Assessment of in vitro particle dosimetry models at the single cell and particle level by scanning electron microscopy.

BACKGROUND: Particokinetic models are important to predict the effective cellular dose, which is key to understanding the interactions of particles with biological systems. For the reliable establishment of dose-response curves in, e.g., the field of pharmacology and toxicology, mostly the In vitro Sedimentation, Diffusion and Dosimetry (ISDD) and Distorted Grid (DG) models have been employed. Here, we used high resolution scanning electron microscopy to quantify deposited numbers of particles on cellular and intercellular surfaces and compare experimental findings with results predicted by the ISDD and DG models. RESULTS: Exposure of human lung epithelial A549 cells to various concentrations of differently sized silica particles (100, 200 and 500 nm) revealed a remarkably higher dose deposited on intercellular regions compared to cellular surfaces. The ISDD and DG models correctly predicted the areal densities of particles in the intercellular space when a high adsorption ("stickiness") to the surface was emulated. In contrast, the lower dose on cells was accurately inferred by the DG model in the case of "non-sticky" boundary conditions. Finally, the presence of cells seemed to enhance particle deposition, as aerial densities on cell-free substrates were clearly reduced. CONCLUSIONS: Our results further validate the use of particokinetic models but also demonstrate their limitations, specifically, with respect to the spatial distribution of particles on heterogeneous surfaces. Consideration of surface properties with respect to adhesion and desorption should advance modelling approaches to ultimately predict the cellular dose with higher precision.

Significance of the histological and ultrastructural features of elastic fibers in diagnosis of bronchioloalveolar carcinoma.

OBJECTIVES: The accurate identification of bronchioloalveolar carcinoma (BAC) from adenocarcinoma (AC) and other types of lung cancer is important from clinical perspectives; especially, when BAC is histologically-mixed with AC. We hypothesized that the elastic fibers (EF) pattern could be used as a differential marker to identify BAC from other lung cancers. The aim was to characterize the EF pattern in different types of lung cancer and evaluate its significance for differential diagnosis of BAC. PATIENTS AND METHODS: Clinical samples of different types of lung cancers were collected. The samples were stained by hematoxylin-eosin (H&E) staining for histopathological comparison. Then, modified Weigert's staining of the EF was performed to characterize its patterns. The EFs were semi-quantified and compared among different types of lung cancer. Further, transmission electronic microscopy (TEM) was performed and ultrastructural features of the EFs were compared between BAC and adenocarcinoma (AC). RESULTS: H&E staining histopathology could differentiate the most types of lung cancer except certain types, such as histologically-mixed BAC and AC. The EF pattern in BAC was uniquely different from other types of lung cancer as > 95% of BAC was + or ++ for EF staining while > 95% of other types of lung cancer were--or ± type. TEM study further confirmed the EF pattern difference between BAC and AC. CONCLUSIONS: As the data show, as > 95% BAC specimens can be identified from other lung cancers based on EF (Weigert's) staining. The EF pattern in BAC is uniquely different from other types of lung cancer and, therefore, can be used as a differential clinical marker of BAC.

Ultrastructural findings in metastatic bronchioloalveolar carcinoma.

This study was prompted by the recent revision of the definition of bronchioloalveolar carcinoma (BAC) that defines BAC, light microscopically, as a non-invasive carcinoma. Doubt has been raised whether BACs retain certain specific microscopic features after becoming invasive or metastatic. We studied 7 cases of metastatic, non-mucinous BAC by electron microscopy. Of these cases, 5 showed Clara cell granules and 1 revealed lamellar bodies. The remaining case did not show ultrastructural features of BAC. These findings suggest that most BACs retain some of their ultrastructural features after becoming metastatic neoplasms.

The concept of bronchioloalveolar cell adenocarcinoma: redefinition, a critique of the 1999 WHO classification, and an ultrastructural analysis of 155 cases.

Bronchioloalveolar cell adenocarcinoma (BACA) is bronchioloalveolar because (1) it arises in bronchioles and alveoli and (2) differentiates into bronchiolar and alveolar cells. Every entity possesses unique characteristics that separate it from other entities. The unique characteristic of BACA is its cell type. Lepidic growth is a clue to the cell type and, even though present in the vast majority, is not unique or absolutely essential. Because of the algebraic nature of concepts, the degree of differentiation, the extent of lepidic growth, and the degree of stromal desmoplasia cannot be used as definitional requirements. Likewise, in malignant tumors, absence of stromal invasion cannot be required. An epistemologically valid definition of BACA is proposed and a study of 155 cases defined this way and examined ultrastructurally is presented.

Establishment of an immortalized cell line and transplantable xenograft from a bronchioloalveolar lung carcinoma of a cat.

OBJECTIVE: To establish an immortalized cell line and transplantable xenograft of feline bronchioloalveolar lung carcinoma (BAC). SAMPLE POPULATION: Pleural effusion from a 12-year-old Persian male cat with BAC. PROCEDURE: Tumor cells from the pleural effusion were grown in monolayer cell culture and injected into severe combined immunodeficient (SCID) mice to establish an immortalized cell line as well as a transplantable xenograft. RESULTS: Both the primary lung carcinoma, the derived cell line, and the transplantable xenograft had evidence of a type-II pneumocyte origin expressing lamellar bodies ultrastructurally and thyroid transcription factor-1 and surfactant immunocytochemically. All 3 also expressed nuclear p53 immunoreactivity. A metaphase spread of the cell line (SPARKY) probed with fluorescein-labeled genomic feline DNA gave evidence of its feline origin. Flow cytometric studies indicated aneuploidy with a DNA index of 1.6. An R-banded karyotype revealed a modal number of 66 including the feline Y chromosome. The cell line had a doubling time of 16 hours. The xenograft (SPARKY-X) reached a diameter of 1 cm in 3 weeks in SCID mice. Deoxyribonucleic acid fingerprint analysis revealed that SPARKY and SPARKY-X were novel and strongly matched each other, except for the murine component found in SPARKY-X. Interestingly, SPARKY-X manifested the characteristic lepidic growth pattern of pulmonic BAC. CONCLUSIONS: Both the cell line and xenograft retained their autochthonous BAC phenotype, making them useful for the subsequent dissection of molecular abnormalities in feline BAC and in vitro screening of chemotherapeutic agents.

Ultrastructural comparison of "alveolar" and "solid" areas in bronchioloalveolar carcinoma.

The purpose of this study was to compare the ultrastructural features of bronchioloalveolar carcinomas, contrasting the well-differentiated alveolar component and the poorly-differentiated solid component in the same tumor. We studied 7 cases of non-mucinous bronchioloalveolar carcinomas by electron microscopy. Two of these cases showed lamellar bodies in both the alveolar and solid components and the remaining 5 cases revealed Clara cell granules in both components. We conclude that the neoplastic cells in bronchioloalveolar carcinoma retain their ultrastructural phenotypes after becoming invasive carcinoma with loss of alveolar differentiation.

Aerogenous spread of primary lung adenocarcinoma induces ultrastructural remodeling of the alveolar capillary endothelium.

To determine whether pulmonary alveolar capillaries manifest ultrastructural remodeling at areas of neoplastic invasion of primary lung adenocarcinomas, we examined 17 well-differentiated adenocarcinomas of lung (2 bronchioloalveolar and 15 papillary adenocarcinomas) by electron microscopy. The expression of vascular endothelial growth factor (VEGF) and proliferating cell nuclear antigen (PCNA) was demonstrated by immunohistochemical stainings. VEGF messenger RNA (mRNA) isoforms were detected by reverse-transcription polymerase chain reaction (RT-PCR) in alveolar walls microdissected from normal and tumor-associated tissues. Cytoplasm of neoplastic cells expressed VEGF protein in all patients. Endothelial cell nuclei of alveolar capillaries showed positive reaction for PCNA. Alveolar capillary lumina were distended like venules, and some intercellular junctions remained open. The cytoplasm of the capillary endothelial cells was enlarged and developed numerous organelles such as Weibel-Palade bodies and vesiculovacuolar organelles, in contrast to marked attenuation in their normal counterpart. Capillary sprouting occurred from proper alveolar capillaries in 2 patients. Cytoplasmic segments became extremely attenuated and developed diaphragm-like fenestrae in 65% of the patients. A relatively higher expression of diffusable isoforms of VEGF mRNA was seen in the tumor-bearing alveolar walls than in normal walls. Expression of KDR (one of the VEGF receptors) mRNA in tumor exceeded that in normal tissues. These results suggest that diffusable isoforms of VEGF mRNA released from the neoplastic cells are deeply involved in the induction of growth activity of alveolar capillary endothelial cells as much as in the characterization of tumor-associated microvessels in primary lung adenocarcinomas.

Ultraestructura del carcinoma bronquioalveolar de pulmón. Estudio de 17 casos / Bronchioloalveolar carcinoma of the ling. Report of 17 cases

Introducción: El carcinoma bronquioloalveolar (CBA) se divide en mucinoso, no mucínoso y mixto. En microscopia electrónica (ME) muestra células mucinosos (CM), de Clara (CC), ciliadas, neumocitos tipo II (PN) y células indeterminadas (C1) de significado incierto. Material y métodos: Comparamos la microscopia óptica con la ultraestructura en 17 casos de CBA. Resultados: Por microscopia óptica, 4 casos eran de tipo mucinoso, 11 no mucinosos y 2 mixtos. Por ME, los 4 casos mucinosos mostraban diferenciación mucinosa y los 2 mixtos una mezcla de CC y CM (uno con abundantes Cl y PN). De los 11 casos no mucinosos, 7 mostraban diferenciación hacia CC (4 de tipo cilíndrico y 3 cúbicas), 2 mucinosa, otro mixta y el último exclusivamente Cl y PN. Conclusiones: Las Cl se encuentran en relación con PN y tienen cierta similitud con las CC cúbicas, lo que sugiere que pudieran ser el origen de algunos CBA (AU)

Unique cellular features in atypical adenomatous hyperplasia of the lung: ultrastructural evidence of its cytodifferentiation.

Atypical adenomatous hyperplasia (AAH) of the lung could be a good material to understand the histogenesis of peripherally occurring, well-differentiated adenocarcinoma. However, its true biological significance remains to be clarified. The authors present the histomorphological studies of this lesion and compare the ultrastructure with that of nonmucinous bronchioloalveolar carcinoma (BAC) to define characteristic features of AAH. Light microscopy showed the well-preserved pulmonary architecture, proliferated neoplastic cells without marked cellular atypia, and no transitional area to obvious adenocarcinoma. Intranuclear inclusion was present in a large number of neoplastic cells. Electron microscopy revealed that cuboidal or low columnar neoplastic cells proliferated actively but were not crowded on slightly thickened fibrous alveolar septa with both Clara-like granules and small lamellar bodies in the cytoplasm resembling that of Clara cell and type 2 pneumocyte. Some of the nuclei had characteristic invaginations of its nuclear membrane. Although the findings appear to be nonspecific for AAH, the authors emphasize that AAH is an alveolar intraepithelial neoplasia that represents a very early stage in the continuous developmental spectrum of adenomatous neoplasia in the bronchioloalveolar region corresponding to dysplasia or intraepithelial neoplasia in other organs, and will give the significance to speculate its histogenesis.

Analysis of cell type and radiographic presentation as predictors of the clinical course of patients with bronchioalveolar cell carcinoma.

STUDY OBJECTIVES: Bronchioloalveolar carcinoma is a primary lung neoplasm of variable histopathologic, radiologic, and clinical expression. There are three cell types described in bronchioloalveolar carcinoma: Clara cells, mucin-producing cells, and alveolar type II epithelial cells. It is unclear whether these three tumor cell types are associated with a specific radiologic presentation and clinical course. In this study, we investigated whether tumor cell type, identified by transmission electron microscopy, correlated with a specific radiologic pattern, and whether tumor cell type or radiologic presentation correlated with the patient's clinical course and outcome. DESIGN: Transmission electron microscopy was used to restudy tissue blocks from the original surgical histopathologic specimens in 54 patients with primary bronchioloalveolar carcinoma diagnosed over a 10-year period (1980 to 1990). The pretreatment radiographs were reviewed in each case, and the first chest radiograph obtained at the time of the discovery of the tumor in each patient was compared with the results of the ultrastructural study. The medical records of each patient were examined to obtain pertinent radiologic, clinical, and patient outcome information. MEASUREMENT AND RESULTS: There were 32 Clara cell tumors, 10 mucin-producing cell tumors, and 1 alveolar type II epithelial cell tumor in this series. Eleven additional tumors had mixtures of two or more cell types. No statistically significant relationship was detected between tumor cell type and radiologic presentation or patient mortality pattern. There was increased mortality among patients who presented radiologically with segmental, lobar, multifocal, or diffuse disease compared with those patients exhibiting a solitary pulmonary nodule at presentation. CONCLUSION: Radiologic presentation, rather than tumor cell type, provides prognostic information that aids in predicting patient outcome.

Bronchiolar-alveolar carcinoma in a cow.

Bronchiolar-alveolar carcinoma was observed in the lung of an 8-year-old Holstein cow. Grossly, the lung contained multiple tumour masses, which were solid, yellowish-white in colour, and firm in consistency. These tumours also occurred in the liver, pancreas, uterus and lymph nodes in the thoracic, abdominal and pelvic cavities. Histologically, the masses were composed of abundant fibrous stroma and proliferating atypical cuboidal epithelial cells, occasionally forming glandular structures. Electron microscopy revealed that the neoplastic cells had microvilli and lamellar bodies in the cytoplasm, suggesting that they originated from immature respiratory epithelial cells differentiating towards either Clara cells or type II pneumocytes.

Cytodifferentiation of atypical adenomatous hyperplasia and bronchioloalveolar lung carcinoma: immunohistochemical and ultrastructural studies.

We used immunohistochemistry and electron microscopy to evaluate the differentiation of cells comprising atypical adenomatous hyperplasia (AAH; n = 26), early bronchioloalveolar lung carcinoma (BAC; n = 11), and overt BAC (n = 16), which are assumed to constitute a continuous spectrum of developmental steps of BAC. Surfactant apoprotein (SAP), a marker for type 2 alveolar cells, was expressed in cells from all the lesions of AAH, early BAC, and overt BAC. However, the proportion of SAP-positive cells decreased and their distribution became more heterogeneous with advancing lesion grade. Urine protein 1, which is identical to the Clara cell-specific 10 kDa protein, was expressed in 70% of overt BAC, whereas only 20% of early BAC showed weak reactivity and none of AAH lesions showed any reactivity at all. Ultrastructurally, type 2 alveolar cell differentiation was predominant among cells from AAH and early BAC. Our results suggest that precursor cells of BAC differentiate predominantly towards type 2 alveolar cells. Cells comprising overt BAC retain this differentiation phenotype, but to a reduced extent. In contrast, concomitantly with progression, cells with Clara cell differentiation emerge and their proportion increases. Such phenotypic changes may reflect metaplasia occurring in tumour cells during the development of BAC.

[Bronchiolo-alveolar carcinoma]. Histologic analysis of 3 cases, one of which with ultrastructural study]. / Carcinoma bronchiolo-alveolare. Analisi istologica di 3 casi uno dei quali con esame ultrastrutturale.

A histological analysis was performed on 3 cases of bronchiolo-alveolar carcinoma (BAC) in one of which an electronmicroscopic research was made. It was about a case of BAC with pneumonocytes of type I the lasting in life of which from the ascertainment of the tumor was 1.2 years (autopsy); and of a case of BAC with pneumocytes of type II (operatory case) and of a case of BAC with conventional cells (autoptic case) who survived an average of 4.3 years. All the cases had metastasis in the ilar lymph nodes. The three BAC were localized in the superficial area of the lung and it is also evident that they occupy the alveolar cavities, taking the thin alveolar wall as the only interstice. Besides in the following development there was evident tendency to hypertrophy of the interstizial collagen tissue, in the BAC with pneumocytes of type II and respectively with cells of conventional type; and the subtotal PAS negativity in the BAC with pneumocytes of type I. The included nuclears present in the only BAC with pneumocytes of the second type take the origin from the cytoplasm in the shape of small round masses penetrating into the nucleus, as from electronic microscopy. In the considered BAC, it was about surfactant apoproteins wich, in the nucleus, precipitate in morphologically different shapes.

Alveolar cell carcinoma presenting as malignant pericardial effusion: diagnosis by electron microscopy.

The cytologic, immunocytochemical, and ultrastructural findings in a 68-year-old man who presented with a malignant pericardial effusion are reported. Radiologic studies failed to identify a primary neoplasm over the next 6 months. Ultrastructural examination of a repeat pericardiocentesis fluid specimen revealed cells with intranuclear tubular inclusions and cytoplasmic lamellar bodies typical of alveolar cell carcinoma. Review of the chest radiographs showed nonresolving patchy infiltrates in the upper lobe of the left lung clinically thought to represent pneumonia; in retrospect, however, these were consistent with the pneumonic form of alveolar cell carcinoma.

[An electron microscopic study of the behavioral characteristics of human lung tumor cells cultured on soft agar]. / Elektronno-mikroskopicheskoe issledovanie osobennostei povedeniia kletok opukholei legkogo cheloveka, kul'tiviruemykh v miagkom agare.

The ultrastructure of cells from seven human lung cancers and from the colonies formed by these cells in soft agar was investigated. Tumor cells developed to display the morphofunctional potentials of the initial tumors. Cultured cells of squamous-cell carcinomas contained numerous tonofilaments, those of adenocarcinomas developed microvilli on their apical surfaces and intracellular lumens. On the other hand, cells of squamous-cell carcinomas showed features specific of adenoma epithelium, i.e. well developed microvilli and intracellular lumens. Besides, cells of adenocarcinoma often contained large quantities of tonofilaments considered to be characteristic of epidermoid epithelium. The results obtained suggest a possibility of metaplastic transformation of the lung epithelium.

Bronchioloalveolar adenocarcinoma with myoepithelial cells.

A unique variant of papillary lung carcinoma in a 58-year-old woman with both Clara cell and myoepithelial components is reported. The tumor was characterized by the presence of glandular spaces lined by two cell layers. The superficial (luminal) layer was made up of columnar cells with ultrastructural and immunohistochemical features of Clara cells, and occasional interspersed type 2 pneumocytes. The cells of the basal layer possessed ultrastructural and immunohistochemical features of myoepithelial cells. Myoepithelial cells have not previously been reported in a bronchioloalveolar adenocarcinoma.

[The ultrastructural characteristics of different histological types of bronchiolar-alveolar cancer]. / Ul'trastrukturnye osobennosti razlichnykh gistologicheskikh tipov bronkhiolo-al'veoliarnogo raka.

Morphological study of bronchioloalveolar carcinoma (BAC) was performed by light or electron microscopy in 100 and 33 patients, respectively. BAC-1, 2 and 3 histologic subtypes were established basing on ultrastructural peculiarities which betrayed different cytogenetic origins. Electron microscopic signs of goblet cells were found in cells constituting BAC-1 tumors. BAC-2 tumor cells exhibited ultrastructural characteristics of Clara cells and/or type-II pneumocytes. BAC-3 tumors appeared less differentiated and contained mainly undifferentiated cells and cell clusters displaying various combinations of electron microscopic markers of goblet and Clara cells and type-II pneumocytes.