Morphological and mechanical changes in juxtarenal aortic segment and aneurysm before and after open surgical repair of abdominal aortic aneurysms.

OBJECTIVE: The aim of study was to assess how the ultrastructure of the wall of aortic aneurysms, sac and neck influences aortic wall distensibility and proximal dilatation 2 years after open repair. METHODS: Biopsies for electron microscopy were taken from aneurysmal sac and neck of 30 patients. Patients were assessed by computed tomography (CT) and ultrasound for aneurysm diameter and distensibility (M-mode ultrasonography). RESULTS: Postoperative CT of the aortic stump distinguished two groups. Group I (n = 11) with little enlargement, median 1 mm (1-3 mm) and group II (n = 19) with significant aortic enlargement, median 5.2 mm (4-12 mm). In group II, changes in elastic fibres in the aneurysm neck were comparable to, but as extreme as in the aneurysm sac. For group I, the distensibility of the aneurysmal sac was significantly lower than in the neck or at the renal arteries. For group II, the distensibility in both the neck and sac was significantly lower than at the juxtarenal segment (p = 0.01). The biopsies of group II patients showed the extensive degeneration of normal architecture, which was associated with altered wall distensibility in both the aneurysmal neck and sac. CONCLUSIONS: Disorganisation and destruction of normal aortic architecture at the ultrastructural level are associated with decreasing aortic distensibility. Low aortic neck distensibility is associated with proximal aortic dilatation at 2 years postoperatively.

Correlation between the stage of cystic fibrosis and the level of morphological changes in adult patients.

The aim of this study was to evaluate the correlation between the stage of cystic fibrosis and the intensity of accompanying morphological changes--including transmission electron microscopy--within bronchial mucosa. The stage of the disease was assessed on the basis of clinical status and radiological and endoscopic examination. We focused on morphological changes in epithelial cells, the presence of metaplasia and/or dysplasia, the type of inflammatory infiltrate, and the presence of epithelial ulcerations, thickening of epithelial basement membrane and collagenization of lamina propria. We found two clinically different patients groups. The first one was in a poor clinical condition, advanced inflammatory fiberoptic bronchoscopy and radiological changes, multiple exacerbations, and with chronic inflammation and only focal appearance of ciliated epithelium. Moreover, squamous cell metaplasia and dysplasia was diagnosed in 3 and 4 cases, respectively. The other group, in a better clinical condition, had normal BMI and small changes on chest Xray. In this group the diagnosis of cystic fibrosis was made at later age. Two patients from this group displayed features of acute phase; ciliated epithelium was covering the whole sample. After statistical analysis, we found a correlation between the clinical course and the morphological changes in bronchial mucosa. Bronchial ulcerations, squamous cell metaplasia and dysplasia were found in the group with the more severe clinical course.

Myopathy as the first symptom of hypokalemic periodic paralysis--case report of a girl from a Polish family with CACNA1S (R1239G) mutation.

PURPOSE: Presenting the case of unusual onset hypokalemic periodic paralysis (HypoPP) where myopathy had developed two years before paralysis occurred. MATERIAL AND METHODS: A Polish three-generation family with HypoPP and mutation in CACNA1S (R1239G) has been investigated. Clinical presentation with unusual onset of the disease, biopsy results and genetic research in one family member were described. CONCLUSION: HypoPP is a rare disease it needs to be taken into consideration not only in cases of paroxysmal weakness but also when there is myopathy of unknown origin.

Intracellular expression of pro-inflammatory cytokines (IL-1alpha, TNF-alpha, and IL-6) in chronic hepatitis C.

The study aimed at localizing TNF-alpha, IL-1alpha, IL-6 at light and electron microscope levels in patients with chronic hepatitis C, using the immunocytochemical techniques in biopsy material from patients with chronic hepatitis C and at comparing the expression of the cytokines with histopathological changes. Our studies demonstrated an augmented expression of all cytokines in liver biopsies in chronic hepatitis C, in comparison with respective values, obtained in control biopsy material. The highest expression of the cytokines was observed in hepatocytes. That was confirmed by electron microscopy, which demonstrated the cytokines mainly in altered ER cisterns and in the cytoplasm. In children, the expression of IL-1alpha was negatively correlated with staging, while in adult patients; the staging was positively correlated with the expression of TNF-alpha. The new element involves demonstration of cellular and subcellular expression of TNF-alpha, IL-1alpha and IL-6 in hepatocytes in in vivo infection.

The expression of PDGF, its receptors and capillary morphometry in the developing lung: quantitative studies.

The aim of the present study was the evaluation of the expression levels of PDGF-A and B, and PDGFR-alpha, andchanges of capillary cross-sectional areas during terminal vascular network formation in the developing lung. Pathogen-free fetuses and newborns used for rats were used. Lungs were processed for frozen section immunohistochemical staining and parallel tissue specimens were prepared for transmission electron microscopy. The expression levels of PDGF-A and B, and PDGFR-alpha were obtained using quantitative image analysis. Additionally. the morphometry of capillary cross-sectional area was performed. We found that the expression levels of the studied antigens fluctuated, and the highest values were found in21-day old fetuses (p < 0.0001). Although there were no significant differences of the area of "a single" capillary, the area of the endothelium decreased significantly in perinatal period (p <0.0001). We conclude that the statistically significant changes of the studied antigens' expression as well as the of capillary cross-sectional area occur in the perinatal period,

The influence of simultaneous whole body exposure to cigarette smoke and low-protein diet feeding on fertility, reproduction, and progeny of rats.

The aim of the present study was to evaluate the toxicological influence of different concentrations of tobacco smoke and low-protein diet on fertility, reproduction, and survival of offspring of Wistar female rats. We studied three generations, two litters in each. There were 192 animals in parent generation. Animals were exposed to three different concentrations of tobacco smoke that were reflected by the carbon monoxide (CO) concentration (500, 1000, and 1500 mg CO/m3 air) during 11 weeks (six before and two weeks during mating, and three weeks of pregnancy). Additionally, animals were divided into two subgroups that were fed normal or low-protein diet. In the present study, we concentrated on the indices describing the reproduction ability, fertility, and rearing of pups of mothers exposed to tobacco smoke. The parameters of newborns survival were estimated also. We concluded that neither tobacco smoke exposure nor the low-protein diet changed the duration of pregnancy. The tobacco smoke in all doses and low-protein diet had a negative influence on fertility, reproduction, and survival of newborns from exposed mothers. This influence was mainly attributed to the tobacco smoke exposure. The levels of carboxyhaemoglobin were increasing with increasing tobacco smoke concentration.

Additional cell damage after transpupillary thermotherapy in choroidal malignant melanoma.

The aim of this study was to evaluate the effectiveness of transpupillary thermotherapy (TTT) in generating tumour necrosis by light and electron microscopy, as well as to evaluate additional cell damage in the area directly adherent to the necrotic zone. Four eyes of four patients diagnosed with intraocular malignant melanoma of the uvea were treated experimentally with diode laser TTT. In all cases a standard technique was used. All eyes were enucleated: one eye the day after TTT, two eyes 2 days after TTT, and one eye 6 weeks after TTT. Immediately after enucleation the eyes were immersed in standard Karnovsky's fixative with cocodylate buffer and prepared for light and electron microscopy. In the treated area of all four melanomas we found a dense band of necrotic tissue (zone A) consisting of an amorphous mass of dead cells sharply demarcated from the rest of the neoplastic tissue. Next to this zone was a more eosinophilic and also sharply demarcated band (zone B) that consisted of similar but less intensive changes. In the next band (zone C), marked injury to the cellular membrane and subcellular structures were seen on electron microscopy. The next band (zone D) consisted of changes mainly observed only within the cytoplasm of neoplastic cells and significantly less intensive than those in zone C. Outside zone D tumour cells that were normal in appearance were seen. No scleral alterations induced by heat were found. We concluded that after TTT the cytotoxic effect gradually decreases in proportion to the distance from the central point of the diode laser spot, with additional cell damage in the area adjacent to the necrotic zone. The interval between TTT and enucleation had no influence on the histological results.

[The Lublanska classification of precancerous lesions of the larynx]. / Klasyfikacja Lublanska stanów przedrakowych krtani.

Laryngeal carcinomas are preceded by precancerous lesions in about 20% of cases. The macroscopical appearance of these lesions is not enough characteristic to define their malignant potential. The accurate identification of epithelial abnormalities of the laryngeal mucosa requires biopsy and microscopic evaluation. There are many histological classifications of laryngeal precancerous lesions used at present. Many of them are highly subjective and have low reproducibility. Moreover, the different grades of these classifications not always give distinct guidelines for clinician concerning the treatment modality. The Ljubljana classification seems to be easier, more readily applied and more reproducible. It uses the name "epithelial hyperplastic laryngeal lesion" (EHLL) which includes all alterations in laryngeal squamous epithelium. The four grades of EHLL are: 1. simple hyperplasia (thickening of epithelium due to augmentation of normal prickle cells), 2. abnormal hyperplasia (with increase of basal-like cells), 3. atypical, or risky hyperplasia (epithelium thickened by increase of basal-like cells with pronounced atypical features), and 4. carcinoma in situ (i.e. full thickness change with the features of malignancy but without stromal invasion). The criteria of Ljubljana classification are precise and gives a possibility to make a more clear-cut separation of cases with risk of developing carcinoma from those without it.

Quantitative morphometry--an easy and inexpensive method for comparative studies of antigen expression during development.

The morphometric studies due to the development of computerized techniques became a very suitable tool for quantitative analysis. In the present paper we describe an easy and relatively inexpensive method useful for comparative studies on changes of expression of examined molecules during development of tissue. We used with a success this method for studies on the expression of CD31 adhesion molecule during the fetal and newborn lung development. We found that expression of CD31 is regulated during the perinatal period. It might be related to the dynamic changes in developing vascular bed in rat lung.

Correlation of morphological alterations of light and electron microscopy in chronic type B and C hepatitis.

Chronic type B and C hepatitis involves inflammatory lesions of a variable intensity and variably advanced fibrosis. Considering current, progressively growing requirements for correct evaluation of lesions in liver biopsies, an attempt was made to appraise suitability of selected techniques for a broadened histopathological diagnosis. The lesions were evaluated at the level of light and electron microscopy. Material for the study consisted of liver biopsies obtained from adults and children (n = 60) with serological markers of chronic type B or type C hepatitis. Routine techniques of staining for light and electron microscopy, as well as the techniques of Brachet and Feulgen, were applied. HBcAg expression and HBV-DNA detection in children with chronic type B hepatitis were studied employing the avidin-biotin peroxidase complex (ABC) technique and in situ hybridisation with the ImmunoMax signal amplification. Slight or moderately intense inflammatory lesions (grading of 1 to 2 points) and a low level of fibrosis advancement (staging of 1 to 2 points) prevailed in the material, independently of the etiologic agent involved and age of the patient. Both in children and in adults, extensive lesions in the nuclear chromatin represented the common trait of chronic type B and type C hepatitis examined by light microscopy. Ultrastructural patterns confirmed the lesions and demonstrated virus-resembling particles in the cell nuclei. In HCV infection, hepatocyte cytoplasm contained tubular and horseshoe-shaped structures with lesions of mitochondria, while in HBV infection Dane's particles and tubular forms of HBsAg were detected. For cognitive reasons and due to frequently equivocal literature data, our data on ultrastructural lesions in chronic type C hepatitis seem to be of particular interest. Using the ImmunoMax signal amplification, we were able to diagnose HBV infection under light microscope and to define stage of the infection. Their sensitivity, specificity and relatively short time required for performing the tests makes them advisable in the routine diagnosis of the two infections.

Detection of DNA, mRNA and early antigen of the human cytomegalovirus using the immunomax technique in autopsy material of children with intrauterine infection.

The present study focuses on the immunomax technique in association with the avidin-biotin-peroxidase complex (ABC) technique and a non-isotopic variation of in situ hybridisation (ISH) for optimal microscopical detection of human cytomegalovirus (HCMV). The studies were performed on an archival paraffin material originating from five children deceased due to intrauterine infection. The results of immunocytochemical and hybridocytochemical studies, with or without amplification using biotinylated tyramine, were compared with the routine histopathological results and results obtained using the polymerase chain reaction (PCR). Early antigen (EA)-HCMV was demonstrated in approximately twice as many cells as detected in the routine staining and also in cells that seemed morphologically intact. The hybridocytochemical studies confirmed the presence of HCMV DNA in cells that were positive in the immunocytochemical tests and, in addition (using the ISH-immunomax technique), in cell nuclei of intact myocardial myocytes. In general, fewer cells manifested the presence of HMCV mRNA than the presence of HCMV DNA. The immunomax technique was found to be more sensitive than the techniques of classical immunocytochemistry or of ISH. The former technique permitted the documentation of a higher number of HCMV replication sites than could be detected using the latter techniques. However, the clinical course of HCMV infection or the cause of death of the children was not directly related to the intensity of HCMV expression in tissues.

A comprehensive analysis of selected diagnostic methods with respect to their usefulness in evaluating the biology of neoplastic cells in patients with laryngeal cancer.

The difficult and complicated mechanism of cancer development with little knowledge about the biology of existing cancers can lead to a permanent search for new examination techniques to improve the precision of life expectancy in patients and the selection of the most efficient methods of treatment. The aim of this study was to analyze certain prognostic factors, i.e., p53, Ki67, proliferating cell nuclear antigen (PCNA), DNA ploidy and cell proliferating activity, as well as the degree of morphological differentiation and cell maturity evaluated on an ultrastructural level in patients with laryngeal cancers in connection with data obtained from follow-up examinations and the clinical course of the disease. Neoplastic tissue was taken from 120 patients with laryngeal cancers. All underwent surgical treatment, radiotherapy and combined treatment in the Department of Otolaryngology. Karol Marcinkowski University School of Medical Sciences, Poznan, Poland, and the Department of Otolaryngology-Head and Neck Surgery, Haukeland University, Bergen, Norway. Before beginning treatment all patients underwent histological verification of their neoplastic tissues. Histopathological examination proved that the majority of cases (95%) had a squamous cell carcinoma. The occurrence of changes within the lymph nodes of the neck (N) was significantly correlated with T, S, Ki67, metastases to lymph nodes, DNA ploidy, site and surgery performed. The degree of clinical progression (S) was intercorrelated with T, N, p53, Ki67, PCNA, DNA ploidy, site and laryngectomy. The occurrence of oncoprotein p53 in neoplastic cells was measured by the staining degree of their nuclei and was correlated with T, S, DNA ploidy, metastases to lymph nodes, PCNA and site. The degree of staining of neoplastic cells for the nuclear antigen Ki67 was correlated to T, N, G, S, DNA ploidy, metastases to lymph nodes and surgical treatment. The proliferative antigen PCNA in the examined population of patients was intercorrelated with T, p53, Ki67, metastases to lymph nodes and surgical treatment. The results obtained from DNA flow cytometry could be associated with N, G, p53, Ki67 and metastases to lymph nodes. On the basis of the results obtained, the techniques suggested for the morphological and biological evaluation of neoplastic cells in cancer of the larynx should include TNM classification + G + DNA + p53 + Ki67.

The experimental investigations of the toxic influence of tobacco smoke affecting progeny during pregnancy.

1. Tobacco smoke contains around 4000 substances, most of which are described as toxic, and they may have an influence on the development of progeny. 2. The present studies concentrate on the measurement and calculation of indices describing the new-born's survival, rearing of pups, weight of foetuses, young animals, placenta and females in relation to different doses of tobacco smoke (carbon monoxide levels). The morphological studies of placenta, foetal and newborn lungs were done as a supplement. Biochemical placenta study was also done. 3. The results of the experiment proved that some indices for animals in groups which were passively exposed to the highest concentrations of tobacco smoke were lower, others fluctuated (4 day, 12 day and total survival) and some did not reveal any changes (rearing). Direct correlation between maternal passive exposure to tobacco smoke and the presence of acute respiratory distress syndrome symptoms in new-borns was observed. A decrease of body weight of pregnant females passively exposed to tobacco smoke was also observed. An increase of placenta-foetal factor was found. A decrease of rat weight was observed after passive exposure to tobacco smoke. 4. We concluded that there is correlation between passive exposition to tobacco smoke during pregnancy and delayed lung maturation in the offspring. Exposure of the pregnant rats to cigarette smoke increases the activity of isocitric and glucose-6-phosphate dehydrogenases in placenta.

CD56(NCAM) antigen in glandular epithelium of human thyroid: light microscopic and ultrastructural study.

CD56 antigen, an isoform of the neural cell adhesion molecule (NCAM) was previously found by us in human thyroid by APAAP immunohistochemistry in light microscopy on frozen tissue sections. In the current study, it was attempted to trace the antigen in question using another light microscopic immunohistochemical procedure and to validate the results at the ultrastructural level. For light microscopy, cryostat sections of 12 surgical samples of human thyroid were subjected to ABC (preformed avidin-biotin-peroxidase complex) method. For immunoelectron microscopy, immunoperoxidase reaction was carried out on prefixed, small thyroid tissue blocks. Following preliminary inspection of semithin sections, ultrathin sections were examined in the transmission electron microscope. ABC reaction revealed distinct specific CD56 staining of thyrocyte cell membranes. The staining was weak or absent in thyroid papillary carcinoma cells. The results were confirmed in semithin sections by indirect immunoperoxidase. The latter reaction in ultrathin sections at the ultrastructural level has shown that specific reaction product was confined to free and lateral surfaces of thyroid follicular cells. Endothelial cell membranes of thyroid capillary vessels were totally devoid of the reaction product. The reaction was weakly positive in thyroid follicular and papilllary carcinomas but absent from medullary carcinoma.

Ultrastructural changes in the lung alveoli after cardiac surgical operations with the use of cardiopulmonary bypass (CPB).

Cardiac surgical procedures with the use of cardiopulmonary bypass (CPB) are commonly complicated by pulmonary dysfunction. The mechanisms of such injury are not well understood. The aim of the present study is to analyze morphologically (mainly ultrastructurally) alveolar injury, which occurred during cardiac surgical operations involving CPB, equipped with a hollow fiber oxygenator. Our study included 20 patients, aged 45-72, who underwent coronary artery bypass grafting. Lung biopsies were taken from the left upper lobe 20 minutes after stopping CPB. Pre-CPB biopsies served as controls. Tissue specimens used for electron microscopy were processed according to standard procedures. Light microscopy revealed only a few alterations in the terminal part of the respiratory tract. Frank edema was seen in some of the alveoli. Extravasated erythrocytes as well as some neutrophils were present in the alveoli and several alveolar capillaries were congested. Ultrastructural observations confirmed the above mentioned changes. Moreover, in many alveoli, extensive injury to air-blood barrier was observed. Type I pneumocytes and endothelial cells appeared swollen or necrotically changed. The cytoplasm of type II pneumocytes was swollen. In many alveoli, pulmonary surfactant could not distribute over the alveolar surface because of edema. Structures of pulmonary surfactant were also seen in alveolar capillaries. The results of this investigation suggest that CPB is associated with some injury to lung tissue. However, this injury seems to be temporary since all examined patients had an uneventful post-operative course.

Studies on tissue expression of HBV in children with chronic hepatitis type B using Immunomax technique.

The study aimed at employing the Immunomax technique to detect the markers of HBV replication (HBcAg and HBV-DNA) in liver biopsy material, obtained from children with chronic hepatitis type B. In line with the currently modified classification of chronic hepatitis and with the increasing potential of antiviral therapy it seemed purposeful to supplement routine staining techniques with studies at the molecular level. Our studies demonstrated the effective detection of both the core antigen and HBV-DNA in liver tissue in children using immunocytochemical techniques and in situ hybridization, amplified with the Immunomax technique. HBcAg was detected in 26 out of 27 liver biopsies from patients with chronic hepatitis type B and with replication of the virus. HBV-DNA was detected in all study children with HBV infection and in 2 out of 5 cases of chronic hepatitis of a distinct etiology. No significant relationships could be found between the detection of tissue HBV markers on the one hand and the intensity of inflammatory lesions or severity of fibrosis on the other.

Development of the fetal and newborn lung--morphometric studies (rat model).

The aim of this study was to investigate the development of the fetal lung by morphometric means. Lung specimens from 16-20 day-old fetuses, and 1, 3, 5, 14 and 21 day-old newborns were used. Tissue specimens were evaluated by light microscopy and transmission electron microscopy. The material was divided into four groups--related to four main stages of lung development (pseudoglandular, tubular, saccular and alveolar). For each developmental stage, 30 photographs of semi-thin sections were saved on computer hard disk for future morphometric measurements. OPTIMAS 4.02 software was used for evaluation of the relative area occupied by vascular, epithelial, stromal and fluid/gaseous compartments. GraphPad InStat software was used for statistical analysis. All measurements and calculations were done per observation field. The area occupied by vessels was greatest at the pseudoglandular stage (11.7%), and smallest at alveolar stage (5.7%). The stroma also occupied the largest area at the pseudoglandular stage (58.4%) and the smallest at alveolar stage (17.0%). The area occupied by epithelial cells was most extensive at the tubular stage (52.5%) and smallest at the alveolar stage (6.1%). The fluid/gas compartment was largest at the alveolar stage (70.5%) and smallest at the pseudoglandular stage (0.58%). The present results indicate that the most dynamic lung development occurs during thin air-blood barrier formation (19th day of intrauterine life to 5th day after delivery--saccular and the beginning of alveolar stage).

[The value of DNA flow cytometry in evaluation of proliferating activity of neoplasm cells in patients with laryngeal cancer]. / Wartosc cytometrii przeplywowej DNA w ocenie zdolnosci proliferacyjnej komórek nowotworowych u chorych na raka krtani.

A study of DNA ploidy was carried out including 84 cases of laryngeal squamous cell carcinoma. Material was fixed during 30 min in 3% paraformaldehyde and embedded in low-melting-point paraffin. The histologic studies were done for diagnosis and general view. Results were as follows. The highest percentage of cancers with DNA diploidy was found in well- and medium-differentiated tumors. Among well-differentiated carcinomas there were 18% diploid versus 5% polyploid tumors. In the medium-differentiated cancers there were 23.5% diploid, 6% polyploid, and 6% hyperdiploid tumors. The highest percentage of DNA hyperdiploidy and poliploido-aneuploidy was found in low-differentiated cancers. Additionally, tumor proliferative activity was evaluated in relation to the percentage of cells in phase S of cellular cycle. Special attention was paid to tumors which revealed the phenomenon of DNA poliploidy-aneuploidy beside DNA diploidy.