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This short overview and updates expresses our brain-and-heart homage to George Emil Palade, “the most influential cell biologist ever”. In his 1971 paper Palade wrote for Albert Claude, the founder of biological electron microscopic method: “Seldom has a field owed so much to a single man”. Herein, we articulate the same words for George Palade, the Teacher of many generations in cell biology research and education. Accordingly, we focus on two paradigm shifts in the cell biology, namely (i) the transition from light to transmission electron microscopy in studying cell protein secretion made by George Palade, and (ii) the transition from contractile to secretory phenotype of vascular smooth muscle cells made by Maria Daria Haust followed and developed by our research group. Altogether, we argue that one of the present challenges in vascular biology is to cultivate secreto-centric thinking and thus further focusing on how we could make the vascular muscle’s secretory pathways work for the benefit of human’s cardiovascular health.

Claudins (CLDNs), of which 24 types have been identified, are integral transmembrane proteins of the tight junctions that are critical for maintaining cell adhesion and polarity. They also act as selective barriers. Cells and tissues are characterized by individual CLDN patterns; the composition and levels of expression change during differentiation and tumor formation. Alterations in the expression of individual CLDNs have been detected in several carcinomas and shown to be related to progression and invasion; however, their role in carcinogenesis is controversial. Using a panel of polyclonal (CLDNs 1, 3, and 7) and monoclonal (CLDNs 2 and 4) antibodies, CLDN pattern and expression were studied by immunohistochemistry in 105 cervical tissue specimens, including normal epithelia (n = 20), cervical intraepithelial neoplasias (CINs; CIN 1/2, n = 27; CIN 3, n = 10), carcinoma in situ (CIS, n = 15), and 33 squamous keratinizing and nonkeratinizing invasive carcinomas. No CLDN 3 was observed in normal or intraepithelial neoplastic cells, but significantly increased expression of CLDNs 1, 2, 4, and 7 was detected in the CIN/CIS lesions and invasive carcinomas compared with the normal tissues (P < .001) and reduced reactivity of CLDNs 1 and 2 was observed in invasive cervical cancers compared with CIN 3/CIS (P = .0001) and of CLDNs 2, 4, and 7 compared with CIN 1/2. These results indicate increased expression of CLDNs in the early phase of carcinogenesis in intraepithelial lesions, which decreases during progression to invasive disease. Expression of CLDN 1 was strongest in premalignant stages; thus, it may serve as a good diagnostic marker for the detection of CIN.

Summary The expression and the distribution of tenascin as well as the extent of blood vessel formation (angiogenesis) were investigated in 70 invasive human breast carcinomas. Formalin-fixed, paraffin-embedded specimens were stained with monoclonal antibody against tenascin-C (DAKO and Biogenex). Anti-CD31 antibody (Biogenex), an acknowledged marker of stromal angiogenesis, was used to detect endothelial cells. Tenascin immunostaining was positive in the tumours around the persisting normal ducts, around tumour-cell nests, in the neostroma, in some tumour cells, and it was found in or around vascular channels. Tumour vascularity was assessed by quantitative vascular grading (Chalkley point count) and was related to the localization and intensity of tenascin immunoreactivity. 19 tumours (27.1%) were scored as low, 35 (50%) as medium, and 16 (22.9%) as having a high vascular grade. The positive correlation between the vascular grade and the tenascin immunopositivity in tumour stroma was observed. Our results suggest that tenascin expression may be associated with endothelial cell activation and may play an important role in tumour angiogenesis.

A case of black insulinoma is reported. The color was due to a cytoplasmic pigment. Immunostaining for neuron-specific enolase and chromogranin was positive in the tumor cells, and the pigment granules themselves reacted with the chromogranin antibody. Numerous beta cell type dense core granules as well as atypical granules were found by electron microscopy. An important finding was that the dense core granules contribute to the lipofuscin pigment formation.

Cell-cell and cell-extracellular matrix interaction is crucial in tumor progression. Tight junction (TJ) proteins as occludin and claudins (CLDNs) play important role in this process together with several extracellular matrix components, as syndecan. Our previous work suggested significant changes in the expression of claudins even in the early stages of cervical carcinogenesis. The aim of our present work was to study the expression of occludin and syndecan-1, as compared to CLDNs, in early phases of cervical carcinogenesis. Paraffin sections of 50 samples were studied by immunohistochemistry, including cervical intraepithelial neoplasias (CINI-II-III), in situ carcinomas (CIS) and normal cervical samples. Occludin and CLDN-2 were found colocalized in the basal layer, while syndecan-1 and CLDN-1, -4 and -7 were coexpressed in the parabasal and intermedier layers in normal epithelia. Intensity of occludin staining decreased in CIN/CIS lesions, although it was more extended towards the upper epithelial layers with inverse relation with grades, as seen in the case of CLDN-2 expression. CLDN-1, -2, -4 and -7 were detected in the entire epithelium in CIN, showing decrease in CIS. The progression of CIN was associated with reduced syndecan-1 expression, in contrast to CLDN-1, -4 and -7 which increased toward CIS. The obtained data suggest that significant changes occur in the composition of cell adhesion complexes even in early stages of cervical carcinogenesis. The pattern of expression is characteristic for the alteration, the changes in the different components, however, are not parallel with each other.

The pathogenic role of Chlamydia pneumoniae in late coronary bypass graft failure has not yet been extensively investigated. We examined failed and new arterial/venous bypass grafts using immunohistochemistry, polymerase chain reaction (PCR), and serology. Thirty-four long-term failed grafts and 28 new grafts were examined in 21 patients undergoing redo coronary artery bypass grafting (CABG). Immunohistochemically, 28 (82%) failed grafts were positive in the intimal–medial compartment, and 33 grafts (97%) were positive for C. pneumoniae in the adventitia. Thirteen (46%) and 27 (96%) new grafts showed infection in the intima–media and in the adventitia, respectively ( p 0.05 ). Immunohistochemically, the overall presence of C. pneumoniae in all vessels examined was 66% in the intima–media and 97% in the adventitia ( p 0.05 ). C. pneumoniae was detected by PCR in 19 (31%) of all the vessels examined. C. pneumoniae seems to be frequently present in grafts of patients considered for redo CABG in Hungary. The adventitia of both failed, and new grafts particularly often contained C. pneumoniae. The results suggest that there exists an adventitial baseline infection from which infection of the inner wall layers develops, depending on local microenvironmental conditions. This is the first study to evaluate chlamydial infection in arterial/venous coronary grafts by immunohistochemistry, PCR, and serology.

In non-atheromatous segments of coronary arteries a sequence of preatherosclerotic changes was identified which consisted of medial thickening followed by intimal thickening. More recently, Chlamydia pneumoniae seropositivity was associated with enhanced intima-media thickness of arteries. In the present study the intimal and medial thickness of coronary artery of young adults were measured, and were correlated with the presence of Chlamydia pneumoniae antigens. Proximal and distal segments of the left anterior descending coronaries (LAD) obtained at autopsy from young adults (15-34 years) were studied. The thickness and cellular density of the intima and of the media without clear-cut atherosclerotic changes were measured by image analysis. The hypertrophy index was calculated as the ratio of cell density and the thickness of the respective layer. Atherosclerotic lesions occurring elsewhere in the same coronary were noted and graded by severity. The presence of Chlamydia pneumoniae verified by immunohistochemistry was correlated with the severity of lesions and with the hypertrophy index. In the proximal segments, atherosclerosis of LAD was associated with the widening of both the intima and the media of lesion free-sites. In the distal coronary segments the proportion of the intimal thickening had a significant association with atherosclerosis. Compared to non-infected arteries, Chlamydia pneumoniae infection was associated with higher hypertrophy index in the intima as well as in the media. The rate of Chlamydia pneumoniae positivity increased with the severity of lesions. As a conclusion: in the LAD coronary, the intimal thickening is the main preatherosclerotic change. Chlamydia pneumoniae may favour arterial wall hypertrophy and plays a role in lesion progression.

Osteopontin, a non-collagen, Ca-binding phosphoprotein, plays roles in both physiological and pathological mineralisation processes. Its expression may be severely altered in malignant tumours.To investigate the presence of this protein in breast tumours showing mammographic microcalcifications.In this study the authors investigated immunohistochemically the osteopontin expression in 26 breast tumours. The elemental composition of the microcalcifications of the same tumours was also examined with energy dispersive spectrometer linked to a scanning electron microscope.Osteopontin expression was found within the tumours in the tumour cells, in vascular smooth muscle cells, in the myoepithelial cells of the normal ducts, and in macrophages. Extracellular osteopontin positivity was seen around calcified foci. There was no correlation between osteopontin positivity and the examined prognostic factors: grade, type and size of the tumour, axillary lymph nodes status, or age. Two types of microcalcifications were seen: 1. Ca-phosphate only, 2. Those composed of various other elements.The presence of osteopontin in the vicinity of calcified foci and in the vessel walls seems to support the hypothesis that osteopontin takes part in calcification process.

The authors determined serum PSA levels in combination with digital rectal examination (DRE) and evaluated their role in the differential diagnosis of prostate diseases with special reference to cancer. The possible causes of differences between the observed cut-off level of PSA and the standard level PSA were analyzed. In the last few years the PSA determination found its clinical role in the diagnosis of prostate cancer.

A malignant process interferes with the normal 'programme' of extracellular matrix biosynthesis and can modify extensively the structure and composition of the matrix. This effect appears to be attributable to several processes such as direct production of some selected matrix macromolecules by malignant cells or indirectly by the production of factors by malignant cells interfering with the regulation of normal matrix production. Other possibilities may also exist, such as the direct action of an environmental carcinogen on otherwise normal mesenchymal cells. The result is a more or less profound modification of tissue structure and composition with possible feedback effects on the malignant process. Some examples will be discussed such as elastin production by some tumours as well as the biosynthesis of some other selected matrix macromolecules as tenascin and osteopontin by breast tumours. Although the detailed mechanisms of these specific matrix productions is not yet completely elucidated, the rapidly increasing knowledge on the regulation of specific matrix production process and deranged matrix production might represent a new area of crosstalk between cancer research and matrix biology.

Summary Chlamydia pneumoniae has emerged as the most likely pathogen to have a causative role in the development and/or for progression of atherosclerosis. Evidence for this is based on epidemiological and pathological studies. In an effort to better understand the significance of finding C. pneumoniae in atheromata, we examined coronary artery segments of young adults (15–34 years) with and without atherosclerosis. Left anterior descending coronary arteries (LAD) of 74 young adults who died suddenly were examined histologically and for the presence of C. pneumoniae by immunohistochemistry. C. pneumoniae was identified in advanced lesions (Stary types III to VI) in 17 of 32 cases (53%), and in early lesions (Stary type I–II) in 8 of 37 cases (21%), mainly at the proximal segments of the LAD. C. pneumoniae was not found in the intimal and medial layer of normal-appearing coronary arteries. C. pneumoniae was detected in the adventitia in 51 (67%) coronary arteries:in 27 of normal arteries and early lesions (64%), and in 24 of atherosclerotic lesions (75%). C. pneumoniae was found most often in macrophages, less offen in smooth muscle cells. We also observed a correlation between C.pneumoniae positivity and cigarette smoking. In conclusion: C. pneumoniae may relate to the severity of atherosclerosis in young people, and it may thus initiate atherosclerotic injury or facilitate its progression with other risk factors.

Summary The purpose of this study was to determine the effects of ovariectomy and long-term combined sexual hormone replacement on the gap junctional protein, connexin 43 (Cx43) of aortic medial smooth muscle cells in rats. Twenty non-pregnant mature Wistar female rats were divided into five groups (four animals in each group). Group A underwent ovariectomy, Group B underwent ovariectomy and received estradiol propionate, Group C underwent ovariectomy and received medroxyprogesterone acetate and Group D underwent ovariectomy and received both hormones. Group E was sham-operated and used as control. After 15 weeks of treatment, thoracic aortas were removed and immunohistochemistry was carried out using a specific fluorescent antibody against Cx43. Tissue sections were examined by confocal laser scanning microscopy and analysed by the Scion Image program. All five different groups had the same distribution and extent of Cx43 in the aorta. Neither the ovariectomy nor the hormone replacement had any effect on the Cx43 expression of aortic smooth muscle cells in rats as compared to control animals. These results indicate that sexual steroids do not influence the gap junctional protein Cx43 of the medial layer of aorta in rats. They may suggest that the beneficial effects of estrogen are not mediated via gap junctions in the human aorta either.

Our knowledge of the anatomy of lower extremity veins is based on description of three main groups of veins more than a century ago by John Gay.1 Since Gay’s original communication, important data have accumulated on venous components of the cutaneous micro circulation, the functional anatomy of venous muscle pump, distribution and function of venous valves and the location and possible role of perforating veins. These observations have allowed a functional approach to understand the complex and intriguing anatomy of the venous system of the lower extremity.

A thorough knowledge of topographical anatomy is essential for learning new surgical procedures. Operations on the venous system of the legs require understanding the anatomy of the main groups of veins, their location and relationship to adjacent structures within the different compartments of the leg. Veins in few regions of the human body have more variability than in the lower limbs, and few areas in the body have a more complex topographical anatomy than the subfascial leg compartments.

In our investigations we accomplished quantitative measurements using energy dispersive X-ray analysis on mineral elements of pollen grain samples belonging to different fruit species such as apple (Malus domestica Borkh.), pear (Pyrus communis L.), cherry (Cerasus avium L.), sour cherry (Cerasus vulgaris Mill.), European plum (Prunus domestica L.), Japanese plum (Prunus salicina Lindi.), almond (Amygdalus communis L.), apricot (Armeniaca vulgaris Lam.), quince (Cydonia oblonga Mill.) and peach (Persica vulgaris Mill.). Pollen grains are particles but, as a result of their size ranging in our samples from 19.2 to 31.2 μm in diameter, they are considered as bulk samples. Link’s ZAF-P/B program was used for the quantitative evaluation of data. The main constituents were Mg, P, S, Cl, K, Ca, while others such as Na, Al, Si and Cu, whose intensity in the spectra did not exceed the standard deviation, were excluded from further quantitative evaluation. Differences in the elemental concentrations of different fruit pollens were significant (p > 5%).

Summary Out of 54 open renal biopsies performed on children, 17 were selected and studied not only bylight microscopy but also by electron microscopy. Statistical-mathematical analysis of basement membrane thickness-measurements was carried out. It is concluded that electron microscopic investigation of the renal biopsy material is required in selected cases involving diagnostic or therapeutic problems. In nephrotic syndrome it may help above all to separate minimal changes from early forms of glomerular lesion of the focal sclerosing type. Electron microscopy can also be useful in any of the mono- and oligosymptomatic renal diseases. In those cases only electron microscopy makes it possible to provide exact morphological diagnosis which is necessary to the proper therapy.

Summary The main feature of an atherosclerotic plaque is the formation of a new tissue in the arteries. In this respect atherosclerosis is similar to other conditions where non-neoplastic tissue formation occurs like in embryogenesis, in healing or in repair processes. A progressive intimal thickening occurs in the early phase of human atherosclerotic lesions and also in certain experimental models. Long-standing aortic intimal thickening could be induced by mechanical injury to the inner surface of the aorta with a microsurgical instrument, which causes controlled endothelial denudation. The injury is followed by an arterial remodeling. The latter process is caused by the development of an intimal plaque which consists of two main components: the smooth muscle cell (SMC) and the intercellular matrix. The matrix components are mostly synthetized by the SMC. Two distinct SMC populations could be distinguished by morphological means in the intimal proliferation: the synthetizing type and the proliferating type. Their role will be discussed and their morphological appearance will be compared with SMC present in other lesions.

SMC-s both in vivo and in vitro have a function in matrix macromolecule production. Under certain conditions the synthesizing function of the SMC-s seems to be more important than their contractile function. The modified less differentiated SMC-s participate in the synthesis of connective tissue macromolecules while the differentiated "resting" SMC-s maintain the functional activity.

alpha-Elastin methylester forms with proteoglycan subunits and structural glycoproteins water-insoluble complexes which after staining with phosphotungstic acid (pH 7.0), reveal at large magnification bundles of slender smooth fibrils with a diameter from 4 to 5 nm. Similar fibrils can be observed after interaction of alpha-elastin methylester with chondroitin sulfate, but only if fixation with glutaraldehyde is used prior to staining. Phosphotungstic acid alters the organization of the complexes, in particular those formed with ligands of low molecular weight. The possible biological significance of the interactions studied is discussed.

The development of intimal thickening in the rat aorta was follwed up in model experiments and the ultrastructure of its cell components was studied. The surface of the proliferation was covered by endothelial cells, the majority of which had obviously crept there from the intact part of the aortic wall. In a lesser degree also the blood's cellular elements appeared to participate in the formation of the endothelial cell row. The multilayered proliferation which had developed between the row of endothelial cells and the damaged internal elastic lamina consisted exclusively of smooth muscle cells; at least we failed to identify any other cell type within it. In the smooth muscle cells of aged proliferation, degeneration was signified by the presence of myelin figures. Neutral fat droplets appeared also in the extracellular space, having apparently passed through the endothelium from the blood plasma to the thickening. It seems, therefore, that there is no notable morphological difference between the intimal thickening serving as experimental model and the early arteriosclerotic lesion and accordingly a closer study of regenerative intimal proliferation may contribute some new information on the pathomechanism of degenerative and regenerative processes taking place in the vessel wall.

Finger pulp biopsies of 31 patients suffering from Raynaud's syndrome (RS) were investigated by light and electron microscopy. The patients suffered from various underlying diseases. Interest was focused on the vascular changes and nerve and connective tissue alterations, especially the elastic fiber changes. In the dermal capillaries, endothelial swelling, narrowing of the lumen, increased numbers of intracytoplasmic filaments and Weibel-Palade bodies, and various types of basement membrane alterations were observed. The basement membrane of the unmyelinated nerves showed similar alterations to the capillary basement membrane. Severe degenerative changes were found in the myelin sheath of the myelinated axons. In the dermal connective tissue, severe elastotic degeneration and alterations of the collagen fibers were observed. The data probably represent the ultrastructural picture of the end stage of this syndrome.

Physicochemical and ultrastructural properties of cholesterol ester complexes of fibrous elastin and of K-elastin were studied using differential scanning calorimetry (DSC) and electron-microscopy. The number of molecules of the different fatty acids retained in these elastin complexes varied between large limits according to the nature of the fatty acid of the cholesterol ester, ranging from 0.1 microgram/200 mg of elastin for cholesterol arachidonate to 48 micrograms/200 mg elastin for cholesterol palmitate. The ultrastructural studies confirmed the association of cholesterol esters with fibrous elastin and soluble K-elastin. The DSC-data showed that the temperature of transition between the crystalline and the liquid crystalline state shifts towards higher temperatures (well above body temperature) when cholesterol esters are bound to elastin: no liquid crystalline mesophase was observed in cholesterol oleate or linoleate-elastin complexes at 37 degrees C; melting of the crystalline structures took place at 51 degrees C and at 42 degrees C for the cholester Holeate and linoleate-elastin complexes respectively. These results indicate that in the elastin-bound form the crystalline-liquid crystalline transition is inhibited. The "stabilization" of the crystalline structure of cholesterol esters by fibrous elastin may be of biological significance and account for the crystalline deposits seen in advanced atherosclerotic lesions at sites of elastic fiber breakdown.

Chlamydia trachomatis infection has been investigated in the prostate gland using in-situ DNA hybridization and transmission electron microscopy. Sixty-four samples of tissue removed by trans-urethral resection or by open surgery from men with benign prostatic hypertrophy (BPH), were examined histologically and 20 of the cases were found to be positive for mononuclear cell infiltration. Some samples were then investigated using in-situ hybridization and transmission electron microscopy. Out of 20 tissue samples with mononuclear cell infiltration, nine were found to be positive for C. trachomatis infection.