Studies on vertebrate kidney. V. PAS-positive lipids in kidneys of fishes from different habitats.

Five fish species living in different habitats, i.e. fresh water, estuarine and marine, were studied for the distribution of PAS-positive materia in various regions of the kidney, 10 minutes' oxidation with 0-5 per cent. HIO4 being employed prior to treatment with Schiff's reagent. PAS-positive material was detected in different sites of the kidney, i.e. brush border of proximal tubules, proximal tubule cells' cytoplasm, distal tubule cells' cytoplasm, glomerulus, basal cell border of proximal tubules and the interstitial cells. Of these sites, the brush border of the proximal tubule of Scoliodon sorrakowah showed the presence of PAS-positive lipids. Elsewhere the PAS-positive reaction was due to carbohydrates. Free aldehyde groups were absent. In Tilapia mossambica and Labeo rohita, PAS staining was enhanced after chloroform-methanol extraction, particularly in the brush border of the proximal tubule. The significance of these findings is discussed.     

Endogenous cytochrome c and cytochrome oxidase in rat and mouse kidney: light microscopic histochemical study.

Activity of cytochrome c oxidase and the level of endogenous cytochrome c were investigated light microscopically in adult rat and mouse kidney by incubating unfixed frozen sections with diaminobenzidine (DAB) in the absence or presence of exogenous cytochrome c. The results suggest that DAB staining intensity mainly reflects the local density of mitochondria and only occasionally visualizes the differences in cytochrome oxidase activity and/or endogenous cytochrome c content. Most intense reaction was observed in proximal and distal tubules both in rat and mouse. Finer differentiation of reactivity in particular nephron segments and interspecies differences between rat and mouse kidney are also described.     

中华鳖肾脏的组织化学特性研究

利用光镜组织化学反应对中华鳖肾单位的结构和组织化学特性进行了详细的观察和分析。结果表明,中华鳖肾脏为分叶形的实质器官,肾小叶由被膜和实质组成,实质无髓质和皮质之分,但可以区分为外侧区和内侧区。外侧区嗜酸性,主要分布有近端小管和集合管。内侧区呈弱嗜酸性,肾小体、颈段、中间段和远端小管主要分布在内侧区。肾小球PAS反应呈阳性,但其琥珀酸脱氢酶(SDH)弱阳性,碱性磷酸酶(ALPase)、Na+/K+-ATPase和阿利新兰(AB)反应为阴性。足细胞酸性磷酸酶(ACPase)反应呈阳性。近端小管刷状缘嗜伊红,PAS反应以及ALPase、ACPase和Na+/K+-ATPase酶反应呈阳性,而SDH弱阳性。中间段、远端小管、集合管弱嗜酸性,SDH阳性。中间段Na+/K+-ATPase弱阳性。远端小管细胞侧面呈PAS阳性,腔面显示AB阳性。集合管胞质含有许多ACPase阳性颗粒,腔面呈PAS强阳性,AB阳性。甲苯胺兰(TB)染色可见集合管腔面有阳性颗粒,肾小管上皮含有亮、暗两种细胞。上述组化反应和分布结果表明,鳖的肾小管细胞类型较多,近端小管在原尿的重吸收中起主要作用,远端小管和集合管具有分泌黏液作用。中华鳖肾单位的结构与组化特性不仅与哺乳类和鸟类有一定差异,也与其他爬行动物不完全相同。       

A note on the sensitivity of the histochemical plasmal reaction

Summary Comparison of the results from both histochemical and biochemical determinations of the plasmalogen concentration in several tissues and organs shows that the two methods are in general agreement. A comparison of the two methods also indicates that the lowest concentration of plasmalogen detectable with the histochemical procedure is about 0.7 moles per gram when 10 sections are employed.This investigation was supported by USPHS Grant 5TI-GM-407.A portion of this paper was prepared from data in a dissertation submitted to the Faculty of the Graduate School of the State University of New York at Buffalo, in partial fulfillment of the requirements for the degree of Doctor of Philosophy.     

Neurosecretion in the distal stump of the sectioned hypothalamic-hypophyseal tract in the toad

Summary After section of the hypothalamic-hypophyseal neurosecretory pathway of the toad accumulation of neurosecretory material both proximally and distally to the plane of section was observed. When, in other animals, a second section close to the neurohypophysis was performed, neurosecretory material was absent in the distal stump resulting from the first section. The pile-up of neurosecretion in the distal stump of the unisectioned tract is attributable to a backward flow from the neurohypophysis to the lesion rather than to local synthesis of neurosecretion.The present investigation was carried out as a consequence of a discussion by the senior author with ProfessorWilliam Etkin, Department of Anatomy, Albert Einstein College of Medicine, New York (USA).     

Biomorphosis of the human fallopian tube mucosa; a contribution to the problem of morphology and aging of the uterine tube. II. Histochemical findings

The present histochemical findings of investigations on the biomorphosis of the human tubal mucosa indicate that mucosubstances and glycogen localized in the epithelium exhibit age-dependent changes with regard to occurrence and localization. In the embryo-fetal time PAS-positive, diastase-resistant substances are localized in the epithelium, at first basally, and later perinuclearly. In the neonatal phase the distal tubal epithelium has only a weak PAS reaction, and the proximal epithelium has a detectable supranuclear activity. In the end of the 1st decade of the life the epithelium possesses a periodate reactive diastase-sensitive material densely deposited in the preampullar and ampullar parts of the uterine tube, preferably. Afterwards PAS-positive diastase-sensitive and diastase-resistant substances, respectively, are regularly present, in which in the fertile age of the women a regular pattern of the PAS activity can be demonstrated. In the period of the regressive age it is possible to establish a increasing disturbance of the usual cellular picture of the tubal epithelium. In connection with the structural changes a increase of histochemical different reacting cell groups is evident. As a result, a dissociated cellular picture has developed. Epithelial glycoproteins and glycogen can be detected in the mucosa up to the phase of the senium.     

Immunohistochemical identification of tubular segments in percutaneous renal biopsies

Summary To identify the renal cortical tubular segments involved in tubulo-interstitial disease in formalin-fixed, paraffin-embedded percutaneous kidney biopsies, we developed multiple immunolabeling protocols using segment-specific tubular markers. The present study of biopsies from patients with minimal change or thin basement membrane nephropathy provides a baseline for interpretation of histopathology. Proximal tubules were stained either by the PAS reaction or by the biotinylated Phaseolus vulgaris erythroagglutinin (PHA-E)-streptavidin-gold-silver system (brush borders black). The anti-Tamm-Horsfall (THP) antibody-immunoperoxidase (aminoethylcarbazole, AEC-IPO), and anti-epidermal cytokeratins (ECK) antibodies-immunoalkaline-Fast Blue BB methods marked the distal straight tubules and the cortical collecting system red-brown and blue, respectively. When these immunolabelings were combined, the coapplication of AEC-PO-labeled peanut agglutinin (PNA) or anti-epithelial membrane antigen antibody-AEC-IPO technique (both are markers for distal nephron) visualized the apical membranes of distal convoluted tubules. In the protocol PHA-E + PNA + THP + ECK, the tubular basement membranes were outlined by the anti-laminin antibody-AEC-IPO staining, carried out simultaneously. The protocol PNA + THP + ECK + PAS was found to be a quite appropriate multiple immunolabeling method for the tubules, and is recommended for use as a tool in the study of tubulo-interstitial diseases.Abbreviations PAS periodic acid-Schiff reaction - PHAE Phaseolus vulgaris erythroagglutinin - PNA Peanut agglutinin - EMA epithelial membrane antigen - THP Tamm-Horsfall glycoprotein - ECK epidermal cytokeratins - PO peroxidase - Biot-PHA-E biotinylated PHA-E - APAAP complexes of alkaline phosphatase and mouse monoclonal anti-alkaline phosphatase - SWARI swine anti-rabbit immunoglobulins - FCS fetal calf serum - TBS Tris-buffered saline - AEC aminoethylcarbazole - DAB diaminobenzidine - FBBB Fast Blue BB - IA immunoalkaline - GL glomerulus - PT proximal tubule - DST distal straight tubule - DCT distal convoluted tubule - CCS cortical collecting system - CT connecting tubule - CD collecting duct     

Spermatophores of the salamander Ambystoma texanum

Living spermatozoa were observed in freshly deposited spermatophores and in fluid from vasa deferentia. In the distal, but not proximal, vas deferens spermatozoa moved together in whorls with heads and tails in alignment. Around the entire periphery of the spermatophore cap, similar slowly undulating groups of spermatozoa had their heads aligned and directed outward. Over time, some individual spermatozoa left the cap of the spermatophore and moved into the surrounding water (cap deterioration). Microscopical observations were made on spermatophore squashes and paraffin sections of spermatophores and vasa deferentia. Spermatozoa around the periphery of the cap were underlain by a PAS-positive membrane-like material. Cytoplasmic droplets, which were attached to spermatozoan necks in the vas deferens, were accumulated in the center of the spermatophore cap deep to the PAS-positive membrane. Spermatophore stalks were strongly PAS and Alcian blue positive and showed positive reaction for RNA. Tests for lipids and proteins were negative in the whole spermatophore. Electron microscopic observations showed the stalk of the spermatophore to be composed of rounded ‘balls’ of fibrous material. At the juncture of the stalk and cap a less dense fibrous material impacted the stalk enclosing many sperm tails and some heads and, although no attachment devices were visualized, the sperm were closely apposed to this material as was the spermatophore stalk. This finely filamentous material encircled the cap and was more prominent in some regions than others. The PAS-positive material detected with the light microscope was also observed with the electron microscope. It was circumferentially oriented and was composed of 200 Å packed filamentous densities. Sperm heads and tails were found lying external to the membrane, whereas only tails and cytoplasmic droplets occupied the core of the spermatophore. Cytoplasmic droplets were usually free of the sperm tail and contained membranous sacs and two types of nuage material.     

Immunohistochemical identification of tubular segments in percutaneous renal biopsies.

To identify the renal cortical tubular segments involved in tubulo-interstitial disease in formalin-fixed, paraffin-embedded percutaneous kidney biopsies, we developed multiple immunolabeling protocols using segment-specific tubular markers. The present study of biopsies from patients with minimal change or thin basement membrane nephropathy provides a baseline for interpretation of histopathology. Proximal tubules were stained either by the PAS reaction or by the biotinylated Phaseolus vulgaris erythroagglutinin (PHA-E)-streptavidin-gold-silver system (brush borders black). The anti-Tamm-Horsfall (THP) antibody-immunoperoxidase (aminoethylcarbazole, AEC-IPO), and anti-epidermal cytokeratins (ECK) antibodies-immunoalkaline-Fast Blue BB methods marked the distal straight tubules and the cortical collecting system red-brown and blue, respectively. When these immunolabelings were combined, the coapplication of AEC-PO-labeled peanut agglutinin (PNA) or anti-epithelial membrane antigen antibody-AEC-IPO technique (both are markers for distal nephron) visualized the apical membranes of distal convoluted tubules. In the protocol PHA-E + PNA + THP + ECK, the tubular basement membranes were outlined by the anti-laminin antibody-AEC-IPO staining, carried out simultaneously. The protocol PNA + THP + ECK + PAS was found to be quite appropriate multiple immunolabeling method for the tubules, and is recommended for use as a tool in the study of tubulo-interstitial diseases.     

Bronchioalveolar carcinoma with renal and hepatic metastases in a degu (Octodon degus)

A primary bronchioloalveolar carcinoma with renal and hepatic metastases was diagnosed in a mature male degu (Octodon degus) that was found dead in a zoological exhibit (Buffalo Zoological Gardens, Buffalo, New York, USA). Grossly, a discrete 0.5 cm diameter nodule was seen in the lung. Smaller, but similar nodules were present scattered in the liver and kidneys. Histologically, nests and sheets of an infiltrating population of cuboidal to low columnar neoplastic epithelial cells partially effaced pulmonary architecture. Vascular invasion was evident. Similar nests and sheets of neoplastic cells were present within the renal cortex and medulla, and a small nest was present within the hepatic parenchyma. This is the first record of this neoplasm in a degu.     

La réaction de la chitine à l'acide-periodique-schiff

Résumé La réaction positive de la chitine à l'APS nécessite l'oxydation des radicaux hydroxy-acétyl-amino des unités d'N-acétylglucosamine par l'acide periodique. L'étude de l'oxydation périodique de dérivés N-acylés d'acides hydroxyaminés et de différents sucres montre que ce radical n'est pas attaqué dans les conditions de l'APS. La chitine sous forme de poly-N-acétylglucosamine doit donc être considérée comme APS-négative. Le cas des structures APS+ contenant de la chitine est discuté.

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The PAS-positive reaction of chitin

Summary The PAS-positive reaction of chitin requires periodic oxidation of hydroxyacetyl-amino groups of N-acetyl-glucosamine units. Periodic oxidation of N-acyl-hydroxyamino acids and of different sugars shows that hydroxy-acetyl-amino groups are not clived under PAS conditions. Thus, chitin, considered as poly-N-acetylglucosamine, is PAS-negative.The PAS reaction of different chitinous structures (especially insect cuticles) is studied.When it is not linked to protein or only weakly, chitin is PAS-positive. Acetylation and deacetylation tests and oxidation with sodium bismuthate suggest that the positive reaction is due to hydroxy-amino groups of glucosamine residues included in the poly-N-acetylglucosamine chain.In stabilized insect cuticles, chitin is linked to form a glycoprotein and reacting groups are masked: PAS reaction is negative. Generally, hot dilute alkali is used to remove the protein fraction. But the Tables 1 and 2 show that chitin undergoes deacetylation and becomes PAS-positive if purification by alcali is too long or concentration too high.It appears that KOH N 24 h at 100° C does not cause deacetylation, and removes enough protein to unmask reacting groups. After such treatment, hard sclerotized cuticles remain PAS-negative, and endocuticles become PAS-positive. The groups responsible for PAS reaction are identified as hydroxy-amino groups by complementary tests.These results confirm that chitin may contain glucosamine, and suggest that during the stabilization process the hydroxy-amino groups are blocked by strong protein or water linkage, or simply acetylated. These two hypothesis are discussed.

     

Cellular patterns of post-mortem glycogenolysis in longissimus dorsi muscles of pigs at different live weights

Synopsis Serial frozen sections of longissimus dorsi muscles from twelve Canadian Yorkshire breed pigs at different live weights (13–86kg) were stained for glycogen by the periodic acid-Schiff (PAS) reaction and reacted for NAD tetrazolium reductase to determine the mitochondrial content of muscle fibres. Sections from muscles taken immediatelypost mortem and at 5 hrpost mortem were compared and the patterns of glycogenolysis in high, intermediate and low mitochondrial content fibres were assessed on the basis of the percentage of initially PAS-positive fibres which became PAS-negative 5 hrpost mortem. This form of assessment was necessary because not all fibres were PAS-positive immediatelypost mortem. In smaller pigs, only a few muscle fibres depleted their stainable glycogen by 5 hr and most of these fibres had a high mitochondrial content. In larger pigs, most or all initially PAS-positive fibres became PAS-negative by 5 hrpost mortem. Fibres with a low mitochondrial content accounted for most of the glycogenolysis detected histochemically in larger animals. The overall percentage of PAS-positive fibres was related to glycogen concentration (mg/g),r=0.52,P<0.025, when samples with all PAS-positive fibres were excluded.     

小熊猫肾脏和输尿管的组织学研究

小熊猫的肾脏呈蚕豆形,表面光滑不分叶,只有1个肾锥体和1个肾盏,无肾盂。肾脏皮质内可见皮质迷路和髓放线。皮质迷路内有近曲小管、远曲小管和肾小体等结构。髓放线内有近端小管直部和远端小管直部。髓质可分为外髓和内髓两个区域。外髓有较多的集合管断面,少量的远端小管直部和细段,较多的直小血管束。内髓部位有大量的细段和乳头管。各种泌尿小管之间有少量的疏松结缔组织构成的间质,间质内有丰富的毛细血管。输尿管横切面呈圆形或卵圆形,管腔呈不规则的裂隙状。管壁由粘膜、肌肉层和外膜组成。并与大熊猫肾脏和输尿管的组织结构作了比较研究。     

An improved periodic acid-thiosemicarbazide-osmium technique to reveal glycoconjugates at the molecular level in situ

The periodic acid-thiocarbohydrazide or thiosemicarbazide-OsO4 method (Seligman AM, Hanker JS, Wasserkrug H, Katzoff L: J Histochem Cytochem 13:629, 1965) has been modified in order to obtain a periodic acid-Schiff (PAS)-like reaction for electron microscopy capable of visualizing structures at the molecular level in situ. Thiocarbohydrazide (TCH) and thiosemicarbazide (TSC) have been used dissolved in distilled water and bubbled with SO2. Treatment of previously oxidized thin sections with TCH (SO2) or TSC (SO2), followed by osmification, resulted in selective and very good staining of all the PAS-positive structures examined: glycogen, intestinal mucopolysaccharides, plasma membrane glycoproteins, basement membranes, Golgi apparatus, and collagen. The staining reaction was highly specific when TSC was used on thin sections from paraformaldehyde-fixed samples. The non-particulate end-reaction product made possible visualization of a periodic distribution of sugar residues in the 64-nm unit of collagen and the structural organization of the PAS-positive glycoconjugate components in the glomerular basement membrane.     

Signet ring cells in gastric carcinomas are derived from neuroendocrine cells.

Adenocarcinomas are malignant tumors with glandular growth and/or supposed intracellular mucin as identified by periodic acid-Schiff (PAS) positivity. Gastric signet ring cell carcinomas are classified as diffuse type. A proportion of diffuse-type adenocarcinomas have previously been suggested to be of neuroendocrine origin. In the present study we examined gastric signet ring cell carcinomas for neuroendocrine differentiation. Of 11 gastric signet ring cell carcinomas, 8 contained areas with PAS-positive signet ring cells that also were immunoreactive for one or several neuroendocrine markers: synaptophysin, chromogranin A, and histidine decarboxylase, the latter an enterochromaffin-like (ECL) cell marker. Whereas PAS positivity was located in the central cytoplasm, neuroendocrine immunoreactivity was often located as a rim surrounding an otherwise non-immunoreactive cytoplasm, presumed to represent the area with PAS-positive material. These findings indicate that signet ring cell carcinomas could be of neuroendocrine origin. We propose that signet ring cell carcinomas develop by gradual dedifferentiation from ECL cells via signet ring cells with neuroendocrine immunoreactivity toward signet ring cells where the cytoplasm mainly consists of PAS-positive material. This finding could have implications for the classification and understanding of gastric carcinogenesis.     

An interview with Dr. Zbigniew Darzynkiewicz on his highly cited paper published in Cell Cycle

Zbigniew Darzynkiewicz is the Director of the Brander Cancer Research Institute and Professor of Pathology and Medicine at the New York Medical College. He received his M.D. (with the highest honors) and Ph.D. degrees from the Medical University of Warsaw in Warsaw, Poland and completed post-graduate studies at the State University of New York at Buffalo and at the Medical Nobel Institute of Karolinska Institute, Stockholm, Sweden. His research is focused on the regulatory mechanisms associated with cell proliferation, apoptosis and sensitivity to anti-tumor drugs. He published over 550 original articles in peer-reviewed journals, over 100 chapters and reviews, authored/edited 12 books and holds 7 US patents. His publications were cited over 23,000 times and 79 of them have over 79 citations each, which ranks him at number 79 in Hirsch’s impact “h-index.”     

Membrane Instability, Plasmalogen Content, and Alzheimer's Disease

Abstract: The normal stability of the cell membrane bilayer depends on its lipid composition being appropriate to the ambient (physiological) temperature, T_p. Membrane lipid composition may be altered by disease such that the bilayer is only stable at a new critical temperature, T^⋆, which may differ from T_p. In Alzheimer's disease (AD) temporal cortex, a defect of lipid composition has previously been identified, namely, a decrease in the ratio of plasmalogen to nonplasmalogen ethanolamine glycerophospholipids. Furthermore, for AD temporal cortex neural membranes, T^⋆≪ T_p, a finding confirmed in the present study in a larger series than previously, using a new method for obtaining T^⋆. This inequality between T^⋆ and T_p has been proposed as a putative contributory pathogenetic mechanism leading to membrane destabilisation in AD brain. The plasmalogen deficiency could account for the change in T^⋆ in AD, as shown by experiments where T^⋆ was measured for artificial lipid mixtures simulating brain membranes with varying plasmalogen/nonplasmalogen ratios. The critical temperature was found to be very sensitive to small alterations in plasmalogen content.     

Alcianblue/PAS or PAS/alcianblue? Remarks on a classical technic used in carbohydrate histochemistry

The combined alcian blue (AB)/PAS technic is widely used for the detection and characterization of mucosubstances in tissue sections. Mostly the sequence AB/PAS is used, occasionally also the reserved sequence PAS/AB. The present study shows clearly that the sequence of the combined technic, i.e. AB/PAS or PAS/AB is substantially influencing the results. So it could be demonstrated that by using the combination PAS/AB originally PAS-positive and AB-negative reacting mucosubstances become AB-positive. This could be caused by periodic acid oxidation followed by addition of hydrogen sulfite to aldehyde group thus providing secondary basophilic resp. AB positive material.     

Alcianblue/PAS or PAS/alcianblue?

Summary The combined alcian blue (AB)/PAS technic is widely used for the detection and characterization of mucosubstances in tissue sections. Mostly the sequence AB/PAS is used, occasionally also the reserved sequence PAS/AB. The present study shows clearly that the sequence of the combined technic, i.e. AB/PAS or PAS/AB is substantially influencing the results. So it could be demonstrated that by using the combination PAS/AB originally PAS-positive and AB-negative reacting mucosubstances become AB-posltive. This could be caused by periodic acid oxidation followed by addition of hydrogen sulfite to aldehyde group thus providing secondary basophilic resp. AB positive material.     

缩醛磷脂提取和制备的实验研究

通过将新鲜猪心切块、绞碎,制成组织匀浆后,再经一系列过程包括总脂的提取、从总脂提取物中制取磷脂酰乙醇胺、碱水解进一步纯化缩醛磷脂酰乙醇胺以及对获得的缩醛磷脂酰乙醇胺制剂鉴定其纯度。通过Ｉｏｄｉｎｅ ｄｉｓａｐｐｅａｒａｎｃｅ法测出缩醛磷脂酰乙醇胺制剂中的含量为１５．３５ｍｍｏｌ／Ｌ,通过Ｂａｒｔｅｌｌ法测出缩醛磷脂酰乙醇胺制剂中无机磷含量为１６．１０ｍｍｏｌ／Ｌ。因此,获得缩醛磷脂酰乙醇胺制剂的纯