PubMed

Sheng, Xiu-zhen; Zhan, Wen-bin; Wang, Yue

2007-03-29

In April 2004 white nodular lesions were found on the fins of whitespotted puffer Arothron hispidus (Linnaeus). Diagnostic studies were carried out to confirm the disease using light and electron microscopy, histochemical methods and PCR. The results revealed that the nodules were composed of giant cells up to 400 microm in diameter. These cells were surrounded by a periodic acid-Schiff (PAS)-positive hyaline capsule containing dot-shaped, Feulgen-positive inclusion bodies in the cytoplasm and an irregular nucleus. Numerous virus particles 200 nm in diameter and with hexagonal profiles were observed in the cytoplasm. These features were consistent with those of lymphocystis disease. Additionally, based on the gene sequences of major capsid protein (MCP) of lymphocystis disease virus (LCDV) from Japanese flounder Paralichthys olivaceus, 2 pair primers were designed; after a nested PCR was performed for detection of LCDV in A. hispidus, a positive amplified product was obtained showing the presence of LCDV. Therefore, the white nodules were the lymphocystis lesions caused by LCDV infection and A. hispidus was demonstrated to be a new host for LCDV.

Does liver damage explain the inverse association between vitamin D status and mortality?

PubMed

Skaaby, Tea; Husemoen, Lise Lotte N; Linneberg, Allan

2013-12-01

Several observational studies have linked vitamin D deficiency with an increased risk of all cause mortality. Vitamin D deficiency is common among patients with liver diseases. In a random sample of the general population, we investigated whether the inverse association between vitamin D status and all-cause mortality could be explained by liver damage as reflected by increased levels of liver enzymes. We included a total of 2649 persons examined in 1993e1994. Vitamin D status was assessed as serum 25-hydroxyvitamin D and liver enzyme levels were measured. Information on all-cause mortality was obtained from the Danish Central Personal Register until July 2011. Median follow-up time was 17.0 years, and there were 736 deaths. Multivariable Cox regression analyses with age as underlying time axis and delayed entry showed lower mortality risk with higher vitamin D levels and this was essentially unaffected by adjustment for liver enzyme levels with hazard ratio, 0.96 (95% confidence interval, 0.93e0.99) for a 10 nmol/L higher vitamin D level. The present study did not support our hypothesis that the well-known association between low vitamin D status and mortality is explained by liver damage as reflected by levels of liver enzymes. 2013 Elsevier Inc. All rights reserved.

Molecular aetiology of primary hyperoxaluria type 1.

PubMed

Danpure, Christopher J

2004-01-01

Primary hyperoxaluria type 1 (PH1) is a rare autosomal-recessive disorder, caused by a deficiency of the liver-specific intermediary-metabolic enzyme alanine:glyoxylate aminotransferase (AGT). AGT deficiency results in increased synthesis and excretion of the metabolic end-product oxalate and the deposition of insoluble calcium oxalate in the kidney and urinary tract. Numerous mutations and polymorphisms have been identified in the gene (AGXT) that encodes AGT, some of which interact synergistically to cause a variety of complex enzyme phenotypes, including AGT intraperoxisomal aggregation, accelerated degradation, and peroxisome-to-mitochondrion mistargeting. The latter is the single most common cause of PH1 and results from the functional interaction between a common Pro11Leu polymorphism and a disease-specific Gly170Arg mutation. The recent solution of the crystal structure of AGT has enabled the effects of several mutations and polymorphisms to be rationalised in terms of their likely effects on AGT conformation. Increased understanding of the molecular aetiology of PH1 has led to significant improvements in all aspects of the clinical management of the disorder, including diagnosis (by enzyme assay of percutaneous needle liver biopsies), prenatal diagnosis (by DNA analysis of chorionic villus samples) and treatment (by liver transplantation as a form of enzyme replacement therapy). Copyright (c) 2004 S. Karger AG, Basel.

Non-specific gastrointestinal features: Could it be Fabry disease?

PubMed

Hilz, Max J; Arbustini, Eloisa; Dagna, Lorenzo; Gasbarrini, Antonio; Goizet, Cyril; Lacombe, Didier; Liguori, Rocco; Manna, Raffaele; Politei, Juan; Spada, Marco; Burlina, Alessandro

2018-05-01

Non-specific gastrointestinal symptoms, including pain, diarrhoea, nausea, and vomiting, can be the first symptoms of Fabry disease. They may suggest more common disorders, e.g. irritable bowel syndrome or inflammatory bowel disease. The confounding clinical presentation and rarity of Fabry disease often cause long diagnostic delays and multiple misdiagnoses. Therefore, specialists involved in the clinical evaluation of non-specific upper and lower gastrointestinal symptoms should recognize Fabry disease as a possible cause of the symptoms, and should consider Fabry disease as a possible differential diagnosis. When symptoms or family history suggest Fabry disease, in men, low alpha-galactosidase A enzyme levels, and in women, specific Fabry mutations confirm the diagnosis. In addition to symptomatic treatments, disease-specific enzyme replacement therapy with recombinant human alpha-galactosidase A enzyme or chaperone therapy (migalastat) in patients with amenable mutations can improve the disease, including gastrointestinal symptoms, and should be initiated as early as possible after Fabry disease has been confirmed; starting enzyme replacement therapy at as young an age as possible after diagnosis improves long-term clinical outcomes. Improved diagnostic tools, such as a modified gastrointestinal symptom rating scale, may facilitate diagnosing Fabry disease in patients with gastrointestinal symptoms of unknown cause and thus assure timely initiation of disease-specific treatment. Copyright © 2018 The Authors. Published by Elsevier Ltd.. All rights reserved.

[Intensity of pentose phosphate metabolism of carbohydrates in various brain areas in normal and starved animals].

PubMed

Kerimov, B F

2002-01-01

The activities of key enzymes of pentose phosphate pathway, glucose-6-phosphate dehydrogenase (G-6 PD) and 6-phosphogluconate dehydrogenase (6-PGD), were studied in cytoplasmatic fractions of brain cortical (limbic, orbital, sensorimotor cortex) and subcortical (myelencefalon, mesencefalon, hypothalamus) structures of rats subjected to starvation for 1, 2, 3, 5 and 7 days. Short-term starvation (1-3 days) caused activation of 6-GPD and 6-PGD both in cortical and subcortical structures. Long-term starvation for 5-7 days caused a decrease of activities of the pentose phosphate pathway enzymes in all studied structures. It is suggested that enzymes of pentose phosphate pathway in nervous tissues are functionally and metabolically related to glutathione system and during starvation they indirectly participate in the regulation lipid peroxidation processes.

Muscle enzyme release does not predict muscle function impairment after triathlon.

PubMed

Margaritis, I; Tessier, F; Verdera, F; Bermon, S; Marconnet, P

1999-06-01

We sought to determine the effects of a long distance triathlon (4 km swim, 120 km bike-ride, and 30 km run) on the four-day kinetics of the biochemical markers of muscle damage, and whether they were quantitatively linked with muscle function impairment and soreness. Data were collected from 2 days before until 4 days after the completion of the race. Twelve triathletes performed the triathlon and five did not. Maximal voluntary contraction (MVC), muscle soreness (DOMS) and total serum CK, CK-MB, LDH, AST and ALT activities were assessed. Significant changes after triathlon completion were found for all muscle damage indirect markers over time (p < 0.0001). MVC of the knee extensor and flexor muscles decreased over time (p < 0.05). There is disparity in the time point at which peak values where reached for DOMS, MVC and enzyme leakage. There is no correlation between serum enzyme leakage, DOMS and MVC impairment which occur after triathlon. Long distance triathlon race caused muscle damage, but extent, as well as muscle recovery cannot be evaluated by the magnitude of changes in serum enzyme activities. Muscle enzyme release cannot be used to predict the magnitude of the muscle function impairment caused by muscle damage.

Cholinergic and nitrergic neuronal networks in the goldfish telencephalon.

PubMed

Giraldez-Perez, Rosa M; Gaytan, Susana P; Pasaro, Rosario

2013-01-01

The general organization of cholinergic and nitrergic elements in the central nervous system seems to be highly conserved among vertebrates, with the involvement of these neurotransmitter systems now well established in sensory, motor and cognitive processing. The goldfish is a widely used animal model in neuroanatomical, neurophysiological, and behavioral research. The purpose of this study was to examine pallial and subpallial cholinoceptive, cholinergic and nitrergic populations in the goldfish telencephalon by means of histochemical and immunohistochemical techniques in order to identify neurons containing acetylcholinesterase (AChE), choline acetyltransferase (ChAT), NADPH-diaphorase (NADPHd), and neuronal nitric oxide synthase (nNOS), and to relate their distribution to their putative functional significance. Regions containing AChE-labeled neurons represented terminal fields of cholinergic inputs as well as a widespread distribution of AChE-related enzymes; these regions also usually contained NADPHd-labeled neurons and often contained small numbers of nNOS-positive cells. However, the ventral subdivisions of the medial and lateral parts of the dorsal telencephalic area, and the ventral and lateral parts of the ventral telencephalic area, were devoid of nNOS-labeled cells. ChAT-positive neurons were found only in the lateral part of the ventral telencephalic area. ChAT- and nNOS-positive fibers exhibited a radial orientation, and were seen as thin axons with en-passant boutons. The distribution of these elements could help to elucidate the role of cholinergic and nitrergic neuronal networks in the goldfish telencephalon.

Physiological characterization of a mouse model of cachexia in colorectal liver metastases.

PubMed

Murphy, Kate T; Struk, Adam; Malcontenti-Wilson, Cathy; Christophi, Christopher; Lynch, Gordon S

2013-05-15

Loss of skeletal muscle mass and function (cachexia) is severe in patients with colorectal liver metastases because of the large increase in resting energy expenditure but remains understudied because of a lack of suitable preclinical models. Our aim was to characterize a novel preclinical model of cachexia in colorectal liver metastases. We tested the hypothesis that mice with colorectal liver metastases would exhibit cachexia, as evidenced by a reduction in liver-free body mass, muscle mass, and physiological impairment. Twelve-week-old male CBA mice received an intrasplenic injection of Ringer solution (sham) or murine colorectal cancer cells (MoCR) to induce colorectal liver metastases. At end-point (20-29 days), the livers of MoCR mice were infiltrated completely with metastases, and MoCR mice had reduced liver-free body mass, muscle mass, and epididymal fat mass compared with sham controls (P < 0.03). MoCR mice exhibited impaired rotarod performance and grip strength (P < 0.03). Histochemical analyses of tibialis anterior muscles from MoCR mice revealed muscle fiber atrophy and reduced oxidative enzyme activity (P < 0.001). Adipose tissue remodeling was evident in MoCR mice, with reduced adipocyte diameter and greater infiltration of nonadipocyte tissue (P < 0.05). These findings reveal the MoCR mouse model exhibits significant cachexia and is a suitable preclinical model of cachexia in colorectal liver metastases. This model should be used for identifying effective treatments for cachexia to improve quality of life and reduce mortality in patients with colorectal liver metastases.

Ethanol Extract of Pinus koraiensis Leaf Ameliorates Alcoholic Fatty Liver via the Activation of LKB1-AMPK Signaling In Vitro and In Vivo.

PubMed

Hong, Sang-Hyuk; Lee, Hyemin; Lee, Hyo-Jung; Kim, Bonglee; Nam, Min-Ho; Shim, Bum-Sang; Kim, Sung-Hoon

2017-05-01

Although Pinus koraiensis leaf (PKL) was reported for its anti-diabetes, anti-obesity and anticancer effects as a folk remedy, the inhibitory effect of PKL on alcoholic fatty liver has never been elucidated yet. This study investigated the molecular mechanisms of PKL on alcoholic fatty liver in HepG2 cells, Sprague Dawley (SD) rats and Imprinting Control Region (ICR) mice. Pinus koraiensis leaf increased phosphorylation of liver kinase B1 (LKB1)/AMP-activated protein kinase signaling, low-density lipoprotein receptor and decreased fatty acid biosynthesis-related proteins such as sterol regulatory element-binding protein 1c, fatty acid synthase, 3-hydroxy-3-methylglutaryl-CoA reductase in HepG2 cells. In SD rats with 25% alcohol-induced fatty liver, PKL suppressed the levels of aspartate aminotransferase and triglyceride and also enhanced the activities of antioxidant enzymes including superoxide dismutase, glutathione peroxidase and glutathione s-transferase compared with untreated control. Furthermore, PKL increased serum alcohol dehydrogenase and serum aldehyde dehydrogenase, but decreased serum alcohol concentration in ICR mice after alcohol administration. Consistently, histochemical analysis revealed that PKL attenuated alcohol-induced fatty liver in SD rats. Overall, these findings suggest that PKL ameliorates alcohol-induced fatty liver via activation of LKB1-AMP-activated protein kinase and modulation of proteins related to lipogenesis synthesis, cholesterol synthesis and fatty acid oxidation. Copyright © 2017 John Wiley & Sons, Ltd. Copyright © 2017 John Wiley & Sons, Ltd.

Alkaline phosphatase in nasal secretion of cattle: biochemical and molecular characterisation.

PubMed

Ghazali, M Faizal; Koh-Tan, H H Caline; McLaughlin, Mark; Montague, Paul; Jonsson, Nicholas N; Eckersall, P David

2014-09-05

Nasal secretion (NS) was investigated as a source of information regarding the mucosal and systemic immune status of cattle challenged by respiratory disease. A method for the collection of substantial volumes (~12 ml) of NS from cattle was developed to establish a reference range of analytes that are present in the NS of healthy cattle. Biochemical profiles of NS from a group of 38 healthy Holstein-Friesian cows revealed high alkaline phosphatase (AP) activity of up to 2392 IU/L. The character and source of the high activity of AP in bovine NS was investigated. Histochemical analysis confirmed the localization of the AP enzyme activity to epithelial cells and serous glands of the nasal respiratory mucosa. Analysis of mRNA levels from nasal mucosa by end point RT-PCR and PCR product sequencing confirmed that the AP was locally produced and is identical at the nucleotide level to the non-specific AP splice variant found in bovine liver, bone and kidney. Analysis by isoelectric focussing confirmed that AP was produced locally at a high level in nasal epithelium demonstrating that AP from nasal secretion and nasal mucosa had similar pI bands, though differing from those of the liver, kidney, bone and intestine, suggesting different post-translational modification (PTM) of AP in these tissues. A nasal isozyme of AP has been identified that is present at a high activity in NS, resulting from local production and showing distinctive PTM and may be active in NS as an anti-endotoxin mediator.

Impairment of Host Liver Repopulation by Transplanted Hepatocytes in Aged Rats and the Release by Short-Term Growth Hormone Treatment.

PubMed

Stock, Peggy; Bielohuby, Maximilian; Staege, Martin S; Hsu, Mei-Ju; Bidlingmaier, Martin; Christ, Bruno

2017-03-01

Hepatocyte transplantation is an alternative to whole liver transplantation. Yet, efficient liver repopulation by transplanted hepatocytes is low in livers of old animals. This restraint might be because of the poor proliferative capacity of aged donor hepatocytes or the regenerative impairment of the recipient livers. The age-dependent liver repopulation by transplanted wild-type hepatocytes was investigated in juvenile and senescent rats deficient in dipeptidyl-peptidase IV. Repopulation was quantified by flow cytometry and histochemical estimation of dipeptidyl-peptidase IV enzyme activity of donor cells in the negative host liver. As a potential pathway involved, expression of cell cycle proteins was assessed. Irrespective of the age of the donor hepatocytes, large cell clusters appeared in juvenile, but only small clusters in senescent host livers. Because juvenile and senescent donor hepatocytes were likewise functional, host-derived factor(s) impaired senescent host liver repopulation. Growth hormone levels were significantly higher in juvenile than in senescent rats, suggesting that growth hormone might promote host liver repopulation. Indeed, short-term treatment with growth hormone augmented senescent host liver repopulation involving the growth hormone-mediated release of the transcriptional blockade of genes associated with cell cycle progression. Short-term growth hormone substitution might improve liver repopulation by transplanted hepatocytes, thus augmenting the therapeutic benefit of clinical hepatocyte transplantation in older patients. Copyright © 2017 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

Melanosome metabolism in the retinal pigmented epithelium of the opossum.

PubMed

Herman, K G; Steinberg, R H

1982-01-01

Melanosomal metabolism, including both formation and degradation of melanosomes, was studied in the retinal pigmented epithelium (RPE) of the adult opossum. The majority of the observations were made on a transitional zone between the tapetal and non-tapetal RPE, the region where melanosome metabolism was at its highest level. Formation of melanosomes, demonstrated ultrastructurally by the presence of stage-II and -III premelanosomes, was also examined autoradiographically following the incorporation of the melanin precursor, dihydroxyphenylalanine. The autoradiographic evidence indicated that many newly formed melanosomes were rapidly incorporated into complexes. Ultrastructural observations suggested that melanosome complexes were formed by at least two methods, via the fusion of melanosomes with phagosomes derived from outer segments of photoreceptors, or by the sequestration of melanosomes by cisternae. A central finding of this study, supported by both ultrastructural and histochemical data, is that there are specialized cellular regions that vary in melanosomal formation and lysosomal activity. Stage-II premelanosomes were observed only in the basal parts of the RPE cells, whereas stage-III and -IV melanosomes were found primarily in the apical RPE. Both ultrastructural and cytochemical observations indicated that degradation of melanosomes occurs only in the basal RPE. These findings are interpreted in terms of the expression of both tapetal and nontapetal characteristics in transitional cells. Finally, this study illustrates the role of lysosomal enzymes in shaping the pattern of pigmentation, and shows that the association of lysosomal activity with melanosomes depends on the functional state of the melanosome.

Development of transgenic finger millet (Eleusine coracana (L.) Gaertn.) resistant to leaf blast disease.

PubMed

Ignacimuthu, S; Ceasar, S Antony

2012-03-01

Finger millet plants conferring resistance to leaf blast disease have been developed by inserting a rice chitinase (chi11) gene through Agrobacterium-mediated transformation. Plasmid pHyg-Chi.11 harbouring the rice chitinase gene under the control of maize ubiquitin promoter was introduced into finger millet using Agrobacterium strain LBA4404 (pSB1). Transformed plants were selected and regenerated on hygromycin-supplemented medium. Transient expression of transgene was confirmed by GUS histochemical staining. The incorporation of rice chitinase gene in R0 and R1 progenies was confirmed by PCR and Southern blot analyses. Expression of chitinase gene in finger millet was confirmed by Western blot analysis with a barley chitinase antibody. A leaf blast assay was also performed by challenging the transgenic plants with spores of Pyricularia grisea. The frequency of transient expression was 16.3% to 19.3%. Stable frequency was 3.5% to 3.9%. Southern blot analysis confirmed the integration of 3.1 kb chitinase gene. Western blot analysis detected the presence of 35 kDa chitinase enzyme. Chitinase activity ranged from 19.4 to 24.8. In segregation analysis, the transgenic R1 lines produced three resistant and one sensitive for hygromycin, confirming the normal Mendelian pattern of transgene segregation. Transgenic plants showed high level of resistance to leaf blast disease compared to control plants. This is the first study reporting the introduction of rice chitinase gene into finger millet for leaf blast resistance.

Immunolocalization of antioxidant enzymes in high-pressure frozen root and stem nodules of Sesbania rostrata.

PubMed

Rubio, Maria C; Becana, Manuel; Kanematsu, Sumio; Ushimaru, Takashi; James, Euan K

2009-01-01

The activities and localizations of superoxide dismutases (SODs) were compared in root and stem nodules of the semi-aquatic legume Sesbania rostrata using gel-activity assays and immunogold labelling, respectively. Nodules were fixed by high-pressure freezing and dehydrated by freeze substitution. Stem nodules showed more total and specific SOD activities than root nodules because of the presence of chloroplastic CuZnSOD. Most of the total SOD activity of stem and root nodules resulted from 'cytosolic' CuZnSOD, localized in the cytoplasm and chromatin, and from MnSOD in the bacteroids and in the mitochondria of vascular tissue. FeSOD was present in nodule plastids and in leaf chloroplasts, and was found to be associated with chromatin. Superoxide production was detected histochemically in the vascular bundles and in the infected tissue of stem and root nodules, whereas peroxide accumulation was observed in the cortical cell walls and intercellular spaces, as well as within the infection threads of both nodule types. These data suggest a role of CuZnSOD and FeSOD in protecting nuclear DNA from reactive oxygen species and/or in modulating gene activity. The enhanced levels of CuZnSOD, MnSOD and superoxide production in vascular bundle cells are consistent with a role of CuZnSOD and superoxide in the lignification of xylem vessels, but also suggest additional functions in coping with superoxide production by the high respiratory activity of parenchyma cells.

A marked animal-vegetal polarity in the localization of Na(+),K(+) -ATPase activity and its down-regulation following progesterone-induced maturation.

PubMed

Mohanty, Basant Kumar; Gupta, Brij L

2012-02-01

The stage-VI Xenopus oocyte has a very distinct animal-vegetal polarity with structural and functional asymmetry. In this study, we show the expression and distribution pattern of Na(+),K(+) -ATPase in stage-VI oocytes, and its changes following progesterone-induced maturation. Using enzyme-specific electron microscopy phosphatase histochemistry, [(3) H]-ouabain autoradiography, and immunofluorescence cytochemistry at light microscopic level, we find that Na(+),K(+) -ATPase activity is mainly confined to the animal hemisphere. Electron microscopy histochemical results also suggest that polarized distribution of Na(+),K(+) -ATPase activity persists following progesterone-induced maturation, and it becomes gradually more polarized towards the animal pole. The time course following progesterone-induced maturation suggests that there is an initial up-regulation and then gradual down-regulation of Na(+),K(+) -ATPase activity leading to germinal vesicle breakdown (GVBD). By GVBD, the Na(+),K(+) -ATPase activity is completely down-regulated due to endocytotic removal of pump molecules from the plasma membrane into the sub-cortical region of the oocyte. This study provides the first direct evidence for a marked asymmetric localization of Na(+),K(+) -ATPase activity in any vertebrate oocyte. Here, we propose that such asymmetry in Na(+),K(+) -ATPase activity in stage-VI oocytes, and their down-regulation following progesterone-induced maturation, is likely to have a role in the active state of the germinal vesicle in stage-VI oocytes and chromosomal condensation after GVBD. Copyright © 2011 Wiley Periodicals, Inc.

Inhibitors of enzymes catalyzing modifications to histone lysine residues: structure, function and activity.

PubMed

Lillico, Ryan; Stesco, Nicholas; Khorshid Amhad, Tina; Cortes, Claudia; Namaka, Mike P; Lakowski, Ted M

2016-05-01

Gene expression is partly controlled by epigenetic mechanisms including histone-modifying enzymes. Some diseases are caused by changes in gene expression that can be mitigated by inhibiting histone-modifying enzymes. This review covers the enzyme inhibitors targeting histone lysine modifications. We summarize the enzymatic mechanisms of histone lysine acetylation, deacetylation, methylation and demethylation and discuss the biochemical roles of these modifications in gene expression and in disease. We discuss inhibitors of lysine acetylation, deacetylation, methylation and demethylation defining their structure-activity relationships and their potential mechanisms. We show that there are potentially indiscriminant off-target effects on gene expression even with the use of selective epigenetic enzyme inhibitors.

Function of the Membrane Water Channel Aquaporin-5 in the Salivary Gland

PubMed Central

Matsuzaki, Toshiyuki; Susa, Taketo; Shimizu, Kinue; Sawai, Nobuhiko; Suzuki, Takeshi; Aoki, Takeo; Yokoo, Satoshi; Takata, Kuniaki

2012-01-01

The process of saliva production in the salivary glands requires transepithelial water transfer from the interstitium to the acinar lumen. There are two transepithelial pathways: the transcellular and paracellular. In the transcellular pathway, the aquaporin water channels induce passive water diffusion across the membrane lipid bilayer. It is well known that aquaporin-5 (AQP5) is expressed in the salivary glands, in which it is mainly localized at the apical membrane of the acinar cells. This suggests the physiological importance of AQP5 in transcellular water transfer. Reduced saliva secretion under pilocarpine stimulation in AQP5-null mice compared with normal mice further indicates the importance of AQP5 in this process, at least in stimulated saliva secretion. Questions remain therefore regarding the role and importance of AQP5 in basal saliva secretion. It has been speculated that there would be some short-term regulation of AQP5 such as a trafficking mechanism to regulate saliva secretion. However, no histochemical evidence of AQP5-trafficking has been found, although some of biochemical analyses suggested that it may occur. There are no reports of human disease caused by AQP5 mutations, but some studies have revealed an abnormal subcellular distribution of AQP5 in patients or animals with xerostomia caused by Sjögren’s syndrome and X-irradiation. These findings suggest the possible pathophysiological importance of AQP5 in the salivary glands. PMID:23209334

Characterization of human arterial tissue affected by atherosclerosis using multimodal nonlinear optical microscopy

NASA Astrophysics Data System (ADS)

Baria, Enrico; Cicchi, Riccardo; Rotellini, Matteo; Nesi, Gabriella; Massi, Daniela; Pavone, Francesco S.

2016-03-01

Atherosclerosis is a widespread cardiovascular disease caused by the deposition of lipids (such as cholesterol and triglycerides) on the inner arterial wall. The rupture of an atherosclerotic plaque, resulting in a thrombus, is one of the leading causes of death in the Western World. Preventive assessment of plaque vulnerability is therefore extremely important and can be performed by studying collagen organization and lipid composition in atherosclerotic arterial tissues. Routinely used diagnostic methods, such as histopathological examination, are limited to morphological analysis of the examined tissues, whereas an exhaustive characterization requires immune-histochemical examination and a morpho-functional approach. Instead, a label-free and non-invasive alternative is provided by nonlinear microscopy. In this study, we combined SHG and FLIM microscopy in order to characterize collagen organization and lipids in human carotid ex vivo tissues affected by atherosclerosis. SHG and TPF images, acquired from different regions within atherosclerotic plaques, were processed through image pattern analysis methods (FFT, GLCM). The resulting information on collagen and cholesterol distribution and anisotropy, combined with collagen and lipids fluorescence lifetime measured from FLIM images, allowed characterization of carotid samples and discrimination of different tissue regions. The presented method can be applied for automated classification of atherosclerotic lesions and plaque vulnerability. Moreover, it lays the foundation for a potential in vivo diagnostic tool to be used in clinical setting.

Characterization and mapping of complementary lesion-mimic genes lm1 and lm2 in common wheat.

PubMed

Yao, Qin; Zhou, Ronghua; Fu, Tihua; Wu, Weiren; Zhu, Zhendong; Li, Aili; Jia, Jizeng

2009-10-01

A lesion-mimic phenotype appeared in a segregating population of common wheat cross Yanzhan 1/Zaosui 30. The parents had non-lesion normal phenotypes. Shading treatment and histochemical analyses showed that the lesions were caused by light-dependent cell death and were not associated with pathogens. Studies over two cropping seasons showed that some lines with more highly expressed lesion-mimic phenotypes exhibited significantly lower grain yields than those with the normal phenotype, but there were no significant effects in the lines with weakly expressed lesion-mimic phenotypes. Among yield traits, one-thousand grain weight was the most affected by lesion-mimic phenotypes. Genetic analysis indicated that this was a novel type of lesion mimic, which was caused by interaction of recessive genes derived from each parent. The lm1 (lesion mimic 1) locus from Zaosui 30 was flanked by microsatellite markers Xwmc674 and Xbarc133/Xbarc147 on chromosome 3BS, at genetic distances of 1.2 and 3.8 cM, respectively, whereas lm2 from Yanzhan 1 was mapped between microsatellite markers Xgwm513 and Xksum154 on chromosome 4BL, at genetic distances of 1.5 and 3 cM, respectively. The linked microsatellite makers identified in this study might be useful for evaluating whether potential parents with normal phenotype are carriers of lesion-mimic alleles.

Primary Systemic Amyloidosis and High Levels of Angiotensin-Converting Enzyme: Two Case Reports

PubMed Central

Praena-Segovia, J.; Sanchez-Gastaldo, A.; Bernabeu-Wittel, M.; Ocete-Pérez, R.; Ávila-Polo, R.; Martino, M. L.

2013-01-01

Infiltrative heart diseases are caused by a heterogeneous group of disorders; amyloidosis and sarcoidosis are two frequent causes of myocardial infiltration, which differ in clinical and biological outcome and treatment issues. The presence of high levels of angiotensin-converting enzyme (ACE) in a patient with infiltrative heart disease may increase suspicion of sarcoidosis. Nevertheless, no mention about increased ACE levels in extracerebral primary systemic amyloidosis is available. We present two cases of primary systemic amyloidosis, which are cardiac involvement and elevated ACE levels. PMID:24826302

CD1 Mouse Retina Is Shielded From Iron Overload Caused by a High Iron Diet

PubMed Central

Bhoiwala, Devang L.; Song, Ying; Cwanger, Alyssa; Clark, Esther; Zhao, Liang-liang; Wang, Chenguang; Li, Yafeng; Song, Delu; Dunaief, Joshua L.

2015-01-01

Purpose High RPE iron levels have been associated with age-related macular degeneration. Mutation of the ferroxidase ceruloplasmin leads to RPE iron accumulation and degeneration in patients with aceruloplasminemia; mice lacking ceruloplasmin and its homolog hephaestin have a similar RPE degeneration. To determine whether a high iron diet (HID) could cause RPE iron accumulation, possibly contributing to RPE oxidative stress in AMD, we tested the effect of dietary iron on mouse RPE iron. Methods Male CD1 strain mice were fed either a standard iron diet (SID) or the same diet with extra iron added (HID) for either 3 months or 10 months. Mice were analyzed with immunofluorescence and Perls' histochemical iron stain to assess iron levels. Levels of ferritin, transferrin receptor, and oxidative stress gene mRNAs were measured by quantitative PCR (qPCR) in neural retina (NR) and isolated RPE. Morphology was assessed in plastic sections. Results Ferritin immunoreactivity demonstrated a modest increase in the RPE in 10-month HID mice. Analysis by qPCR showed changes in mRNA levels of iron-responsive genes, indicating moderately increased iron in the RPE of 10-month HID mice. However, even by age 18 months, there was no Perls' signal in the retina or RPE and no retinal degeneration. Conclusions These findings indicate that iron absorbed from the diet can modestly increase the level of iron deposition in the wild-type mouse RPE without causing RPE or retinal degeneration. This suggests regulation of retinal iron uptake at the blood-retinal barriers. PMID:26275132

An Enzyme from Aristolochia indica Destabilizes Fibrin-β Amyloid Co-Aggregate: Implication in Cerebrovascular Diseases

PubMed Central

Bhattacharjee, Payel; Bhattacharyya, Debasish

2015-01-01

Fibrinogen and β-amyloid (Aβ) peptide independently form ordered aggregates but in combination, they form disordered structures which are resistant to fibrinolytic enzymes like plasmin and cause severity in cerebral amyloid angiopathy (CAA). A novel enzyme of 31.3 kDa has been isolated from the root of the medicinal plant Aristolochia indica that showed fibrinolytic as well as fibrin-Aβ co-aggregate destabilizing properties. This enzyme is functionally distinct from plasmin. Thrombolytic action of the enzyme was demonstrated in rat model. The potency of the plant enzyme in degrading fibrin and fibrin-plasma protein (Aβ, human serum albumin, lysozyme, transthyretin and fibronectin) co-aggregates was demonstrated by atomic force microscopy, scanning electron microscopy and confocal microscopy that showed better potency of the plant enzyme as compared to plasmin. Moreover, the plant enzyme inhibited localization of the co-aggregate inside SH-SY5Y human neuroblastoma cells and also co-aggregate induced cytotoxicity. Plasmin was inefficient in this respect. In the background of limited options for fragmentation of these co-aggregates, the plant enzyme may appear as a potential proteolytic enzyme. PMID:26545113

HISTOCHEMICAL TESTS OF DISCOVERY OF FAT PEROXIDES IN THE EPIDERMIS OF RATS IN LOCAL IRRADIATION

DOE Office of Scientific and Technical Information (OSTI.GOV)

Godlewski, H.

1958-01-01

Fat peroxides were determined according to the method of Dubulo and Duma 0, 3, 6, and 24 hr after single local irradiation of the hip with a dose of 2400 r. Negative results were obtained. However, the peroxides were discovered in irradiated as well as in control animals in places of prolonged restorative degeneration of unknown etiology (panniculus carnosus?).

Targeting Microvascular Pericytes in Angiogenic Vessels of Prostate Cancer

DTIC Science & Technology

2006-04-01

Schlingemann RO. 2004. In vivo angiogenic phenotype of endothelial cells and pericytes induced by vascular endothelial growth factor -a. J Histochem Cytochem...R, McDonald DM. Age-related changes in vascular endothelial growth factor dependency and angiopoietin-1-induced plasti- city of adult blood vessels...hematopoietic progenitor cells and their progeny in vivo . We used the basic fibroblast growth factor (bFGF)- induced mouse corneal neovascularization

The pathology of innactivation in monkeys.

NASA Technical Reports Server (NTRS)

Bourne, G. H.; Golarz De Bourne, M. N.; Mcclure, H.; Keeling, M.

1973-01-01

Progress report on a long-term experiment using rhesus monkeys and designed to study the effects of isolation up to one year, as well as the effects of bed rest simulated by immobilization in a plaster cast for six months. The investigation includes histopathological and histochemical studies of these effects on various internal organs and tissues, and some of the preliminary results of these studies are presented and discussed.

Histological and histochemical studies of the liver of rats flown aboard Kosmos-690 biosatellite. [Prolonged space flight has no effect on the mmorphological changes induced by. gamma. rays in rat liver

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yakovleva, V.I.

1978-10-26

This work is part of a comprehensive study of the biological effects of long-term radiation on rats flown aboard Kosmos-690 for 20.5 days. The results of morphological studies of the rat liver irradiated aboard the biosatellite are discussed.

Detectability and medico-legal value of the gunshot residues in the intracorporeal channel.

PubMed

Migliorini, Arnaldo Stanislao; Andreola, Salvatore; Battistini, Alessio; Gentile, Guendalina; Muccino, Enrico; Vancheri, Giulia; Zoja, Riccardo

2015-11-01

The application of the histochemical stain of sodium rhodizonate to the entrance wound for the detection of the lead (Pb) residues coming from the gunshot may be affected by false positive cases due to the contamination of the environmental Pb. The aim of the Authors is to histochemically search the Pb of GSR in a region which should be more protected by the contamination: the intracorporeal channel. Two hundreds and eighteen serial histological specimens of the intracorporeal channels coming from 25 subjects (dead due to gunshots and being autopsied at the Section of Legal Medicine of the Milan University, in the years 2013-2014) were stained with the sodium rhodizonate and sodium rhodizonate in acid environment (HCl 5%), and then observed by the microscope. The sodium rhodizonate showed a positivity for the Pb residues in the intracorporeal channel, with the detection of the particles within the first 2 cm beyond the entrance wound in 6 cases over the total number of 25 (24%). Victims were characterized by common features: short-barreled weapon; contact shots or short-distance shots; involvement of regions that were not covered by clothing; preservation of the microscopic structure of organs interested by the intracorporeal channel. The searching of GSR in the intracorporeal channel, even in conditions securing a high sensitivity, could represent an important test for the discrimination between an environmental contamination of Pb and the presence of Pb residues by GSR: once confirmed the presence of GSR in the intracorporeal channel by the histochemical analysis, the diagnostic process should require the application of the SEM-EDX for the confirmation of the results. Although not yet studied, this combination could be applied to cadavers exposed to the environment, with advanced post-mortal phenomena permitting at least the suspects of the existence of gunshot wounds at the macroscopic autopsy evaluation. Indeed, in some cases, the putrefaction is so advanced that no suspect of gunshot injuries could be derived from the soft tissue. Copyright © 2015 Elsevier Ltd and Faculty of Forensic and Legal Medicine. All rights reserved.

Inhibition of ligand exchange kinetics via active-site trapping with an antibody fragment.

PubMed

Oyen, David; Steyaert, Jan; Barlow, John N

2014-04-01

We describe the first example of an inhibitory antibody fragment (nanobody ca1697) that binds simultaneously to an enzyme (the enzyme dihydrofolate reductase from Escherichia coli) and its bound substrate (folate). Binding of the antibody to the substrate causes a 20-fold reduction in the rate of folate exchange kinetics. This work opens up the prospect of designing new types of antibody-based inhibitors of enzymes and receptors through suitable design of immunogens.

Defense reactions of bean genotypes to bacterial pathogens in controlled conditions

NASA Astrophysics Data System (ADS)

Uysal, B.; Bastas, K. K.

2018-03-01

This study was focused on the role of antioxidant enzymes and total protein in imparting resistance against common bacterial blight caused by Xanthomonas axonopodis pv. phaseoli (Xap) and halo blight caused by Pseudomonas syringae pv. phaseolicola (Psp) in bean. Activities of Ascorbate peroxidase (APX), Catalase (CAT) and total protein were studied in resistant and susceptible bean genotypes. Five-day-old seedlings were inoculated with a bacterial suspension (108 CFU ml-1) and harvested at different time intervals (0, 12, 24 and 36 up to 72 h) under controlled growing conditions and assayed for antioxidant enzymes and total protein. Temporal increase of CAT, APX enzymes activities showed maximum activity at 12 h after both pathogens inoculation (hpi) in resistant cultivar, whereas in susceptible it increased at 72 h after both pathogens inoculation for CAT and 12, 24 h for APX enzymes. Maximum total protein activities were observed at 12 h and 24 h respectively after Xap, Psp inoculation (hpi) in resistant and maximum activities were observed at 24 h and 72 h respectively after Xap, Psp inoculation (hpi) in susceptible. Increase of antioxidant enzyme and total protein activities might be an important component in the defense strategy of resistance and susceptible bean genotypes against the bacterial infection. These findings suggest that disease protection is proportional to the amount of enhanced CAT, APX enzyme and total protein activity.

A review of enzyme changes in serum and urine due to treatment with drugs (tuberculostatics, contraceptive medication, diagnostics and drugs in real diseases).

PubMed

Haschen, R J

1980-01-01

Serum enzymes are sensitive tools for demonstration of injury to different organs of the body, including lesions that are side effects of drugs or the application of certain compounds for diagnostic purposes. It has been reported that tuberculosis therapy can cause liver damage of the unpredictable drug-induced liver lesion-type. The liver lesions due to tuberculostatics resemble an unspecific or viral hepatitis. Oral contraceptives can also cause hepatic lesions, some of them being quite serious. They have been known to cause hepatosis with or without cholestasis, drug-induced hepatitis, circulatory disturbances such as hepatic peliosis and Budd-Chiari syndrome, and benign and malignant tumors. Therapy control should be instituted in these cases. Enzymes can be used as indicators. A considerable increase of coeruloplasmin (or serum copper) is a useful marker for monitoring contraceptive medication; the aminotransferases initially show a slight increase while later, they may be decreasing to subnormal values. The appearance of serious morphological findings may necessitate delineation of normal ranges and alarm ranges. Serial enzyme determinations are suggested in: 1) high risk patients who have been suffering from hepatitis or intrahepatic cholestasis due to pregnancy or drugs, and 2) in persons complaining of signs or symptoms suggestive of liver involvement. The kidney is another organ which might be influenced by certain drugs leaving the organism via the urinary tract. The presence of enzymes in urine can serve as sensitive indicators of a proximal tubular lesion. The effects of certain hormones on the kidneys are discussed.

Hereditary sensory neuropathy type 1-associated deoxysphingolipids cause neurotoxicity, acute calcium handling abnormalities and mitochondrial dysfunction in vitro.

PubMed

Wilson, Emma R; Kugathasan, Umaiyal; Abramov, Andrey Y; Clark, Alex J; Bennett, David L H; Reilly, Mary M; Greensmith, Linda; Kalmar, Bernadett

2018-05-18

Hereditary sensory neuropathy type 1 (HSN-1) is a peripheral neuropathy most frequently caused by mutations in the SPTLC1 or SPTLC2 genes, which code for two subunits of the enzyme serine palmitoyltransferase (SPT). SPT catalyzes the first step of de novo sphingolipid synthesis. Mutations in SPT result in a change in enzyme substrate specificity, which causes the production of atypical deoxysphinganine and deoxymethylsphinganine, rather than the normal enzyme product, sphinganine. Levels of these abnormal compounds are elevated in blood of HSN-1 patients and this is thought to cause the peripheral motor and sensory nerve damage that is characteristic of the disease, by a largely unresolved mechanism. In this study, we show that exogenous application of these deoxysphingoid bases causes dose- and time-dependent neurotoxicity in primary mammalian neurons, as determined by analysis of cell survival and neurite length. Acutely, deoxysphingoid base neurotoxicity manifests in abnormal Ca 2+ handling by the endoplasmic reticulum (ER) and mitochondria as well as dysregulation of cell membrane store-operated Ca 2+ channels. The changes in intracellular Ca 2+ handling are accompanied by an early loss of mitochondrial membrane potential in deoxysphingoid base-treated motor and sensory neurons. Thus, these results suggest that exogenous deoxysphingoid base application causes neuronal mitochondrial dysfunction and Ca 2+ handling deficits, which may play a critical role in the pathogenesis of HSN-1. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

Aberrant expression and distribution of enzymes of the urea cycle and other ammonia metabolizing pathways in dogs with congenital portosystemic shunts.

PubMed

van Straten, Giora; van Steenbeek, Frank G; Grinwis, Guy C M; Favier, Robert P; Kummeling, Anne; van Gils, Ingrid H; Fieten, Hille; Groot Koerkamp, Marian J A; Holstege, Frank C P; Rothuizen, Jan; Spee, Bart

2014-01-01

The detoxification of ammonia occurs mainly through conversion of ammonia to urea in the liver via the urea cycle and glutamine synthesis. Congenital portosystemic shunts (CPSS) in dogs cause hyperammonemia eventually leading to hepatic encephalopathy. In this study, the gene expression of urea cycle enzymes (carbamoylphosphate synthetase (CPS1), ornithine carbamoyltransferase (OTC), argininosuccinate synthetase (ASS1), argininosuccinate lyase (ASL), and arginase (ARG1)), N-acetylglutamate synthase (NAGS), Glutamate dehydrogenase (GLUD1), and glutamate-ammonia ligase (GLUL) was evaluated in dogs with CPSS before and after surgical closure of the shunt. Additionally, immunohistochemistry was performed on urea cycle enzymes and GLUL on liver samples of healthy dogs and dogs with CPSS to investigate a possible zonal distribution of these enzymes within the liver lobule and to investigate possible differences in distribution in dogs with CPSS compared to healthy dogs. Furthermore, the effect of increasing ammonia concentrations on the expression of the urea cycle enzymes was investigated in primary hepatocytes in vitro. Gene-expression of CPS1, OTC, ASL, GLUD1 and NAGS was down regulated in dogs with CPSS and did not normalize after surgical closure of the shunt. In all dogs GLUL distribution was localized pericentrally. CPS1, OTC and ASS1 were localized periportally in healthy dogs, whereas in CPSS dogs, these enzymes lacked a clear zonal distribution. In primary hepatocytes higher ammonia concentrations induced mRNA levels of CPS1. We hypothesize that the reduction in expression of urea cycle enzymes, NAGS and GLUD1 as well as the alterations in zonal distribution in dogs with CPSS may be caused by a developmental arrest of these enzymes during the embryonic or early postnatal phase.

Effects of silver nanoparticles on soil enzyme activities with and without added organic matter.

PubMed

Peyrot, Caroline; Wilkinson, Kevin J; Desrosiers, Mélanie; Sauvé, Sébastien

2014-01-01

The effects of silver nanoparticles (AgNPs) on terrestrial ecosystems need to be better understood and assessed. Cationic silver (Ag+) has well-documented toxicity against bacteria, but it is not clear what will be the effect of nanoscale Ag. In the present study, the potential effects of AgNPs were investigated in soils by measuring activity of the enzymes phosphomonoesterase, arylsulfatase, β-D-glucosidase, and leucine-aminopeptidase. The toxicity of AgNPs was compared with that of ionic Ag, and the ameliorating effects of soil organic matter were evaluated. To this end, 2 soils with different organic matter contents were artificially contaminated with either AgNPs or Ag-acetate at equivalent total Ag concentrations. In general, enzyme activities were inhibited as a function of the Ag concentration in the soil. In the AgNP exposures, only a small fraction of the AgNP was actually truly dissolved (found in the <1-nm fraction), suggesting that the particulate forms of AgNPs resulted in a significant inhibition of soil enzymes. The addition of organic matter to the soils appeared to enhance enzyme activities; however, the mechanism of organic matter action is not clear given that dissolved Ag concentrations were similar in both the organic-matter–amended and unamended soils. The present study shows that the AgNP produces significant negative effects on the soil enzyme activities tested. The Ag chemical speciation measurements suggested that the AgNP caused greater toxic effects to the soil enzymes at the low Ag concentrations. For the larger concentrations of total soil Ag, causes of the negative effects on enzyme activities are less obvious but suggest that colloidal forms of Ag play a role.

Aberrant Expression and Distribution of Enzymes of the Urea Cycle and Other Ammonia Metabolizing Pathways in Dogs with Congenital Portosystemic Shunts

PubMed Central

van Straten, Giora; van Steenbeek, Frank G.; Grinwis, Guy C. M.; Favier, Robert P.; Kummeling, Anne; van Gils, Ingrid H.; Fieten, Hille; Groot Koerkamp, Marian J. A.; Holstege, Frank C. P.; Rothuizen, Jan; Spee, Bart

2014-01-01

The detoxification of ammonia occurs mainly through conversion of ammonia to urea in the liver via the urea cycle and glutamine synthesis. Congenital portosystemic shunts (CPSS) in dogs cause hyperammonemia eventually leading to hepatic encephalopathy. In this study, the gene expression of urea cycle enzymes (carbamoylphosphate synthetase (CPS1), ornithine carbamoyltransferase (OTC), argininosuccinate synthetase (ASS1), argininosuccinate lyase (ASL), and arginase (ARG1)), N-acetylglutamate synthase (NAGS), Glutamate dehydrogenase (GLUD1), and glutamate-ammonia ligase (GLUL) was evaluated in dogs with CPSS before and after surgical closure of the shunt. Additionally, immunohistochemistry was performed on urea cycle enzymes and GLUL on liver samples of healthy dogs and dogs with CPSS to investigate a possible zonal distribution of these enzymes within the liver lobule and to investigate possible differences in distribution in dogs with CPSS compared to healthy dogs. Furthermore, the effect of increasing ammonia concentrations on the expression of the urea cycle enzymes was investigated in primary hepatocytes in vitro. Gene-expression of CPS1, OTC, ASL, GLUD1 and NAGS was down regulated in dogs with CPSS and did not normalize after surgical closure of the shunt. In all dogs GLUL distribution was localized pericentrally. CPS1, OTC and ASS1 were localized periportally in healthy dogs, whereas in CPSS dogs, these enzymes lacked a clear zonal distribution. In primary hepatocytes higher ammonia concentrations induced mRNA levels of CPS1. We hypothesize that the reduction in expression of urea cycle enzymes, NAGS and GLUD1 as well as the alterations in zonal distribution in dogs with CPSS may be caused by a developmental arrest of these enzymes during the embryonic or early postnatal phase. PMID:24945279

The relevance of cytochrome P450 polymorphism in forensic medicine and akathisia-related violence and suicide.

PubMed

Eikelenboom-Schieveld, Selma J M; Lucire, Yolande; Fogleman, James C

2016-07-01

Adverse drug reactions and interactions are among the major causes of death in the United States. Antidepressants have been reported as causing suicide and homicide and share the class attribute of frequently producing akathisia, a state of severe restlessness associated with thoughts of death and violence. Medical examiners can now identify some pharmacogenetic interactions that cause drugs, deemed safe for most, to be lethal to others. Such deaths do not yet include medication-induced, akathisia-related suicides and homicides. An extrapyramidal side effect, akathisia is a manifestation of drug toxicity whose causes lie, inter alia, in drugs, doses, and co-prescribed medications that inhibit and compete for metabolizing enzymes, which may themselves be defective. In this paper, we report our investigation into adverse drug reactions/interactions in three persons who committed homicide, two also intending suicide, while on antidepressants prescribed for stressful life events. Their histories of medication use, adverse reactions and reasons for changes in medications are presented. DNA samples were screened for variants in the cytochrome P450 gene family; that produce drug metabolizing enzymes. All three cases exhibit genotype-based diminished metabolic capability that, in combination with their enzyme inhibiting/competing medications, decreased metabolism further and are the likely cause of these catastrophic events. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

Protection by Chrysanthemum zawadskii extract from liver damage of mice caused by carbon tetrachloride is maybe mediated by modulation of QR activity

PubMed Central

Seo, Ji Yeon; Lim, Soon Sung; Park, Jia; Lim, Ji-Sun; Kim, Hyo Jung; Kang, Hui Jung; Yoon Park, Jung Han

2010-01-01

Our previous study demonstrated that methanolic extract of Chrysanthemum zawadskii Herbich var. latilobum Kitamura (Compositae) has the potential to induce detoxifying enzymes such as NAD(P)H:(quinone acceptor) oxidoreductase 1 (EC 1.6.99.2) (NQO1, QR) and glutathione S-transferase (GST). In this study we further fractionated methanolic extract of Chrysanthemum zawadskii and investigated the detoxifying enzyme-inducing potential of each fraction. The fraction (CZ-6) shown the highest QR-inducing activity was found to contain (+)-(3S,4S,5R,8S)-(E)-8-acetoxy-4-hydroxy-3-isovaleroyloxy-2-(hexa-2,4-diynyliden)-1,6-dioxaspiro [4,5] decane and increased QR enzyme activity in a dose-dependent manner. Furthermore, CZ-6 fraction caused a dose-dependent enhancement of luciferase activity in HepG2-C8 cells generated by stably transfecting antioxidant response element-luciferase gene construct, suggesting that it induces antioxidant/detoxifying enzymes through antioxidant response element (ARE)-mediated transcriptional activation of the relevant genes. Although CZ-6 fraction failed to induce hepatic QR in mice over the control, it restored QR activity suppressed by CCl4 treatment to the control level. Hepatic injury induced by CCl4 was also slightly protected by pretreatment with CZ-6. In conclusion, although CZ-6 fractionated from methanolic extract of Chrysanthemum zawadskii did not cause a significant QR induction in mice organs such as liver, kidney, and stomach, it showed protective effect from liver damage caused by CCl4. PMID:20461196

Protection by Chrysanthemum zawadskii extract from liver damage of mice caused by carbon tetrachloride is maybe mediated by modulation of QR activity.

PubMed

Seo, Ji Yeon; Lim, Soon Sung; Park, Jia; Lim, Ji-Sun; Kim, Hyo Jung; Kang, Hui Jung; Yoon Park, Jung Han; Kim, Jong-Sang

2010-04-01

Our previous study demonstrated that methanolic extract of Chrysanthemum zawadskii Herbich var. latilobum Kitamura (Compositae) has the potential to induce detoxifying enzymes such as NAD(P)H:(quinone acceptor) oxidoreductase 1 (EC 1.6.99.2) (NQO1, QR) and glutathione S-transferase (GST). In this study we further fractionated methanolic extract of Chrysanthemum zawadskii and investigated the detoxifying enzyme-inducing potential of each fraction. The fraction (CZ-6) shown the highest QR-inducing activity was found to contain (+)-(3S,4S,5R,8S)-(E)-8-acetoxy-4-hydroxy-3-isovaleroyloxy-2-(hexa-2,4-diynyliden)-1,6-dioxaspiro [4,5] decane and increased QR enzyme activity in a dose-dependent manner. Furthermore, CZ-6 fraction caused a dose-dependent enhancement of luciferase activity in HepG2-C8 cells generated by stably transfecting antioxidant response element-luciferase gene construct, suggesting that it induces antioxidant/detoxifying enzymes through antioxidant response element (ARE)-mediated transcriptional activation of the relevant genes. Although CZ-6 fraction failed to induce hepatic QR in mice over the control, it restored QR activity suppressed by CCl(4) treatment to the control level. Hepatic injury induced by CCl(4) was also slightly protected by pretreatment with CZ-6. In conclusion, although CZ-6 fractionated from methanolic extract of Chrysanthemum zawadskii did not cause a significant QR induction in mice organs such as liver, kidney, and stomach, it showed protective effect from liver damage caused by CCl(4).

HIBCH mutations can cause Leigh-like disease with combined deficiency of multiple mitochondrial respiratory chain enzymes and pyruvate dehydrogenase.

PubMed

Ferdinandusse, Sacha; Waterham, Hans R; Heales, Simon J R; Brown, Garry K; Hargreaves, Iain P; Taanman, Jan-Willem; Gunny, Roxana; Abulhoul, Lara; Wanders, Ronald J A; Clayton, Peter T; Leonard, James V; Rahman, Shamima

2013-12-04

Deficiency of 3-hydroxy-isobutyryl-CoA hydrolase (HIBCH) caused by HIBCH mutations is a rare cerebral organic aciduria caused by disturbance of valine catabolism. Multiple mitochondrial respiratory chain (RC) enzyme deficiencies can arise from a number of mechanisms, including defective maintenance or expression of mitochondrial DNA. Impaired biosynthesis of iron-sulphur clusters and lipoic acid can lead to pyruvate dehydrogenase complex (PDHc) deficiency in addition to multiple RC deficiencies, known as the multiple mitochondrial dysfunctions syndrome. Two brothers born to distantly related Pakistani parents presenting in early infancy with a progressive neurodegenerative disorder, associated with basal ganglia changes on brain magnetic resonance imaging, were investigated for suspected Leigh-like mitochondrial disease. The index case had deficiencies of multiple RC enzymes and PDHc in skeletal muscle and fibroblasts respectively, but these were normal in his younger brother. The observation of persistently elevated hydroxy-C4-carnitine levels in the younger brother led to suspicion of HIBCH deficiency, which was investigated by biochemical assay in cultured skin fibroblasts and molecular genetic analysis. Specific spectrophotometric enzyme assay revealed HIBCH activity to be below detectable limits in cultured skin fibroblasts from both brothers. Direct Sanger sequence analysis demonstrated a novel homozygous pathogenic missense mutation c.950G

Marine Sponges and Bacteria as Challenging Sources of Enzyme Inhibitors for Pharmacological Applications

PubMed Central

Ruocco, Nadia; Costantini, Susan; Palumbo, Flora; Costantini, Maria

2017-01-01

Enzymes play key roles in different cellular processes, for example, in signal transduction, cell differentiation and proliferation, metabolic processes, DNA damage repair, apoptosis, and response to stress. A deregulation of enzymes has been considered one of the first causes of several diseases, including cancers. In the last several years, enzyme inhibitors, being good candidates as drugs in the pathogenic processes, have received an increasing amount of attention for their potential application in pharmacology. The marine environment is considered a challenging source of enzyme inhibitors for pharmacological applications. In this review, we report on secondary metabolites with enzyme inhibitory activity, focusing our attention on marine sponges and bacteria as promising sources. In the case of sponges, we only reported the kinase inhibitors, because this class was the most representative isolated so far from these marine organisms. PMID:28604647

Effects of Acifluorfen on Endogenous Antioxidants and Protective Enzymes in Cucumber (Cucumis sativus L.) Cotyledons

PubMed Central

Kenyon, William H.; Duke, Stephen O.

1985-01-01

The herbicide acifluorfen (2-chloro-4-(trifluoromethyl)phenoxy-2-nitrobenzoate) causes strong photooxidative destruction of pigments and lipids in sensitive plant species. Antioxidants and oxygen radical scavengers slow the bleaching action of the herbicide. The effect of acifluorfen on glutathione and ascorbate levels in cucumber (Cucumis sativus L.) cotyledon discs was investigated to assess the relationship between herbicide activity and endogenous antioxidants. Acifluorfen decreased the levels of glutathione and ascorbate over 50% in discs exposed to less than 1.5 hours of white light (450 microeinsteins per square meter per second). Coincident increases in dehydroascorbate and glutathione disulfide were not observed. Acifluorfen also caused the rapid depletion of ascorbate in far-red light grown plants which were photosynthetically incompetent. Glutathione reductase, dehydroascorbate reductase, superoxide dismutase, ascorbate oxidase, ascorbate free radical reductase, peroxidase, and catalase activities rapidly decreased in acifluorfen-treated tissue exposed to white light. None of the enzymes were inhibited in vitro by the herbicide. Acifluorfen causes irreversible photooxidative destruction of plant tissue, in part, by depleting endogenous antioxidants and inhibiting the activities of protective enzymes. PMID:16664506

Infrared-thermographic screening of the activity and enantioselectivity of enzymes.

PubMed

Reetz, M T; Hermes, M; Becker, M H

2001-05-01

The infrared radiation caused by the heat of reaction of an enantioselective enzyme-catalyzed transformation can be detected by modern photovoltaic infrared (IR)-thermographic cameras equipped with focal-plane array detectors. Specifically, in the lipase-catalyzed enantioselective acylation of racemic 1-phenylethanol, the (R)- and (S)-substrates are allowed to react separately in the wells of microtiter plates, the (R)-alcohol showing hot spots in the IR-thermographic images. Thus, highly enantioselective enzymes can be identified at kinetic resolution.

Phytotoxicity of pesticides mancozeb and chlorpyrifos: correlation with the antioxidative defence system in Allium cepa.

PubMed

Fatma, Firdos; Verma, Sonam; Kamal, Aisha; Srivastava, Alka

2018-02-01

Pesticides are a group of chemical substances which are widely used to improve agricultural production. However, these substances could be persistent in soil and water, accumulative in sediment or bio-accumulative in biota depending on their solubility, leading to different types of environmental pollution. The present study was done to assess the impact of pesticides-mancozeb and chlorpyrifos, via morphological and physiological parameters using Allium cepa test system. Phytotoxic effects of pesticides were examined via germination percentage, survival percentage, root and shoot length, root shoot length ratio, seedling vigor index, percentage of phytotoxicity and tolerance index. Oxidative stress on Allium seedlings caused by pesticides was also assessed by investigating the activity of antioxidative enzymes viz. catalase, peroxidase and superoxide dismutase. Correlation was worked out between morphological parameters and antioxidative enzymes to bring out the alliance between them. Mancozeb and chlorpyrifos concentrations were significantly and positively correlated with the activity of antioxidative enzymes and negatively correlated with morphological parameters. Significant positive correlation between various morphological parameters showed their interdependency. However, negative correlation was obtained between activity of antioxidative enzymes and morphological parameters. The enzymes however, showed positive correlation with each other. Based on our result we can conclude that all morphological parameters were adversely affected by the two pesticides as reflected by phytotoxicity in Allium . Their negative correlation with activity of antioxidative enzymes indicates that upregulation of antioxidative enzymes is not sufficient to overcome the toxic effect, thereby signifying the threat being caused by the regular use of these pesticides.

Rice Blast Fungus (Magnaporthe oryzae) Infects Arabidopsis via a Mechanism Distinct from That Required for the Infection of Rice1[W][OA

PubMed Central

Park, Ju-Young; Jin, Jianming; Lee, Yin-Won; Kang, Seogchan; Lee, Yong-Hwan

2009-01-01

Magnaporthe oryzae is a hemibiotrophic fungal pathogen that causes rice (Oryza sativa) blast. Although M. oryzae as a whole infects a wide variety of monocotyledonous hosts, no dicotyledonous plant has been reported as a host. We found that two rice pathogenic strains of M. oryzae, KJ201 and 70-15, interacted differentially with 16 ecotypes of Arabidopsis (Arabidopsis thaliana). Strain KJ201 infected all ecotypes with varying degrees of virulence, whereas strain 70-15 caused no symptoms in certain ecotypes. In highly susceptible ecotypes, small chlorotic lesions appeared on infected leaves within 3 d after inoculation and subsequently expanded across the affected leaves. The fungus produced spores in susceptible ecotypes but not in resistant ecotypes. Fungal cultures recovered from necrotic lesions caused the same symptoms in healthy plants, satisfying Koch's postulates. Histochemical analyses showed that infection by the fungus caused an accumulation of reactive oxygen species and eventual cell death. Similar to the infection process in rice, the fungus differentiated to form appressorium and directly penetrated the leaf surface in Arabidopsis. However, the pathogenic mechanism in Arabidopsis appears distinct from that in rice; three fungal genes essential for pathogenicity in rice played only limited roles in causing disease symptoms in Arabidopsis, and the fungus seems to colonize Arabidopsis as a necrotroph through the secretion of phytotoxic compounds, including 9,12-octadecadienoic acid. Expression of PR-1 and PDF1.2 was induced in response to infection by the fungus, suggesting the activation of salicylic acid- and jasmonic acid/ethylene-dependent signaling pathways. However, the roles of these signaling pathways in defense against M. oryzae remain unclear. In combination with the wealth of genetic and genomic resources available for M. oryzae, this newly established pathosystem allows comparison of the molecular and cellular mechanisms underlying pathogenesis and host defense in two well-studied model plants. PMID:18987215

Expression of digestive enzymes and nutrient transporters in Eimeria acervulina-challenged layers and broilers

USDA-ARS?s Scientific Manuscript database

Avian coccidiosis is a disease caused by the intestinal protozoa Eimeria. Eimeria-infected chickens develop lesions in the intestinal mucosa, which result in reduced feed efficiency and body weight gain. This growth reduction may be due to changes in expression of digestive enzymes and nutrient tran...

Expression of an antimicrobial peptide, digestive enzymes and nutrient transporters in the intestine of E. praecox-infected chickens

USDA-ARS?s Scientific Manuscript database

Coccidiosis is a major intestinal disease of poultry, caused by several species of the protozoan Eimeria. The objective of this study was to examine changes in expression of digestive enzymes, nutrient transporters and an antimicrobial peptide following an Eimeria praecox challenge of chickens at d...

Tissue specific distribution of pyrimidine deoxynucleoside salvage enzymes shed light on the mechanism of mitochondrial DNA depletion.

PubMed

Wang, L; Eriksson, S

2010-06-01

Deficiency in thymidine kinase 2 (TK2) activity due to genetic alterations caused tissue specific mitochondrial DNA (mtDNA) depletion syndrome with symptoms resembling these of AIDS patients treated with nucleoside analogues. Mechanisms behind this mitochondrial effects is still not well understood. With rat as a model we isolated mitochondrial and cytosolic fractions from major organs and studied enzymes involved in thymidine (dT) and deoxycytidine (dC) phosphorylation by using ionic exchange column chromatography. A cytosolic form of TK2 was identified in all tested tissues in addition to mitochondrial TK2. TK1 was detected in liver and spleen cytosolic extracts while dCK was found in liver, spleen and lung cytosolic extracts. Thus, the nature of dT and dC salvage enzymes in each tissue type was determined. In most tissues TK2 is the only salvage enzyme present except liver and spleen. These results may help to explain the mechanisms of mitochondrial toxicity of antiviral nucleoside analogues and mtDNA depletion caused by TK2 deficiency.

Ochratoxin degradation and adsorption caused by astaxanthin-producing yeasts.

PubMed

Péteri, Z; Téren, J; Vágvölgyi, C; Varga, J

2007-05-01

Ochratoxin degrading and adsorbing activities of Phaffia rhodozyma and Xanthophyllomyces dendrorhous isolates were tested. P. rhodozyma CBS 5905 degraded more than 90% of ochratoxin A (OTA) in 15 days at 20 degrees C. The data presented indicate that P. rhodozyma is able to convert OTA to ochratoxin alpha, and this conversion is possibly mediated by an enzyme related to carboxypeptidases. Chelating agents like EDTA and 1,10-phenanthroline inhibited OTA degradation caused by P. rhodozyma indicating that the carboxypeptidase is a metalloprotease, similarly to carboxypeptidase A. The temperature optimum of this enzyme was found to be above 30 degrees C, which is much higher than the temperature optimum for growth of P. rhodozyma cells, which is around 20 degrees C. The enzyme responsible for ochratoxin degradation was found to be cell-bound. Besides, both viable and heat-treated (dead) P. rhodozyma cells were also able to adsorb significant amounts (up to 250 ng ml(-1)) of OTA. Heat treatment enhanced OTA adsorbing activities of the cells. Further studies are in progress to identify the enzyme responsible for OTA degradation in P. rhodozyma.

Bioinformatic Analysis of the Human Recombinant Iduronate 2-Sulfate Sulfatase

PubMed Central

Morales-Álvarez, Edwin D.; Rivera-Hoyos, Claudia M.; Landázuri, Patricia; Poutou-Piñales, Raúl A.; Pedroza-Rodríguez, Aura M.

2016-01-01

Mucopolysaccharidosis type II is a human recessive disease linked to the X chromosome caused by deficiency of lysosomal enzyme Iduronate 2-Sulfate Sulfatase (IDS), which leads to accumulation of glycosaminoglycans in tissues and organs. The human enzyme has been expressed in Escherichia coli and Pichia pastoris in attempt to develop more successful expression systems that allow the production of recombinant IDS for Enzyme Replacement Therapy (ERT). However, the preservation of native signal peptide in the sequence has caused conflicts in processing and recognition in the past, which led to problems in expression and enzyme activity. With the main object being the improvement of the expression system, we eliminate the native signal peptide of human recombinant IDS. The resulting sequence showed two modified codons, thus, our study aimed to analyze computationally the nucleotide sequence of the IDSnh without signal peptide in order to determine the 3D structure and other biochemical properties to compare them with the native human IDS (IDSnh). Results showed that there are no significant differences between both molecules in spite of the two-codon modifications detected in the recombinant DNA sequence. PMID:27335624

The effect of glycosaminoglycan enzymes and proteases on the viscosity of alpaca seminal plasma and sperm function.

PubMed

Kershaw-Young, C M; Stuart, C; Evans, G; Maxwell, W M C

2013-05-01

In order to advance the development of cryopreservation and other assisted reproductive technologies in camelids it is necessary to eliminate the viscous component of the seminal plasma without impairing sperm function. It has been postulated that glycosaminoglycans (GAGs) or proteoglycans are responsible for this viscosity. This study investigated the effect of the GAG enzymes hyaluronidase, chondroitinase ABC and keratanase and the proteases papain and proteinase K on seminal plasma viscosity and sperm function in order to aid identification of the cause of seminal plasma viscosity and propose methods for the reduction of viscosity. Sperm motility, DNA integrity, acrosome integrity and viability were assessed during 2h incubation. All enzymes reduced seminal plasma viscosity compared to control (P<0.001) although papain was most effective, completely eliminating viscosity within 30 min of treatment. Sperm motility and DNA integrity was not affected by enzyme treatment. The proportion of viable, acrosome intact sperm was reduced in all enzyme treated samples except those treated with papain (P<0.001). These findings suggest that proteins, not GAGs are the main cause of alpaca seminal plasma viscosity. Papain treatment of alpaca semen may be a suitable technique for reduction of seminal plasma viscosity prior to sperm cryopreservation. Copyright © 2013 Elsevier B.V. All rights reserved.

Structural and functional features of enzymes of Mycobacterium tuberculosis peptidoglycan biosynthesis as targets for drug development

PubMed Central

Moraes, Gleiciane Leal; Gomes, Guelber Cardoso; de Sousa, Paulo Robson Monteiro; Alves, Cláudio Nahum; Govender, Thavendran; Kruger, Hendrik G.; Maguire, Glenn E.M.; Lamichhane, Gyanu; Lameira, Jerônimo

2015-01-01

SUMMARY Tuberculosis (TB) is the second leading cause of human mortality from infectious diseases worldwide. The WHO reported 1.3 million deaths and 8.6 million new cases of TB in 2012. Mycobacterium tuberculosis (M. tuberculosis), the infectious bacteria that causes TB, is encapsulated by a thick and robust cell wall. The innermost segment of the cell wall is comprised of peptidoglycan, a layer that is required for survival and growth of the pathogen. Enzymes that catalyse biosynthesis of the peptidoglycan are essential and are therefore attractive targets for discovery of novel antibiotics as humans lack similar enzymes making it possible to selectively target bacteria only. In this paper, we have reviewed the structures and functions of enzymes GlmS, GlmM, GlmU, MurA, MurB, MurC, MurD, MurE and MurF from M. tuberculosis that are involved in peptidoglycan biosynthesis. In addition, we report homology modelled 3D structures of those key enzymes from M. tuberculosis of which the structures are still unknown. We demonstrated that natural substrates can be successfully docked into the active sites of the GlmS and GlmU respectively. It is therefore expected that the models and the data provided herein will facilitate translational research to develop new drugs to treat TB. PMID:25701501

Targeting of Several Glycolytic Enzymes Using RNA Interference Reveals Aldolase Affects Cancer Cell Proliferation through a Non-glycolytic Mechanism

PubMed Central

Lew, Carolyn Ritterson; Tolan, Dean R.

2012-01-01

In cancer, glucose uptake and glycolysis are increased regardless of the oxygen concentration in the cell, a phenomenon known as the Warburg effect. Several (but not all) glycolytic enzymes have been investigated as potential therapeutic targets for cancer treatment using RNAi. Here, four previously untargeted glycolytic enzymes, aldolase A, glyceraldehyde 3-phosphate dehydrogenase, triose phosphate isomerase, and enolase 1, are targeted using RNAi in Ras-transformed NIH-3T3 cells. Of these enzymes, knockdown of aldolase causes the greatest effect, inhibiting cell proliferation by 90%. This defect is rescued by expression of exogenous aldolase. However, aldolase knockdown does not affect glycolytic flux or intracellular ATP concentration, indicating a non-metabolic cause for the cell proliferation defect. Furthermore, this defect could be rescued with an enzymatically dead aldolase variant that retains the known F-actin binding ability of aldolase. One possible model for how aldolase knockdown may inhibit transformed cell proliferation is through its disruption of actin-cytoskeleton dynamics in cell division. Consistent with this hypothesis, aldolase knockdown cells show increased multinucleation. These results are compared with other studies targeting glycolytic enzymes with RNAi in the context of cancer cell proliferation and suggest that aldolase may be a useful target in the treatment of cancer. PMID:23093405

alpha-Galactosidase from Bacillus megaterium VHM1 and its application in removal of flatulence-causing factors from soymilk.

PubMed

Patil, Aravind Goud G; K, Praveen Kumar S; Mulimani, Veerappa H; Veeranagouda, Yaligara; Lee, Kyoung

2010-11-01

A bacterial strain capable of producing extracellular alpha-galactosidase was isolated from sugar cane industrial waste soil sample. Microbiological, physiological, and biochemical studies revealed that isolate belonged to Bacillus sp,. Furthermore, 16S rDNA sequence analysis of new isolates was identified as Bacillus megaterium VHM1. The production of alpha-galactosidase was optimized by various physical culture conditions. Guar gum and yeast extract acted as the best carbon and nitrogen source, respectively for the production of alpha-galactosidase. The enzyme showed an optimum pH at 7.5 and was stable over a pH between 5 and 9. The enzyme was optimally active in 55degreesC and the enzyme was thermostable with half life of 120 minutes at 55 degrees C and lost their 90%, residual activity in 120 minutes at 60 degrees C. alpha-Galactosidase was strongly inhibited by Ag2, Cu2, and Hg2+ at 1mM concentration. The metal ions Fe2, Mn2+, and Mg2+ had no effect on alpha-galactosidase activity, Zn2+,Ni2+, and Ca2+ reduced the enzyme activity slightly. The B megaterium VHM1 enzyme treatment completely hydrolyzed flatulence-causing sugars of soymilk within one and half hour.

A Combination Histochemical and Autoradiographic Method for Analysis of Enzymatic and Proliferative Responses to Immunoreactive Osteoinducer

DTIC Science & Technology

1988-01-15

DNA replication and alkaline phosphate activity in the responding cells was shown to result in effective differential labeling of these features in mildly fixed tissue sections. Application of this method with monoclonal antibodies specific for induction-associated determinants and with modifications to permit ultrastructural analyses may provide important information relevant to the mechanism of matrix-induced bone formation. Keywords: Osteogenic implants; Immunohistochemistry;

Glycoconjugate sugar residues in the chick embryo developing lung: a lectin histochemical study.

PubMed

Gheri, G; Sgambati, E; Bryk, S G

2000-03-01

A lectin histochemical study was performed to investigate the distribution and changes of the oligosaccharidic component of the glycoconjugates in the lung of chick embryos, of 1-day-old chick, and of the adult animal. For this purpose, a battery of seven horseradish peroxidase-conjugated lectins (PNA, SBA, DBA, WGA, Con A, LTA, and UEA I) were employed. During the first phase of parabronchi and atria formation, D-galactose-(beta1-->3)-N-acetyl-D-galactosamine, beta-N-acetyl-D-galactosamine, D-glucosamine, alpha-D-mannose, and sialic acid, present at the level of the surface and of cytoplasmic granules of the lining epithelial cells, seem to play a role in regulating morphogenetic phenomena. In the subsequent phases, the parabronchial lumen and the atrial cavities were characterized by the presence of lectin-reactive material rich in terminal D-galactose-(beta1-->3)-N-acetyl-D-galactosamine, beta-N-acetyl-D-galactosamine, D-glucosamine and alpha-D-mannose. From day 18 onwards and immediately after hatching, the free border of the cells lining the air capillaries was characterized by the presence of beta-N-acetyl-D-galactosamine and alpha-D-mannose. The appearance of these sugar residues was concomitant with the beginning of respiratory activity. Copyright 2000 Wiley-Liss, Inc.

Locus ceruleus neurons in people with autism contain no histochemically-detectable mercury.

PubMed

Pamphlett, Roger; Kum Jew, Stephen

2016-02-01

Exposure to environmental mercury has been proposed to play a part in autism. Mercury is selectively taken up by the human locus ceruleus, a region of the brain that has been implicated in autism. We therefore looked for the presence of mercury in the locus ceruleus of people who had autism, using the histochemical technique of autometallography which can detect nanogram amounts of mercury in tissues. In addition, we sought evidence of damage to locus ceruleus neurons in autism by immunostaining for hyperphosphorylated tau. No mercury was found in any neurons of the locus ceruleus of 6 individuals with autism (5 male, 1 female, age range 16-48 years). Mercury was present in locus ceruleus neurons in 7 of 11 (64%) age-matched control individuals who did not have autism, which is significantly more than in individuals with autism. No increase in numbers of locus ceruleus neurons containing hyperphosphorylated tau was detected in people with autism. In conclusion, most people with autism have not been exposed early in life to quantities of mercury large enough to be found later in adult locus ceruleus neurons. Human locus ceruleus neurons are sensitive indicators of mercury exposure, and mercury appears to remain in these neurons indefinitely, so these findings do not support the hypothesis that mercury neurotoxicity plays a role in autism.

Peripheral muscle composition and health status in patients with COPD.

PubMed

Montes de Oca, María; Torres, Sonia H; Gonzalez, Yudith; Romero, Elizabeth; Hernández, Noelina; Mata, Abdón; Tálamo, Carlos

2006-10-01

The present study evaluated the relationship between health status (HS) and peripheral muscle histochemical characteristics in chronic obstructive pulmonary disease (COPD), and identified selected independent respiratory and extrapulmonary variables that predicted the HS of these patients. Cross-sectional study. Outpatient respiratory clinic of a university hospital. We studied 29 patients (63+/-10 yrs) with a forced expiratory volume in 1s (FEV1) of 39+/-12%. All patients underwent vastus lateralis muscle biopsies for histochemical analysis. They also had spirometry, arterial blood gas analysis, body mass index (BMI), dyspnea determined with the MMRC scale and responded to the St. George's Respiratory Questionnaire (SGRQ) for HS assessment. SGRQ total score correlated with fiber type distribution. A stepwise multiple regression identified three independent predictors of SGRQ total score: type I fiber proportion, BMI, and FEV1; r = 0.78 and r2 = 0.61. These results indicate that impaired HS in COPD is related to the peripheral muscle changes characterized by less type I fibers proportion. The findings argue in favor of an important contribution of the systemic consequences on the HS in COPD independently from the airflow limitation severity, and help to explain the observation of the poor correlation between the degree of airflow limitation and SGRQ total score.

In Situ Histochemical Localisation of Alkaloids and Acetogenins in the Endosperm and Embryonic Axis of Annona Macroprophyllata Donn. Sm. Seeds During Germination

PubMed Central

Brechú-Franco, A.E.; Laguna-Hernández, G.; De la Cruz-Chacón, I.; González-Esquinca, A.R.

2016-01-01

Currently, the Annonaceae family is characterised by the production of acetogenins (ACGs), and also by the biosynthesis of alkaloids, primarily benzylisoquinolines derived from tyrosine. The objective of this study was to confirm the presence of alkaloids and acetogenins in the idioblasts of the endosperm and the embryonic axis of A. macroprophyllata seeds in germination. The Dragendorff, Dittmar, Ellram, and Lugol reagents were used to test for alkaloids, and Kedde’s reagent was used to determine the presence of acetogenins in fresh sections of the endosperm and embryonic axis of seeds after twelve days of germination. A positive reaction was observed for all the reagents, and the presence of alkaloids and acetogenins was confirmed in the idioblasts of the endosperm and those involved in the differentiation of the embryonic axis of the developing seedling. We concluded that the idioblasts store both metabolites, acetogenins and alkaloids. Beginning at differentiation, the idioblasts of the embryonic axis simultaneously biosynthesise acetogenins and alkaloids that are characteristic of the species during the development of the seedling. The method used here can be applied to histochemically confirm the presence of acetogenins and alkaloids in tissues and structures of the plant in different stages of its life cycle. PMID:26972713

Molecular characterization and histochemical demonstration of salmon olfactory marker protein in the olfactory epithelium of lacustrine sockeye salmon (Oncorhynchus nerka).

PubMed

Kudo, H; Doi, Y; Ueda, H; Kaeriyama, M

2009-09-01

Despite the importance of olfactory receptor neurons (ORNs) for homing migration, the expression of olfactory marker protein (OMP) is not well understood in ORNs of Pacific salmon (genus Oncorhynchus). In this study, salmon OMP was characterized in the olfactory epithelia of lacustrine sockeye salmon (O. nerka) by molecular biological and histochemical techniques. Two cDNAs encoding salmon OMP were isolated and sequenced. These cDNAs both contained a coding region encoding 173 amino acid residues, and the molecular mass of the two proteins was calculated to be 19,581.17 and 19,387.11Da, respectively. Both amino acid sequences showed marked homology (90%). The protein and nucleotide sequencing demonstrates the existence of high-level homology between salmon OMPs and those of other teleosts. By in situ hybridization using a digoxigenin-labeled salmon OMP cRNA probe, signals for salmon OMP mRNA were observed preferentially in the perinuclear regions of the ORNs. By immunohistochemistry using a specific antibody to salmon OMP, OMP-immunoreactivities were noted in the cytosol of those neurons. The present study is the first to describe cDNA cloning of OMP in salmon olfactory epithelium, and indicate that OMP is a useful molecular marker for the detection of the ORNs in Pacific salmon.

Stromal characterization and comparison of odontogenic cysts and odontogenic tumors using picrosirius red stain and polarizing microscopy: A retrospective and histochemical study.

PubMed

Jahagirdar, P B; Kale, A D; Hallikerimath, S

2015-01-01

Odontogenic lesions represent a range of conditions, the features of which probably depend on the stage of induction towards tooth formation reached prior to neoplastic or hamartomatous proliferation. It has been also suggested that inductive changes may allow progression from one type of odontogenic tumor to another. The epithelium also plays an important role in the pathogenesis of these lesions; even stroma is likely to play an equally important role in the pathogenesis and biological behavior. So, this study was performed to investigate, compare, and correlate different types of collagen fibers in odontogenic cysts and odontogenic tumors. Thirty each pre-diagnosed odontogenic cysts and tumors were histochemically analyzed using a special stain (Picrosirius red stain) and polarizing microscopy. Seven cases (99%) of inflammatory cysts exhibited predominantly greenish-yellow birefringence indicating procollagen, intermediate, or pathologic collagen fibers suggestive of loosely packed collagen fibers. Predominant yellowish-orange birefringence exhibited by 21 cases (99%) of developmental cysts was comparable to the yellowish-orange and orangish-red to red birefringence exhibited by odontogenic tumors suggesting tightly packed fibers. The Picrosirius red stain in conjunction with polarizing microscopy serves as a specific and sensitive tool in characterizing collagen fibers in odontogenic cysts and odontogenic tumor.

An efficient in planta transformation of Jatropha curcas (L.) and multiplication of transformed plants through in vivo grafting.

PubMed

Jaganath, Balusamy; Subramanyam, Kondeti; Mayavan, Subramanian; Karthik, Sivabalan; Elayaraja, Dhandapani; Udayakumar, Rajangam; Manickavasagam, Markandan; Ganapathi, Andy

2014-05-01

An efficient and reproducible Agrobacterium-mediated in planta transformation was developed in Jatropha curcas. The various factors affecting J. curcas in planta transformation were optimized, including decapitation, Agrobacterium strain, pin-pricking, vacuum infiltration duration and vacuum pressure. Simple vegetative in vivo cleft grafting method was adopted in the multiplication of transformants without the aid of tissue culture. Among the various parameters evaluated, decapitated plants on pin-pricking and vacuum infiltrated at 250 mmHg for 3 min with the Agrobacterium strain EHA 105 harbouring the binary vector pGA 492 was proved to be efficient in all terms with a transformation efficiency of 62.66%. Transgene integration was evinced by the GUS histochemical analysis, and the GUS positive plants were subjected to grafting. Putatively transformed J. curcas served as "Scion" and the wild type J. curcas plant severed as "Stock". There was no occurrence of graft rejection and the plants were then confirmed by GUS histochemical analysis, polymerase chain reaction (PCR) and Southern hybridization. Genetic stability of the grafted plants was evaluated by using randomly amplified polymorphic DNA (RAPD), marker which showed 100% genetic stability between mother and grafted plants. Thus, an efficient in planta transformation and grafting based multiplication of J. curcas was established.

Design and Fabrication of Anatomical Bioreactor Systems Containing Alginate Scaffolds for Cartilage Tissue Engineering

PubMed Central

Gharravi, Anneh Mohammad; Orazizadeh, Mahmoud; Ansari-Asl, Karim; Banoni, Salem; Izadi, Sina; Hashemitabar, Mahmoud

2012-01-01

The aim of the present study was to develop a tissue-engineering approach through alginate gel molding to mimic cartilage tissue in a three-dimensional culture system. The perfusion biomimetic bioreactor was designed to mimic natural joint. The shear stresses exerting on the bioreactor chamber were calculated by Computational Fluid Dynamic (CFD). Several alginate/bovine chondrocyte constructs were prepared, and were cultured in the bioreactor. Histochemical and immunohistochemical staining methods for the presence of glycosaminoglycan(GAG), overall matrix production and type II collagen protein were performed, respectively. The dynamic mechanical device applied a linear mechanical displacement of 2 mm to 10 mm. The CFD modeling indicated peak velocity and maximum wall shear stress were 1.706×10−3 m/s and 0.02407 dyne/cm 2, respectively. Histochemical and immunohistochemical analysis revealed evidence of cartilage-like tissue with lacunas similar to those of natural cartilage and the production of sulfated GAG of matrix by the chondrons, metachromatic territorial matrix-surrounded cells and accumulation of type II collagen around the cells. The present study indicated that when chondrocytes were seeded in alginate hydrogel and cultured in biomimetic cell culture system, cells survived well and secreted newly synthesized matrix led to improvement of chondrogenesis. PMID:23408660

Altered T-Cell Balance in Lymphoid Organs of a Mouse Model of Colorectal Cancer.

PubMed

Tanner, Scott M; Daft, Joseph G; Hill, Stephanie A; Martin, Colin A; Lorenz, Robin G

2016-12-01

The adenomatous polyposis coli (APC) gene is a known tumor suppressor gene, and mice with mutations in Apc (Apc Min/+ ) spontaneously form multiple intestinal neoplasms. In this model of human colorectal cancer (CRC), it has been reported that CD4 + T-cell-derived interleukin 17 (IL-17) promotes intestinal tumor development, but it is not known if the Apc mutation actually directly alters T-cell function and subsequently tumor immunosurveillance. To investigate the Apc Min/+ mutation on T-cell function, flow cytometric, histochemical, and immunofluorescent studies on both wild-type (Apc +/+ ) and Apc Min/+ mice were performed. We identified decreased levels of interferon gamma (IFN-γ + )IL-17 + double-positive CD4 + cells in the mesenteric lymph nodes and Peyer's patches of Apc Min/+ mice. In addition, altered levels of CD8 + cells, and changes in CD8 + production of IFN-γ and granzyme B were observed. These T-cell alterations did modify tumor immunosurveillance, as the adoptive transfer of splenocytes from Apc Min/+ animals into a chemically induced CRC model resulted in the inability to prevent epithelial dysplasia. These results suggest an altered T-cell balance in Apc Min/+ mice may disrupt intestinal homeostasis, consequently limiting intestinal tumor immunosurveillance. © 2016 The Histochemical Society.

Safety Assessment and Botanical Standardization of an Edible Species from South America.

PubMed

Traesel, Giseli Karenina; Machado, Camila Dias; Tirloni, Cleide Adriane Signor; Menetrier, Jacqueline Vergutz; Dos Reis Lívero, Francislaine Aparecida; Lourenço, Emerson Luiz Botelho; Oesterreich, Silvia Aparecida; Budel, Jane Manfron; Junior, Arquimedes Gasparotto

2017-05-01

Tropaeolum majus L. (Tropaeolaceae), commonly known as nasturtium, is an important edible plant native to the Andean States and widely disseminated throughout South America. Despite the use of this species is quite widespread, there are no minimum quality control standards or data on its genotoxicity. So, the aim of this study was to present a detailed anatomical and histochemical study for Tropaeolum majus and provide genotoxicity parameters of a preparation routinely used in South American countries. First, three different Tropaeolum majus aqueous extracts (TMAEs) at concentrations of 1.5%, 7%, and 15% were prepared according to the popular use. Then, genetic toxicity of TMAE was evaluated on bacterial reverse mutation, genomic lesions, and micronucleus formation in male rats. Furthermore, a detailed anatomical and histochemical study of the leaves and stems of Tropaeolum majus were performed. No revertant colonies were found in any bacterial cultures examined. In the comet assay, TMAE showed no significant DNA damage in all tested doses. Micronucleus assay showed no significant increases in the frequency of inducing micronuclei in any dose examined. Light and electron microscope images of cross-section of leaves and stems from Tropaeolum majus revealed useful diagnostic features. The presented data showed significant safety parameters for the use of TMAE and provided important data for the quality control of this plant species.

Enzymatic cell wall degradation of Chlorella vulgaris and other microalgae for biofuels production.

PubMed

Gerken, Henri G; Donohoe, Bryon; Knoshaug, Eric P

2013-01-01

Cell walls of microalgae consist of a polysaccharide and glycoprotein matrix providing the cells with a formidable defense against its environment. We characterized enzymes that can digest the cell wall and weaken this defense for the purpose of protoplasting or lipid extraction. A growth inhibition screen demonstrated that chitinase, lysozyme, pectinase, sulfatase, β-glucuronidase, and laminarinase had the broadest effect across the various Chlorella strains tested and also inhibited Nannochloropsis and Nannochloris strains. Chlorella is typically most sensitive to chitinases and lysozymes, both enzymes that degrade polymers containing N-acetylglucosamine. Using a fluorescent DNA stain, we developed rapid methodology to quantify changes in permeability in response to enzyme digestion and found that treatment with lysozyme in conjunction with other enzymes has a drastic effect on cell permeability. Transmission electron microscopy of enzymatically treated Chlorella vulgaris indicates that lysozyme degrades the outer surface of the cell wall and removes hair-like fibers protruding from the surface, which differs from the activity of chitinase. This action on the outer surface of the cell causes visible protuberances on the cell surface and presumably leads to the increased settling rate when cells are treated with lysozyme. We demonstrate radical ultrastructural changes to the cell wall in response to treatment with various enzyme combinations which, in some cases, causes a greater than twofold increase in the thickness of the cell wall. The enzymes characterized in this study should prove useful in the engineering and extraction of oils from microalgae.

A study of overproduction and enhanced secretion of enzymes. Quarterly report

DOE Office of Scientific and Technical Information (OSTI.GOV)

Dashek, W.V.

1993-09-01

Wood decay within forests, a significant renewable photosynthetic energy resource, is caused primarily by Basidiomycetous fungi, e.g., white rot fungi. These organisms possess the ability to degrade lignin, cellulose and hemicellulose, the main organic polymers of wood. In the case of the white rot fungi, e.g., Coriolus versicolor, the capacity results from the fungus` ability to elaborate extracellular cellulolytic and ligninolytic enzymes. With regard to the latter, at least one of the enzymes, polyphenol oxidase (PPO) appears within a defined growth medium. This proposal focuses on the over-production and enhanced secretion of PPO, cellulase and lignin peroxidase. There are twomore » major sections to the proposal: (1) overproduction of lignocellulolytic enzymes by genetic engineering methodologies and hyper-production and enhanced secretion of these enzymes by biochemical/electro microscopical techniques and (2) the biochemical/electron microscopical method involves substrate induction and the time-dependent addition of respiration and PPO enzymes.« less

The interaction of the Eco R1 restriction enzyme E.coli with nucleotides

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hollis, Donald F.

1979-11-01

The Eco R1 restriction enzyme can be shown to be inhibited by nucleotides which correspond to any part of its known site of phosphodiesterase activity. A series of di-, tetra-, and hexa-nucleotide fragments were synthesized and their effect on the activity of the enzyme upon superhelical Co1 E1 DNA studied. The inhibition caused by the individual mononucleotides were also studied. In general all the nucleotide fragments showed some form of interaction with the enzyme system. Tetranucleotides were stronger inhibitors than dinucleotides, which in turn were stronger inhibitors than the mononucleotides. Within each category of inhibitors, those containing the phosphodiester bondmore » which is acted upon by the enzyme were the strongest inhibitors. Only those fragments which were consistent with the enzymes site of activity showed competitive inhibition kinetics. Nucleotides which do not fit within the site of phosphodiesterase activity show non-competitive inhibition kinetics.« less

[Acatalasemia and type 2 diabetes mellitus].

PubMed

Góth, László; Nagy, Teréz; Káplár, Miklós

2015-03-08

The catalase enzyme decomposes the toxic concentrations of hydrogen peroxide into oxygen and water. Hydrogen peroxide is a highly reactive small molecule and its excessive concentration may cause significant damages to proteins, deoxyribonucleic acid, ribonucleic acid and lipids. Acatalasemia refers to inherited deficiency of the catalase enzyme. In this review the authors discuss the possible role of the human catalase enzyme, the metabolism of hydrogen peroxide, and the phenomenon of hydrogen peroxide paradox. In addition, they review data obtained from Hungarian acatalasemic patients indicating an increased frequency of type 2 diabetes mellitus, especially in female patients, and an early onset of type 2 diabetes in these patients. There are 10 catalase gene variants which appear to be responsible for decreased blood catalase activity in acatalasemic patients with type 2 diabetes. It is assumed that low levels of blood catalase may cause an increased concentration of hydrogen peroxide which may contribute to the pathogenesis of type 2 diabetes mellitus.

Mercury(II) binds to both of chymotrypsin's histidines, causing inhibition followed by irreversible denaturation/aggregation.

PubMed

Stratton, Amanda; Ericksen, Matthew; Harris, Travis V; Symmonds, Nick; Silverstein, Todd P

2017-02-01

The toxicity of mercury is often attributed to its tight binding to cysteine thiolate anions in vital enzymes. To test our hypothesis that Hg(II) binding to histidine could be a significant factor in mercury's toxic effects, we studied the enzyme chymotrypsin, which lacks free cysteine thiols; we found that chymotrypsin is not only inhibited, but also denatured by Hg(II). We followed the aggregation of denatured enzyme by the increase in visible absorbance due to light scattering. Hg(II)-induced chymotrypsin precipitation increased dramatically above pH 6.5, and free imidazole inhibited this precipitation, implicating histidine-Hg(II) binding in the process of chymotrypsin denaturation/aggregation. Diethylpyrocarbonate (DEPC) blocked chymotrypsin's two histidines (his 40 and his 57 ) quickly and completely, with an IC 50 of 35 ± 6 µM. DEPC at 350 µM reduced the hydrolytic activity of chymotrypsin by 90%, suggesting that low concentrations of DEPC react with his 57 at the active site catalytic triad; furthermore, DEPC below 400 µM enhanced the Hg(II)-induced precipitation of chymotrypsin. We conclude that his 57 reacts readily with DEPC, causing enzyme inhibition and enhancement of Hg(II)-induced aggregation. Above 500 µM, DEPC inhibited Hg(II)-induced precipitation, and [DEPC] >2.5 mM completely protected chymotrypsin against precipitation. This suggests that his 40 reacts less readily with DEPC, and that chymotrypsin denaturation is caused by Hg(II) binding specifically to the his 40 residue. Finally, we show that Hg(II)-histidine binding may trigger hemoglobin aggregation as well. Because of results with these two enzymes, we suggest that metal-histidine binding may be key to understanding all heavy metal-induced protein aggregation. © 2017 The Protein Society.

A New Strategy for Smoking Cessation: Characterization of a Bacterial Enzyme for the Degradation of Nicotine.

PubMed

Xue, Song; Schlosburg, Joel E; Janda, Kim D

2015-08-19

Smoking is the leading cause of preventable diseases; thus, effective smoking cessation aids are crucial for reducing the prevalence of cigarette smoking and smoking-related illnesses. In our current campaign we offer a nicotine-degrading enzyme from Pseudomonas putida, NicA2, a flavin-containing protein. To explore its potential, a kinetic evaluation of the enzyme was conducted, which included determination of K(m), k(cat), buffer/serum half-life, and thermostability. Additionally, the catabolism profile of NicA2 was elucidated to assess the potential toxicity of the nicotine-derived products. In characterizing the enzyme, a favorable biochemical profile of the enzyme was discovered, making NicA2 a prospective therapeutic candidate. This approach provides a new avenue for the field of nicotine addiction therapy.

Substrate specificity and kinetic properties of alpha-galactosidases from Vicia faba.

PubMed

Dey, P M; Pridham, J B

1969-10-01

1. The hydrolysis of a variety of galactosides and other glycosides by alpha-galactosidases I and II of Vicia faba was studied. 2. The effect of temperature on kinetic parameters was also examined. 3. Both enzymes are inhibited by excess of substrate (p-nitrophenyl alpha-d-galactoside); with enzyme I this is competitive and is caused by the galactosyl moiety. 4. Enzyme I is inhibited by oligosaccharides possessing terminal non-reducing galactose residues and to a smaller extent by l-arabinose and d-fucose. 5. The effect of pH on K(m) and V(max.) values suggests that carboxyl and imidazole groups are involved in the catalytic activity of enzyme I. 6. Photo-oxidation experiments with enzyme I also suggest that an imidazole group is present at the active site.

Isolation of a matrix that binds medial Golgi enzymes

PubMed Central

1994-01-01

Rat liver Golgi stacks were extracted with Triton X-100 at neutral pH. After centrifugation the low speed pellet contained two medial-Golgi enzymes, N-acetylglucosaminyltransferase I and mannosidase II, but no enzymes or markers from other parts of the Golgi apparatus. Both were present in the same structures which appeared, by electron microscopy, to be small remnants of cisternal membranes. The enzymes could be removed by treatment with low salt, leaving behind a salt pellet, which we term the matrix. Removal of salt caused specific re-binding of both enzymes to the matrix, with an apparent dissociation constant of 3 nM for mannosidase II. Re-binding was abolished by pretreatment of intact Golgi stacks with proteinase K, suggesting that the matrix was present between the cisternae. PMID:8106542

Following the compositional changes of fresh grape skin cell walls during the fermentation process in the presence and absence of maceration enzymes.

PubMed

Zietsman, Anscha J J; Moore, John P; Fangel, Jonatan U; Willats, William G T; Trygg, Johan; Vivier, Melané A

2015-03-18

Cell wall profiling technologies were used to follow compositional changes that occurred in the skins of grape berries (from two different ripeness levels) during fermentation and enzyme maceration. Multivariate data analysis showed that the fermentation process yielded cell walls enriched in hemicellulose components because pectin was solubilized (and removed) with a reduction as well as exposure of cell wall proteins usually embedded within the cell wall structure. The addition of enzymes caused even more depectination, and the enzymes unravelled the cell walls enabling better access to, and extraction of, all cell wall polymers. Overripe grapes had cell walls that were extensively hydrolyzed and depolymerized, probably by natural grape-tissue-ripening enzymes, and this enhanced the impact that the maceration enzymes had on the cell wall monosaccharide profile. The combination of the techniques that were used is an effective direct measurement of the hydrolysis actions of maceration enzymes on the cell walls of grape berry skin.

Effects of AISI 316L corrosion products in in vitro bone formation.

PubMed

Morais, S; Sousa, J P; Fernandes, M H; Carvalho, G S; de Bruijn, J D; van Blitterswijk, C A

1998-06-01

Rat bone marrow cells were cultured in experimental conditions that favour the proliferation and differentiation of osteoblastic cells (i.e., 2.52 x 10(-4) mol l(-1) ascorbic acid, 10(-2) mol l(-1) beta-glycerophosphate and 10(-8) mol l(-1) dexamethasone) in the absence and in the presence of stainless-steel corrosion products, for a period of 18 days. An AISI 316L stainless-steel slurry (SS) was obtained by electrochemical means and the concentrations of the major metal ions, determined by atomic absorption spectrometry, were 8.78 x 10(-3) mol l(-1) of Fe, 4.31 x 10(-3) mol l(-1) of Cr and 2.56 x 10(-3) mol l(-1) of Ni. Bone marrow cells were exposed to 0.01, 0.1 and 1% of the SS and at the end of the incubation period, control and treated cultures were evaluated by histochemical assays for the identification of the presence of alkaline phosphatase and also calcium and phosphate deposition. Cultures were further observed by scanning electron microscopy. Levels of total and ionised calcium and phosphorus in the culture media collected from control and metal exposed cell cultures were also quantified. Histochemical staining showed that control cultures presented a strong reaction for the presence of alkaline phosphatase and exhibited formation of calcium and phosphates deposits. The presence of 0.01% SS caused no detectable biological effects in these cultures, 0.1% SS impaired osteoblastic behaviour and, 1% SS resulted in cell death. In the absence of bone cells, levels of total and ionised calcium and phosphorus in the control and metal added culture medium were similar throughout the incubation period. A significant decrease in the levels of ionised calcium and phosphorus were observed in the culture medium of control cultures and also in cultures exposed to 0.01% SS after two weeks of incubation, an event related with the formation of mineral calcium phosphate deposits in these cultures. In cultures grown in the presence of 0.1 and 1% SS corrosion products, levels of calcium and phosphorus were similar to those observed in the absence of cells. Results showed that stainless-steel corrosion products above certain concentrations may disturb the normal behaviour of osteoblast-like rat bone marrow cell cultures.

Improved mortality of the Formosan subterranean termite by fungi, when amended with cuticle-degrading enzymes or eicosanoid biosynthesis inhibitors

USDA-ARS?s Scientific Manuscript database

Formosan subterranean termites (FST) were exposed to spores of the fungus Beauveria pseudobassiana (Bpb) strain 8046 to determine virulence of the fungus. Once Bpb was determined to cause mortality of FST it was combined with enzymes capable of degrading the insect cuticle to measure the potential ...