Proliferating cell nuclear antigen (PCNA) expression in tooth primordia in the field vole (Microtus agrestis,Rodentia)

Cell proliferation in developing tooth germs has been studied particularly using bromodeoxyuridine (BrdU) incorporation into growing tooth primordia and by counting and three-dimensional (3D) reconstruction of mitoses in serial sections of developing teeth. PCNA has been proposed as an alternative marker of proliferation activity. The aim of our study was to detect immunohistochemically locations of PCNA-positive cells in developing tooth germs of Microtus agrestis (Rodentia). PCNA expression could be distinguished in oral epithelium and mesenchyme before first signs of dental lamina elevation. During bud, cap, and bell stages, positive immunostaining could be observed at defined sites in enamel organ, tooth papilla, and dental follicle. Rudimental tooth germs of the upper diastema, enamel knots, and inner enamel epithelium at day of ontogeny 18 and 19 showed negative reaction. PCNA marks cycling and early G0 cells and can be used successfully as a proliferation marker even in collection material.     

大肠粘膜c-myc蛋白和增殖细胞核抗原表达的相关性研究

本文应用c-myc癌基因蛋白和增殖细胞核抗原(PCNA)免疫组织化学方法,研究了它们在正常大肠粘膜、结肠炎、腺瘤和腺癌中的表达。发现结肠炎和肿瘤c-myc蛋白和PCNA阳性率较正常粘膜高。在管状腺瘤、管状绒毛状腺瘤和绒毛状腺瘤,阳性率呈进行性增加,但没有统计学意义。c-myc蛋白和PCNA表达呈相关性。肿瘤中阳性细胞呈异质性,所以要得到可靠的研究结果应多点取材。     

Expression of Early and Late Differentiation Markers (Proliferating Cell Nuclear Antigen, Syndecan-3, Annexin VI, and Alkaline Phosphatase) by Human Osteoarthritic Chondrocytes

Although osteoarthritis is characterized by a progressive loss of the extracellular cartilage matrix, very little is known about the fate of articular chondrocytes during the progression of the disease. In this study we examined the expression of syndecan-3, a marker of early chondrocyte differentiation, and annexin VI, a marker of late chondrocyte differentiation, in mammalian embryonic growth plate cartilage and normal and osteoarthritic human articular cartilage. Whereas syndecan-3 was expressed in the proliferative and hypertrophic zones of growth platecartilage, immunostaining for annexin VI waspredominately found in the hypertrophic and mineralizing zones of fetal bovine growth plate cartilage. Approximately 20% of chondrocytes were immunopositive for syndecan-3 in normal human articular cartilage, the number of syndecan-3-expressing chondrocytes significantly increased during the progression of osteoarthritis with more than 80% syndecan-3-positive cells in the upper zone of severely affected osteoarthritic cartilage. Similarly, the number of annexin VI-expressing cells significantly increased in the upper cartilage zones during the progression of osteoarthritis. Furthermore, immunostaining for proliferating cell nuclear antigen, a marker for cell proliferation, was detected in chondrocytes in the upper zone of osteoarthritic cartilage. Double-labeling experiments with antibodies against syndecan-3 and annexin VI revealed chondrocytes that expressed only syndecan-3, and cells that expressed both syndecan-3 and annexin VI. These results suggest that the expression of early (proliferating cell nuclear antigen, syndecan-3) and late differentiation markers (annexin VI, alkaline phosphatase) is activated in chondrocytes of osteoarthritic cartilage.     

Helminthfauna of Microtus (Microtus) cabrerae (Thomas, 1906) (Rodentia: Arvicolidae) in the Iberian peninsula: faunistic and ecological considerations.

Faunistic and ecological study of parasitic helminths of Microtus (Microtus) cabrerae (Thomas, 1906) (Rodentia: Arvicolidae) in the Iberian Peninsula. 70 specimens have been dissected, coming from 8 enclaves located in three Spanish provinces; 6 species of helminths have been detected (1 Digenetic Trematode, 4 Cestodes and 1 Nematode). Structure of helminthfauna of M. (M.) cabrerae in relation to remaining Iberian Arvicolids and the most conditioning ecological factors of the helminthfauna of the Rodent are analyzed.     

高压力对血管平滑肌细胞的形态、α-肌动蛋白和增殖细胞核抗原表达的影响--一种新的颈总动脉在体受力模型研究

为了研究机械应力在动脉重建中的作用和机制 ,本文建立了以压力改变为启动因素导致动脉重建的大鼠颈总动脉在体受力模型 ,并观察了高压力对血管平滑肌细胞 ( VSMCs)形态、α-肌动蛋白 ( α- actin)和增殖细胞核抗原 ( PCNA)表达的影响。以高压力 ( 160 mm Hg)和正常压力 ( 80 m m Hg)灌流大鼠颈总动脉 6h,组织学和免疫组织化学方法观察 VSMCs的形态、α- actin和 PCNA表达的变化。结果表明 ,大鼠颈总动脉在体受力模型在压力、脉压差和频率方面有较好的可控制性及稳定性 ;高压力使 VSMCs核常染色质增加 ,α- actin染色较正常压力组浅淡 ,表明 α- actin含量减少 ;PCNA表达阳性。大鼠颈总动脉在体受力模型的成功建立 ,为研究机械应力对动脉重建的作用提供了一种新的手段。高压力可诱导 VSMCs向合成型转换 ,细胞趋于增殖     

Distribution of L1-retroposons on the giant sex chromosomes of Microtus cabrerae (Arvicolidae, Rodentia): functional and evolutionary implications

Long interspersed nuclear elements (L1 or LINE-1) are the most abundant and active retroposons in the mammalian genome. Traditionally, the bulk of L1 sequences have been explained by the ‘selfish DNA’ hypothesis; however, recently it has been also argued that L1s could play an important role in genome and gene organizations. The non-random chromosomal distribution of these retroelements is a striking feature considered to reflect this functionality. In the present study we have cloned and analyzed three different L1 fragments from the genome of the rodent Microtus cabrerae. In addition, we have examined the chromosomal distribution of this L1 in several species of Microtus, a very interesting group owing to the presence in some species of enlarged (‘giant’) sex chromosomes. Interestingly, in all species analyzed, L1-retroposons have preferentially accumulated on both the giant- and the normal-sized sex chromosomes compared with the autosomes. Also we have demonstrated that L1-retroposons are not similarly distributed among the heterochromatic blocks of the giant sex chromosomes in M. cabrerae and M. agrestis, which suggest that L1 retroposition and amplification over the sex heterochromatin have been different and independent processes in each species. Finally, we proposed that the main factors responsible for the L1 distribution on the mammalian sex chromosomes are the heterochromatic nature of the Y chromosome and the possible role of L1 sequences during the X-inactivation process.     

Abnormal Nuclear Structures (Micronuclei,Nucleoplasmic Bridges,and Nuclear Buds) in a Pleomorphic Giant Cell Carcinoma of the Stomach

A rare case of pleomorphic giant cell carcinoma of the stomach in a 70-year-old man is reported. Characteristic microscopic findings included a general lack of architectural cohesiveness, aggregates of mononucleated or multinucleated giant cells, extensive areas of coagulative necrosis, and numerous mitoses. Immunohistochemically, tumor cells displayed cytoplasmic immunoreactivity for cytokeratin AE1/AE3 as well as overexpression of p53 and Ki-67. Electron microscopy revealed paranuclear tonofilaments bundles in giant cells confirming their epithelial nature. Furthermore, giant cells contained two or more nuclei with heterogeneous size, nucleoplasmic bridges, nuclear buds, and micronuclei. Similar abnormal nuclear structures have been closely related to breakage-fusion-bridge type of mitotic disturbances in tumor cell lines, and have not been previously reported in a human tumor.     

Expression of proliferating cell nuclear antigen (PCNA) in dysplasia of the bronchial epithelium

Proliferating cell nuclear antigen (PCNA) is expressed in cells in the cell cycle and has been studied as a marker of proliferation in lung and other tumours. We have noted immunocytochemical differences in PCNA expression between normal and neoplastic bronchial cells. As bronchial dysplasia is considered preneoplastic, we have examined PCNA expression in this condition. PCNA staining in 47 cases of bronchial dysplasia and 32 samples of normal bronchial epithelium was compared. Of the dysplasias, three were mild, 11 moderate, and 33 severe. A significant increase in PCNA counts over normal epithelium was seen only in moderate and severe dysplasias. In dysplasia, mitotic indices showed a significant positive correlation with the percentage of PCNA-positive cells. We conclude that in moderate and severe dysplasias there is an increase in the number of cells expressing PCNA and undergoing division, indicating abnormal growth control.     

Expression of carbonic anhydrase,cystic fibrosis transmembrane regulator (CFTR) and V-H-ATPase in the lancelet Branchiostoma lanceolatum (Pallas, 1774)

Sequencing of the amphioxus genome revealed that it contains a basic set of chordate genes involved in development and cell signaling. Despite the availability of genomic data, up till now no studies have been addressed on the comprehension of the amphioxus osmoregulation. Using primers designed on Branchiostoma floridae carbonic anhydrase (CA) II, cystic fibrosis transmembrane regulator (CFTR) and V-H⁺-ATPase, a 100 bp long region, containing the protein region recognized by the respective antibodies, has been amplified and sequenced in B. lanceolatum indicating the presence of hortologous V-ATPase, CFTR and carbonic anhydrase II genes in Branchiostoma lanceolatum. Immunohistochemical results showed that all three transporting proteins are expressed in almost 90% of epithelial cells of the skin in B. lanceolatum adults with a different degree of positivity in different regions of body wall and with a different localization in the cells. The comparison of results between young and adult lancelets showed that the distribution of these transporters is quite different. Indeed, in the young specimens the expression pattern of all tested molecules appears concentrated at the gut level, whereas in adult the gut loses its key role that is mostly supported by skin.     

Expression of Biomolecular Markers (Ki-67, PCNA, Bcl-2, BAX, BclX, VEGF) in Breast Tumors

We carried out a retrospective immunohistochemical study of Ki-67, PCNA, Bcl-2, BAX, BclX, and VEGF expression in tumors of two groups of breast cancer patients with favorable and unfavorable course of the disease. Considerably enhanced VEGF expression was detected in tumors of patients with early relapses of breast cancer. High VEGF expression was paralleled by high level of Ki-67 and PCNA expression in tumors. It can be hypothesized that expression of VEGF, Ki-67, and PCNA in primary tumor can be used for predicting the course of breast cancer or detecting the patients at a high risk of early relapses.     

Morphological peculiarities in the integument of enigmatic anomalurid gliders (Anomaluridae,Rodentia)

Scaly-tailed squirrels, the most poorly known group of gliding mammals, hold the record for variety of remarkable integument peculiarities. One of the most striking of these features is the scales on the tail, which apparently allow them to reduce energy costs when positioning themselves on a tree trunk. No less interesting is a peculiar spur that supports the flying membrane: the unciform element (‘spur’). Despite the peculiarity of such elements, their nature has not yet been studied. Using anatomical, histological methods and scanning electron microscopy we studied the structure of the skin and its derivatives in five of the six species from both genera of extant gliding scaly-tailed squirrels (Anomaluridae, Rodentia): Idiurus macrotis, Idiurus zenkeri, Anomalurus beecrofti, Anomalurus pusillus and Anomalurus derbianus. In addition to the common mammalian skin structures, such as hair, vibrissae, sebaceous glands, meibomian glands of eyelids and eccrine sweat glands of the palmar and plantar pads, these animals have unique species-specific skin derivatives (the tail scaly organ and its specific glands, vibrissae of the withers, patagium and its hair brush) that play a significant role in their adaptation to gliding and to their environment in general. The structure of the elbow spur is also described and hypotheses on its evolutionary origin from the tendon of the triceps muscle are presented.     

Expression of neural cell adhesion molecule (NCAM) during the first molar development in the mouse

NCAM, the neural cell adhesion molecule, was immunolocalized in the mandibular first molar tooth germ of the mouse. NCAM was first detected in the tooth germ of the late bud stage, where only the cells in the outer part of the condensed mesenchyme (primitive dental follicle) exhibited faint immunoreactivity. The entire dental follicle was intensely immunostained for NCAM from cap stage to the stage when root formation started. During root formation, NCAM disappeared from the follicular tissue surrounding the cervical root as well as from the part covering the crown top. This loss of NCAM proceeded in the direction of the root apex, but even after the tooth had achieved functional occlusion, NCAM was still expressed by the mesenchymal cells adjacent to the root apex. On the other hand, NCAM was negative in the dental papilla until birth. After birth, NCAM-immunoreactivity appeared in the basal portion of the dental papilla, but this NCAM-positive area gradually diminished in width during the root elongation. Instead, another NCAM-positive zone appeared in the core of the pulp during root formation. Even in the tooth that had already erupted, the pulp core contained cells that were strongly positive for NCAM immunostaining. In addition to its expression in the above two mesenchymal cell lineages, NCAM was transiently expressed by epithelial components of the tooth germ, some of the cells of the dental lamina and the enamel organ. The results suggest that NCAM participates in several processes of tooth development.     

Receptor Activator of Nuclear Factor Kappa B Ligand (RANKL) Modulates the Expression of Genes Involved in Apoptosis and Cell Cycle in Human Osteoclasts

It has been clearly established that receptor activator of nuclear factor kappa B ligand (RANKL) is a key cytokine involved in the differentiation of osteoclastic precursors of the monocytic/macrophagic lineage. However, relatively little information is available on the ability of RANKL to modulate the expression of genes controlling cell survival/apoptosis and proliferation in human osteoclastic cells in comparison to macrophages. For this purpose, CD14⁺ human peripheral blood mononuclear cells, which express the cognate high affinity receptor activator of nuclear factor kappa B (RANK), were differentiated along the macrophagic or osteoclastic lineage by adding macrophage‐colony stimulating factor (M‐CSF) or M‐CSF plus RANKL in culture for 12 days. RANKL up‐regulated the expression of the chemokine MIP1α, which potentiates osteoclastic differentiation and simultaneously activated both anti‐apoptotic (Bcl‐2) and pro‐apoptotic (CIDEB, PYCARD, and BAK‐1) genes. Moreover, RANKL markedly up‐regulated cylin D2, while it significantly decreased the levels of cyclin A, cyclin‐dependent kinase 2, and other cyclin‐dependent kinases, in keeping with the notion that end‐stage osteoclasts are nondividing cells. Finally, a long‐term exposure of RANKL up‐regulated the adaptor protein TRAF3 but not TRAF6. Anat Rec, 2007. © 2007 Wiley‐Liss, Inc.     

Expression of the neural cell adhesion molecule (NCAM) during second- and third-molar development in the mouse

Distribution of the neural cell adhesion molecule (NCAM) during the development of the mandibular second- and third-molars of the mouse was studied by indirect immunofluorescence techniques. At the initial stage, NCAM was intensely expressed by the mesenchymal cells surrounding the dental lamina, and by the cap stage NCAM expression by the mesenchymal cells became restricted to the dental follicle. After that, in addition to the follicular mesenchyme, some cells in the basal part of the dental papilla showed NCAM-immunoreactivity for a while after the hard tissue formation had started. During root formation, the follicular cells lost NCAM first from the level of the cervical root and later from the coronal part, while an additional NCAM positive area appeared deep in the dental papilla. Even after the teeth had erupted, NCAM was expressed in the tissue surrounding the apical root and in the pulp core. During the initial and bud stages, the pattern of NCAM expression in the second and third molars was different from that in the first molar, where NCAM was found only after the late bud stage; while from the cap stage onward, it changed in the same sequence as in the first molar. The different pattern of NCAM expression implies that there is a difference in developmental events between the early stages of the first and the other two molars. On the other hand, the common sequence of NCAM expression in the tooth germs later than the cap stage suggests that NCAM plays an essential role in the formation of the basic structure of the teeth and periodontal tissues.     

Abnormal Nuclear Structures (Micronuclei,Nuclear Blebs,Strings, and Pockets) in a Case of Anaplastic Giant Cell Carcinoma of the Thyroid: An Immunohistochemical and Ultrastructural Study

The authors report a case of a 70-year-old woman with an anaplastic giant cell thyroid carcinoma, along with immunohistochemical and electron microscopic findings. Histologically, the tumor is characterized by mononucleated and multinucleated giant cells, lack of architectural cohesion, atypical mitoses, and extensive areas of coagulative necrosis. Tumor cells showed AE1/AE3 positivity as well as nuclear overexpression of p53 and ki-67. Semithin sections revealed multiple nuclei with heterogeneous size ranging from micronuclei to large-size (giant) nuclei. Micronuclei were confirmed by electron microscopy that disclosed also the presence of nuclear blebs, strings, and pockets. Morphological findings of these abnormal nuclear structures in conjunction with p53 and Ki-67 nuclear overexpression suggested a faulty mitotic checkpoint/mitotic catastrophe in the progression of anaplastic giant cell thyroid carcinoma.     

Validation of the use of proliferation markers in canine neoplastic and non-neoplastic tissues: comparison of KI-67 and proliferating cell nuclear antigen (PCNA) expression versus in vivo bromodeoxyuridine labelling by immunohistochemistry

In order to evaluate the suitability of Ki-67 and proliferating cell nuclear antigen (PCNA) for determination of proliferative activity, the immunohistochemically determined nuclear expression of these antigens in canine non-neoplastic and neoplastic tissues was compared with the results of in vivo bromodeoxyuridine (BrdU) labelling, which - by measurement of the fraction of S-phase cells - is considered as the standard in the analysis of proliferative activity. The samples investigated consisted of non-neoplastic mammary and lymphoid tissues, and of benign and malignant (primary/metastatic) mammary tumours, and malignant lymphomas. Great regional heterogeneity prevented determination of an overall labelling index (LI) in normal lymphoid tissues. In the remaining combined group of samples, LI values were significantly ranked in the order PCNA>Ki-67>BrdU. However, the correlation of Ki-67 or PCNA as compared to BrdU LI values was only moderate in the combined group [approximately 0.5, Spearman rank test] as well as in most subgroups, whilst it was very poor in the group of primary mammary cancers. These observations indicate that Ki-67 or PCNA LIs as markers of proliferation do not evenly match in vivo BrdU labelling.