Histochemical Distribution Research Articles

Extracellular silicate uptake and deposition induced by oxidative burst in Saccharina japonica sporophytes (Phaeophyceae)

The process of biogenic silicon (Si) deposition in sporophytes of kelp (Saccharina japonica) was investigated by culturing sporophyte disks in the medium with and without hydrogen peroxide (H2O2). Culture with H2O2 remarkably promoted silicate-Si uptake by the disks. Silicate-Si uptake was completely inhibited by the treatment with potassium bromide (KBr), a competitive inhibitor of iodoperoxidase (IPO), even in the medium with H2O2. In addition, disk culture with phenol red in the medium with H2O2 resulted in the formation of bromophenol blue, indicating the presence of extracellular IPO. However, KBr treatment inhibited the formation of bromophenol blue in the medium with H2O2. These results showed that silicate-Si uptake is activated by extracellular IPO in the presence of radical oxygen species (ROS), such as H2O2. Interestingly, 1 mM salicylic acid also promoted silicate-Si uptake, even in the medium without H2O2. Furthermore, the allocation of silicate-Si to the soluble and cell wall-bound phlorotannin fractions using rhodamine 123 (R123), a tracer of biogenic Si, was estimated. Tracer fluorescence was detected in not only the soluble but also the cell wall-bound phlorotannin fraction. R123 fluorescence was histologically detected in the apoplasts of the epidermal and outer cortical layers of the disk, overlapping the histochemical distribution of IPO activity. These results suggested that silicate-Si uptake is activated by an oxidative burst induced by various stresses, and Si deposition is catalyzed by extracellular IPO in the presence of both ROS and phlorotannin in the extracellular matrix of sporophytes. Based on these findings, the hypothetical mechanisms of the extracellular silicate-Si uptake for defense responses in kelp were discussed.

Histochemical characteristics and distribution of lipofuscin and polyglucosan bodies in the brain of dogs more than 10 years old

The ageing process is accompanied by numerous changes in the brain of dogs, such as accumulation of amyloid, fibrosis of blood vessel walls and meninges, accumulation of lipofuscin, and the presence of polyglucosan bodies (PGBs), satellitosis and neuronophagia. In this study, the presence of lipofuscin and PGBs in various parts of the brain in dogs of different sexes and ages was examined. For this purpose, brain samples were stained using haematoxylin eosin, modified Ziehl Neelsen and Periodic acid Schiff (PAS) methods. Lipofuscin was visualised by Ziehl Neelsen and PAS methods of specific staining on the same brain tissue segments. Lipofuscin had accumulated in 93% of old (more than 10 years old) dog brains, mostly in neurons of the medulla oblongata. The percentage of age-related lipofuscin pigment in other examined brain tissue segments was lower than in the medulla oblongata. There was a small difference in the percentage of lipofuscin-positive individuals between the two staining methods. The presence of PGBs was established by the PAS method for the vast majority (about 93%) of the old dogs (more than 10 years old), while PGBs were not detected in the group of young dogs (up to 5 years old). However, PGBs occurred in all examined segments of the dog?s brain tissues (for each of the tissue types, from 90% to 93% of dogs were positive for PGBs). The results obtained the oldest dogs (15 years old) harboured PGBs both extracellularly and intracellularly, while in other dogs, only extracellular PGBs were seen. Lipofuscin was accumulated mostly in large neurons of olivary nuclei of the medulla oblongata. PGBs were confirmed in all examined segments of the brain tissue of dogs more than 10 years old. This is one of the numerous indications that old dogs could be a very good animal model for studying the normal ageing process or neurodegenerative diseases.

Histochemical Distribution of Three Types of Enzymes and Mucous Cells in the Gill, Mantle and Hepatopancreas of the Ark Shell Scapharca subcrenata

The present study was aimed at evaluating the histochemical distribution of alkaline phosphatase (ALP), nonspecific esterase (NSE), peroxidase (POD) and various types of mucous cells in the gill, mantle and hepatopancreas of the ark shell Scapharca subcrenata. The ALP activity was found in the epithelial cells (EP) of the gill filament and the mantle as well as in the basal lamina of the glandular duct in the hepatopancreas. The staining intensity of ALP in the mantle and hepatopancreas was stronger than that in the gill. The NSE activity was detected in the EP in the gill filament, mantle and hepatopancreas, with the hepatopancreas showing a stronger intensity than the gill. The POD activity was located in the EP of the mantle and glandular duct of the hepatopancreas as well as in the connective tissue of the gill. The mantle showed stronger staining intensity of POD than the hepatopancreas. The Alcian blue (pH 2.5) periodic acid Schiff (AB-PAS) histochemical results revealed four types of mucous cells in the gill, mantle and hepatopancreas. Type III cells (nPAS+AB+) and type IV cells (aPAS+AB+) were observed in the gill, mantle and hepatopancreas. Type I cells (PAS+AB–) were not detected in the hepatopancreas but were distributed ubiquitously throughout the gill filament and mantle. Type II cells (PAS–AB+) were not detected in the gill but were distributed ubiquitously in the digestive ducts of the hepatopancreas.

Histochemical distribution of four types of enzymes and mucous cells in the intestine of koi carp (Cyprinus carpio var. koi).

The main purpose of this study was to investigate the distribution of acid phosphatase (ACP), alkaline phosphatase (ALP), non-specific esterase (NSE), peroxidase (POD), and mucous cells in the intestine of the koi carp Cyprinus carpio var. koi. ACP activity was located in the striated border, enterocytes, and lamina propria of the anterior and middle intestines. The ACP activity in the anterior intestine was higher than that in the middle and posterior intestines. ALP existed in the striated border of enterocytes and lamina propria, serosa, muscular layer, and the junction between muscular layer and submucosa layer of the intestine. The ALP activity in the anterior intestine was higher than that in the middle and posterior intestines. NSE activity was localized in the cytoplasm of enterocytes in the whole intestine, and the middle intestine showed the lower NSE activity than the anterior and posterior intestines. POD activity was localized in the blood cells of the lamina propria and cytoplasm of enterocytes in all intestinal segments. The POD activity among the anterior, middle, and posterior intestines was non-significantly different. Alcian blue periodic acid-Schiff histochemical results revealed three types of mucous cells in the intestine. The total number of mucous cells and percentage of type I cells among the anterior, middle, and posterior intestines were non-significantly different. The percentage of the type II cells was the highest in the posterior intestine, while the lowest in the anterior intestine. The percentage of the type III cells was the highest in the anterior intestine, while the lowest in the posterior intestine.

Biochemical and ultrastructural changes in Raillietina echinobothrida in vitro exposed to extract of Lysimachia ramosa.

Lysimachia ramosa (Primulaceae) is a traditionally used medicinal plant, leaves extract of which is being widely used by the Jaintia tribes of Meghalaya, India for controlling helminthiasis. Preliminary investigation carried out on helminth parasites revealed that the crude extract of the plant causes deformity in the surface topography leading to death of the parasites. Therefore, the present study was conducted to identify the specific fraction of the crude leaf extract of the plant responsible for cestocidal efficacy, through biochemical and ultrastructural studies in Raillietina echinobothrida exposed to crude extract and its different fractions namely hexane, chloroform, ethyl acetate and n-butanol. A dose dependent efficacy, with highest rate of mortality among n-butanol exposed parasites was recorded. The treated parasites exhibited complete erosion of microtriches from the tegument, disintegration of muscle bundles, cellular organelles, plasma membrane, nuclear membrane, nucleolus and vacuolization of mitochondria was also observed. Observations on histochemical distribution of some important tegumental enzymes like adenosine triphosphatase (ATPase), alkaline phosphatase (AlkPase), acid phosphatase (AcPase) and 5'Nucleotidase (5'-Nu) revealed a marked diminished stain intensity in the tegument of R. echinobothrida exposed to the crude extract and n-butanol fraction of the crude extract compared to the control. Highest reduction (77.93%) in the activity of ATPase was observed when the parasites exposed to 6mg n-butanol fraction/ml of PBS. The results suggest that these enzymes act as target for anthelmintic stress caused by the phytochemicals present in the plant.

Histochemical Distribution of Acid phosphatase in the tissues of Indian Major carp, Labeorohita (ham)

Acid phosphatases (AcPs) are known to provide phosphate to tissues that have high energy requirements, especially during development, growth and maturation. AcPs enzyme shows variations in quantity in reproductive phases in gonads, kidney and liver. AcPs is highest in both the sexes in postspawning phase and lowest in preparatory phase. In male, acid phosphatase activity is maximum in all three tissues i.e gonads, kidney and liver in postspawning phase and lowest in the preparatory phase as in female. AcPs staining was either granular or in diffuse form in brown deposits. This study is aimed on the estimation of activity of acid phosphatase in kidney, liver and gonads tissues of Labeorohita

Evaluation of PACE4 isoforms as biomarkers in thyroid cancer

BackgroundTo date, no single molecular marker has been demonstrated as clinically useful in differentiating malignant from benign thyroid nodules when a fine needle aspiration falls in the “unknown significance” categories of the Bethesda Classification. PACE4, a member of the proprotein convertase family of enzymes, has been shown to play a major role in the pathogenesis of prostate cancer, through the formation of an oncogenic isoform named PACE4-altCT. PACE4 isoforms have also been suggested to play a role in other cancers, including thyroid cancer, but have never been investigated in a detailed manner. Our objective is to compare the histochemical distribution of the two major PACE4 isoforms in benign and malignant thyroid nodules, in order to determine their potential usefulness as discriminatory biomarkers.MethodsThyroid tissues of patients who underwent thyroidectomy were classified according to final pathology. Corresponding tissue sections were immunostained, using two previously validated antibodies raised against the C-terminal end of the two PACE4 isoforms, namely the full-length PACE4 protein (PACE4-FL) and its alternative isoform (PACE4-altCT). Nodules were compared with adjacent normal parenchyma and immunostaining was rated as “low” or “high” by a head and neck pathologist.ResultsNon-lesional thyroid parenchyma did not express PACE4-FL (p = 0.002). As a group, malignant (n = 17) nodules expressed PACE4-FL significantly more than benign (n = 24) nodules (percentage of high immunostaining: 52.9% vs 4.2%; p = 0.001). Reciprocally, there was a statistically lower expression of PACE4-altCT in malignant nodules than in adjacent non-lesional parenchyma (p = 0.014). The specificity of a high PACE4-FL immunostaining in determining malignancy was 95.8% (95% CI, 78.9% to 99.9%).ConclusionThis study supports the previously described relationship between PACE4-FL and PACE4-altCT through alternative splicing. It also suggests that PACE4-FL is a promising biomarker for thyroid malignancy. Its high specific expression for malignancy could make it an interesting “rule in” test for thyroid cancer. Further prospective, quantitative studies are currently being designed to address how measurements of PACE4 isoforms could be used in a clinical setting.Trial registrationThis study does not report the results of a health care intervention on human participants. It was nonetheless registered on ClinicalTrials.gov under reference number NCT03160482.

Histochemical distribution of four types of enzymes and mucous cells in the gastrointestinal tract of reared half-smooth tongue sole Cynoglossus semilaevis.

The histochemical distribution of acid phosphatase (ACP), alkaline phosphatase (ALP), non-specific esterase (NSE), peroxidase (POD) and mucous-cell types was evaluated in the gastrointestinal tract of the half-smooth tongue sole Cynoglossus semilaevis. The enzymes were detected in the entire stretch of the gastrointestinal tract. ACP activity was found in the supranuclear region of enterocytes and the lamina propria of the intestine, as well as the cytoplasm of epithelial cells of the stomach. The staining intensity of ACP in the anterior and posterior intestines was stronger than in the stomach. ALP activity was detected in the striated border of enterocytes and muscularis of the whole intestine, lamina propria and supranuclear cytoplasm of the enterocytes in the anterior intestine, as well as in the blood vessels of the stomach. The staining intensity for ALP in the anterior intestine was stronger than in the posterior segment and the latter was stronger than in the stomach. NSE activity was detected in the cytoplasm of the epithelial cells in the entire gastrointestinal tract, with the anterior intestine showing stronger intensity than the stomach. POD activity was located in the blood cells of the lamina propria of the gastrointestinal tract and the levels in the stomach were similar to the anterior and posterior intestines. Alcian blue (pH2·5) periodic acid Schiff (AB-PAS) histochemical results revealed three types of mucous cells in the gastrointestinal tract. Type I cells (PAS+AB-) were observed among the gastric mucosa columnar cells in the stomach and enterocytes in the basal region of the villi and in the middle and top regions of the intestinal villi. Type II cells (PAS-AB+) and type III cells (PAS+AB+) were not detected in the stomach but were distributed ubiquitously among enterocytes in the middle and top regions of the intestinal villi.

Histological and Histochemical Study of the Hepatopancreas of Two Estuarine Crab Species, Cyrtograpsus angulatus and Neohelice granulata (Grapsoidea, Varunidae): Influence of Environmental Salinity.

The aim of this study was to analyze the histology and the histochemical distribution of glycoproteins (GPs) and lipids of the hepatopancreas of Cyrtograpsus angulatus and Neohelice granulata acclimated to salinities of 10 psu (hyperregulation) and 35 psu (osmoconformation). Sections of the hepatopancreas of adult male crabs were treated with hematoxylin-eosin; Masson trichrome; Alcian Blue (pHs 2.8, 1.0, 0.5); Toluidine Blue (pHs 5.6, 4.2); periodic acid Schiff; Sudan Black and Red. At salinity 35 psu, the hepatopancreas of both species exhibited typical histological features, whereas at salinity 10 psu, detachment of the basal lamina, desquamated epithelium, disrupted brush border, loss of intercellular cohesion, hypertrophied tubular lumen, and hemolymph infiltration between cells were observed in some zones. Resorptive cells (R-cells) and vacuoles of blister-like cells (B-cells) of both species show a higher glycogen content at 35 psu than at 10 psu. At lower salinities, the cytoplasm of the different cell types evidence higher contents of carboxylated GPs in N. granulata and of su If at ed GPs in C. angulatus. At both salinities, and at the two pHs in N. granulata and at pH 5.6 in C. angulatus, the brush border, the vacuoles of B-cells and the peritrophic membrane show metachromasia. R-cell vacuoles and the cytoplasm of all cell types--except for the E-cells--at all salinities in both species show abundant lipid droplets. The results of the present study contribute significant data to the histophysiology of crustacean decapods, favoring the comprehension of the complex adjustment mechanisms facing saline stress in euryhaline crabs.

塩化ランタン（La）投与ラット脳組織におけるLaの分布に関する組織化学的研究

塩化ランタン（La）投与ラット脳組織におけるLaの分布に関する組織化学的研究

Effects of Acacia oxyphylla and Securinega virosa on functional characteristics of Raillietina echinobothrida (Phylum: Platyhelminthes; Class: Cestoidea), a poultry cestode parasite

The histochemical distribution and quantification of alkaline phosphatase, acid phosphatase, and adenosine triphosphatase have been investigated in the adult cyclophyllidean cestode, Raillietina echinobothrida after treatment with crude ethanol extract of traditionally used medicinal plants Acacia oxyphylla and Securinega virosa. Estimation of free amino acid pool and vital trace elements like calcium and magnesium were also performed to study the effect of the plant extracts on the parasite. A decrease in the levels of phosphatases and trace elements was noticed in the treated parasites when compared to control groups. At the same time, a lot of disparity was noted in the free amino acid pool as the levels of many amino acids were seen to increase or decrease in relation to the levels observed in control parasites. Role of phyto-products in the changed physiology of cestode and their possible chemotherapeutic targets are discussed.

Histochemical distribution of sialic acids and antimicrobial substances in porcine carpal glands

The localization of sialic acids and antimicrobial products (lysozyme, IgA, lactoferrin, β-defensin 2) as well as Rab3D in the carpal glands of pig was studied by sialoglycoconjugate histochemistry and immunohistochemistry. The secretory epithelium of the carpal glands consisted of dark and clear cells. The dark cells of these glands exhibited high levels of sialoglycoconjugates, including O-acetylated sialic acids, whereas the localization of sialic acids linked to α2-3Gal1-4GlcNAc was confined to a subpopulation of the dark cells. Furthermore, all antimicrobial substances and Rab3D were mainly detectable in a subpopulation of the dark cells. The results obtained are discussed with regard to the functional significance of these glands. Our findings suggested that Rab3D is involved in the secretory regulation of sialoglycoconjugates and antimicrobial substances. These secretory products may create a defensive barrier against microbial invasion and play an essential role in the preservation of skin integrity.

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

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.

Hyaluronan within fascia in the etiology of myofascial pain

The layers of loose connective tissue within deep fasciae were studied with particular emphasis on the histochemical distribution of hyaluronan (HA). Samples of deep fascia together with the underlying muscles were taken from neck, abdomen and thigh from three fresh non-embalmed cadavers. Samples were stained with hematoxylin-eosin, Azan-Mallory, Alcian blue and a biotinylated HA-binding protein specific for HA. An ultrasound study was also performed on 22 voluntary subjects to analyze the thickness of these deep fasciae and their sublayers. The deep fascia presented a layer of HA between fascia and the muscle and within the loose connective tissue that divided different fibrous sublayers of the deep fascia. A layer of fibroblast-like cells that stained prominently with Alcian blue stain was observed. It was postulated that these are cells specialized for the biosynthesis of the HA-rich matrix. These cells we have termed "fasciacytes", and may represent a new class of cells not previously recognized. The ultrasound study highlighted a mean thickness of 1.88 mm of the fascia lata, 1.68 mm of the rectus sheath, and 1.73 mm of the sternocleidomastoid fascia. The HA within the deep fascia facilitates the free sliding of two adjacent fibrous fascial layers, thus promoting the normal function associated with the deep fascia. If the HA assumes a more packed conformation, or more generally, if the loose connective tissue inside the fascia alters its density, the behavior of the entire deep fascia and the underlying muscle would be compromised. This, we predict, may be the basis of the common phenomenon known as "myofascial pain."

Synthesis and Preliminary Evaluation of Piperidinyl and Pyrrolidinyl Iodobenzoates as Imaging Agents for Butyrylcholinesterase

The purpose of this study is to synthesize and evaluate specific agents for molecular imaging of butyrylcholinesterase (BuChE), known to be associated with neuritic plaques and neurofibrillary tangles in Alzheimer's disease (AD). In this study, these agents were tested in a normal rat model. The distribution of radiolabel was compared with known BuChE histochemical distribution in the rat brain. Iodobenzoate esters were synthesized and tested, through spectrophotometric analysis, as specific substrates for BuChE. These compounds were converted to the corresponding (123)I esters from tributyltin intermediates and purified for studies in the rat model. Whole body dynamic scintigraphic images were obtained for biodistribution studies. Autoradiograms of brain sections were obtained and compared to histochemical distribution of the enzyme in this model system. The three iodobenzoate esters studied were specific substrates for BuChE. Whole body biodistribution studies with (123)I-labeled compounds showed rapid disappearance from the body while radioactivity was retained in the head region. Brain section autoradiography of animals injected with these labeled compounds indicated that most areas known to contain BuChE corresponded to areas of radioactivity accumulation. BuChE-specific radiolabeled iodobenzoates enter the brain and, in general, label areas known to exhibit BuChE activity in histochemical studies. Such molecules may represent a new direction for the development of agents for the molecular imaging of BuChE in the living brain, especially in regions where BuChE-containing neuropathological structures appear in AD.

Histochemical Distribution of Lipase and Acid Phosphatase in the Intestinal Tract of the Snow Trout, Schizothorax curvifrons Heckel.

Histochemical Distribution of Lipase and Acid Phosphatase in the Intestinal Tract of the Snow Trout, Schizothorax curvifrons Heckel.

Studies on the Histochemistry of the Proventriculus and Gizzard of Post-Hatch Guinea Fowl (Numida meleagris)

The proventricular surface epithelial cells were positive for both acid and neutral mucins and stained intensive for acid phosphatase and adenosine triphosphatase. The proprial gland cells revealed histochemical distribution and activity of various enzymes and lipids similar to that of the submucosal gland cells. However, the ductular cells of the submucosal glands showed comparatively stronger reaction for acid phosphatase, alkaline phosphatase, adenosine triphosphatase, lipase and carbonic anhydrase except for succinic dehydrogenase. In gizzard, the koilin was positive for both acid and neutral mucin. The epithelial cells of the proprial glands stained strongly for adenosine triphosphatase and moderately for acid and alkaline phosphatases. However, their response for succinic dehydrogenase and lipase was weak. The smooth muscle cells of the proventriculus and gizzard showed a moderate to strong reaction for adenosine triphosphatase.

Histochemical characterization, distribution and morphometric analysis of NADPH diaphorase neurons in the spinal cord of the agouti

We evaluated the neuropil distribution of the enzymes NADPH diaphorase (NADPH-d) and cytochrome oxidase (CO) in the spinal cord of the agouti, a medium-sized diurnal rodent, together with the distribution pattern and morphometrical characteristics of NADPH-d reactive neurons across different spinal segments. Neuropil labeling pattern was remarkably similar for both enzymes in coronal sections: reactivity was higher in regions involved with pain processing. We found two distinct types of NADPH-d reactive neurons in the agouti's spinal cord: type I neurons had large, heavily stained cell bodies while type II neurons displayed relatively small and poorly stained somata. We concentrated our analysis on type I neurons. These were found mainly in the dorsal horn and around the central canal of every spinal segment, with a few scattered neurons located in the ventral horn of both cervical and lumbar regions. Overall, type I neurons were more numerous in the cervical region. Type I neurons were also found in the white matter, particularly in the ventral funiculum. Morphometrical analysis revealed that type I neurons located in the cervical region have dendritic trees that are more complex than those located in both lumbar and thoracic regions. In addition, NADPH-d cells located in the ventral horn had a larger cell body, especially in lumbar segments. The resulting pattern of cell body and neuropil distribution is in accordance with proposed schemes of segregation of function in the mammalian spinal cord.

Autometallographic tracing of Hg–S quantum dots in frogs exposed to inorganic mercury

The histochemical distribution of mercury in the kidneys and gut of frogs (Rana ridibunda) exposed to inorganic mercury was analyzed with autometallography (AMG). It was found that most mercury in the kidneys accumulated in the proximal convoluted tubules as Hg-S nanocrystal, while control animals were totally void of AMG grains. In the gut the highest concentration of mercury was observed in the large intestine. The AMG grains were primarily located in the apical part of the absorptive cells, although rather high concentrations of silver enhanced mercury quantum dots were also detected in a special cell type of gut epithelium and the glycocalyx. A certain amount of AMG grains were detected in the lumen of the gut. We hypothesize that this pool of quantum dots results from sloughed off epithelial cells and macrophages. Such still intact cells and red blood cells containing AMG grains were also found in the lumen of the gut.

Histochemical distribution of digestive enzymes in intestine of goldline, Sarpa salpa L. 1758

Histochemical localization of non-specific esterase, alkaline and acid phosphatase in the intestine of free-living goldline (Sarpa salpa L. 1758) was investigated. Fish were caught in the vicinity of the town of Zadar(Adriatic Sea Croatia), and samples of three parts of the intestine proper (anterior, middle and posterior) as well as the rectum were used for presentation of non-specific esterases, alkaline phosphatase and acid phosphatase. Non-specific esterase activity was found in the cytoplasm and brush border of enterocytes in all investigated intestinal segments and the rectum. The activity was stronger in the middle and posterior part of intestine but weaker in the anterior segment of the intestine as well as in the rectum. Intestinal alkaline phosphatase was detected in brush border and supranuclear cytoplasm of enterocytes of all investigated intestinal segments. Enzymatic activity gradually decreased in a posterior direction. Acid phosphatase activity was observed as fine granular reaction product in supranuclear region of enterocytes and was almost equal in all investigated intestinal segments as well as in the rectum. The possible role of enzymes in intracellular digestion and transport is discussed.