Thioredoxin, a gene found overexpressed in human cancer, inhibits apoptosis in vitro and in vivo.

The redox protein thioredoxin plays an important role in controlling cancer cell growth through regulation of DNA synthesis and transcription factor activity. Thioredoxin is overexpressed by a number of human primary cancers and its expression is decreased during dexamethasone-induced apoptosis of mouse WEHI7.2 thymoma cells. We examined the ability of WEHI7.2 cells stably transfected with human thioredoxin cDNA showing increased levels of cytoplasmic thioredoxin to undergo apoptosis in vitro and in vivo. The cells were protected from apoptosis induced by dexamethasone, staurosporine, etoposide, and thapsigargin, but not by N-acetyl-sphingosine. When inoculated into severe combined immunodeficient mice, the trx-transfected cells formed tumors that showed increased growth compared to wild-type, as well as bcl-2-transfected, WEHI7.2 cells. The trx- and bcl-2-transfected cell tumors both showed less spontaneous apoptosis than tumors formed by the wild-type cells. Unlike tumors formed by the wild-type and bcl-2-transfected WEHI7.2 cells, trx-transfected cell tumors did not show growth inhibition upon treatment with dexamethasone. This study suggests that increased thioredoxin expression in human cancers may result in an increased tumor growth through inhibition of spontaneous apoptosis and a decrease in the sensitivity of the tumor to drug-induced apoptosis.

Catalase-overexpressing thymocytes are resistant to glucocorticoid-induced apoptosis and exhibit increased net tumor growth.

Glucocorticoids are used for the treatment of lymphoid neoplasms, taking advantage of the well-known ability of these compounds to cause apoptosis in lymphoid tissues. Previously, we have shown that dexamethasone, a synthetic glucocorticoid, causes a down-regulation of several antioxidant defense enzymes and proteins, including catalase and thioredoxin, concomitant with the induction of apoptosis in WEHI7.2 mouse thymoma cells. To test whether this down-regulation plays a critical role in the mechanism of steroid-induced apoptosis, WEHI7.2 cells were transfected with rat catalase. Two clones, expressing 1.4-fold and 2.0-fold higher catalase specific activity, respectively, when compared with vectoronly transfectants were selected for further study. An increase to 1.4-fold parental cell catalase activity delayed cell loss after dexamethasone treatment, whereas a 2.0-fold parental catalase activity prevented dexamethasone-induced cell loss for 48 h after treatment. Dexamethasone treatment of the WEHI7.2 cells stimulated a release of cytochrome c into the cytosol. Catalase-overexpressing cells showed a delay or lack of cytochrome c release from the mitochondria, which correlated temporally with the delay or prevention of cell loss in the culture after dexamethasone treatment. A decreased amount of cell death from WEHI7.2 cells overexpressing catalase was also seen in tumor xenografts in severe combined immunodeficient mice when compared with tumors from vector-only transfected cells. Similarly, thioredoxin-overexpressing WEHI7.2 cells, shown previously to be apoptosis resistant, showed decreased cell death in tumor xenografts. This resulted in larger tumors from cells overexpressing these proteins. Cell death in control transfectant tumor xenografts was primarily attributable to apoptosis. In contrast, the cell death we observed in tumors from thioredoxin- or catalase-overexpressing cells had a higher frequency of a nonapoptotic, nonnecrotic type of cell death termed para-apoptosis. These data suggest that: (a) oxidative stress plays a critical role in steroid-induced apoptosis prior to the commitment of the cells to undergo apoptosis; and (b) resistance to oxidative stress can contribute to tumor growth.

The NAD+ precursors, nicotinic acid and nicotinamide protect cells against apoptosis induced by a multiple stress inducer, deoxycholate.

The bile salt, sodium deoxycholate (NaDOC), is a natural detergent that promotes digestion of fats. At high physiologic levels, NaDOC activates many stress-response pathways and induces apoptosis in various cell types. NaDOC induces DNA damage and activates poly(ADP-ribose) polymerase (PARP), an enzyme that utilizes NAD+ as a substrate to repair DNA. NaDOC also induces oxidative stress, endoplasmic reticulum (ER) stress and contributes to protein malfolding. The NAD+ precursors, nicotinic acid (NA) and nicotinamide (NAM) were found to protect cells against NaDOC-induced apoptosis. NA and NAM also decreased constitutive levels of both activated NF-kappaB and GRP78, two proteins that respond to oxidative stress. However, the mechanism by which NA and NAM protects cells against apoptosis does not involve a reduction in constitutive levels of oxidative stress. NA or NAM treatment increased the protein levels of glyceraldehyde-3-phosphate dehydrogense (GAPDH), a multi-functional enzyme, in the nucleus and cytoplasm, respectively. NAM did not activate the promoter/response elements of 13 stress response genes nor reduce intracellular non-protein thiols, suggesting that it is non-toxic to cells. NAM thus has promise as a dietary supplement to help prevent disorders involving excessive apoptosis.

Bile acid activation of the gadd153 promoter and of p53-independent apoptosis: relevance to colon cancer.

Bile acids are strongly implicated in the etiology of colon cancer. Bile acids also induce apoptosis, and this action may be a key to understanding their role in colon cancer. However the mechanism of bile acid induction of apoptosis is not known. We present evidence of bile acid activation of the gadd153 promoter (a promoter activated by DNA damaging agents). We also show that bile acid induction of apoptosis is p53-independent. In addition, bile salts were found to induce blebbing preceding the actual morphological onset of apoptosis, which indicates early cytoskeletal alterations.

The stress-response proteins poly(ADP-ribose) polymerase and NF-kappaB protect against bile salt-induced apoptosis.

Bile salts induce apoptosis and are implicated as promoters of colon cancer. The mechanisms by which bile salts produce these effects are poorly understood. We report that the cytotoxic bile salt, sodium deoxycholate (NaDOC), activates the key stress response proteins, NF-kappaB and poly(ADP-ribose) polymerase (PARP). The activation of NF-kappaB and PARP, respectively, indicates that bile salts induce oxidative stress and DNA damage. The pre-treatment of cells with specific inhibitors of these proteins [pyrrolidine dithiocarbamate (NF-kappaB inhibitor) and 3-aminobenzamide (PARP inhibitor)] sensitizes cells to the induction of apoptosis by NaDOC, indicating that these stress response pathways are protective in nature. Colon cancer risk has been reported to be associated with resistance to apoptosis. We found an increase in activated NF-kappaB at the base of human colon crypts that exhibit apoptosis resistance. This provides a link between an increased stress response and colon cancer risk. The implications of these findings with respect to apoptosis and to colon carcinogenesis are discussed.

Bile acids as carcinogens in human gastrointestinal cancers.

Bile acids were first proposed to be carcinogens in 1939 and 1940. On the basis of later work with rodent models, bile acids came to be regarded as cancer promoters rather than carcinogens. However, considerable indirect evidence, obtained more recently, supports the view that bile acids are carcinogens in humans. At least 15 reports, from 1980 through 2003, indicate that bile acids cause DNA damage. The mechanism is probably indirect, involving induction of oxidative stress and production of reactive oxygen species that then damage DNA. Repeated DNA damage likely increases the mutation rate, including the mutation rate of tumor suppressor genes and oncogenes. Additional reports, from 1994 through 2002, indicate that bile acids, at the increased concentrations accompanying a high fat diet, induce frequent apoptosis. Those cells within the exposed population with reduced apoptosis capability tend to survive and selectively proliferate. That bile acids cause DNA damage and may select for apoptosis-resistant cells (both leading to increased mutation), indicates that bile acids are likely carcinogens. In humans, an increased incidence of cancer of the laryngopharyngeal tract, esophagus, stomach, pancreas, the small intestine (near the Ampulla of Vater) and the colon are associated with high levels of bile acids. The much larger number of cell generations in the colonic (and, likely, other gastrointestinal) epithelia of humans compared to rodents may allow time for induction and selection of mutations leading to cancer in humans, although not in rodents.

Change in the frequency of apoptosis after low- and high-dose X-irradiation of human lymphocytes.

The purpose of the present study was to correlate the type and frequency of cell death in human lymphocytes receiving variable doses of X-irradiation. Monocyte-depleted lymphocyte fractions were exposed in vitro to variable doses of X-rays of 0-20 Gy (0-2000 rads) and incubated for 4 and 16 h. An assessment of the mode of cell death (apoptosis vs. classical necrosis) was carefully evaluated using a multidisciplinary approach using light, fluorescence and electron microscopy (EM), and dye exclusion assays. Eosin Y exclusion assays indicated the absence of classical necrosis occurring in short-term cultures (4 h postirradiation). An assessment of cell counts, however, revealed a mean decrease of 4% at 0 Gy and 13% at 10-20 Gy (1000-2000 rads). The predominant mode of cell death was apoptosis, but the percent apoptotic cells (determined by EM) did not parallel this increase in cell loss with increasing radiation and actually decreased at doses above 5 Gy (500 rads). The discrepancy between percent cell loss and percent apoptosis was explained by a proposed change in overall duration of the apoptotic process. In long-term cultures (16 h postirradiation), a combination of classical necrosis, classical apoptosis, and combined apoptosis and necrosis (secondary necrosis of apoptotic cells) was apparent and was associated with a marked decrease in viability. Irradiation effects on lymphocytes showing none of the morphologic features of apoptosis or classical necrosis in short-term culture were evidenced by an increase in nuclear lobation. The results of this study indicate that the vast majority of peripheral blood lymphocytes are radioresistant. The use of irradiation in an in vitro model to study the biochemical events of the apoptotic process is also evaluated.

The hyfrecator: a treatment for radiation induced telangiectasia in breast cancer patients.

Radiation induced telangiectasia is a common problem in breast cancer survivors. By interfering with choice of clothing and acting as an unpleasant visible reminder of their disease, it negatively affects quality of life. The hyfrecator, based on the principles of electrosurgery, is a standard treatment modality for facial telangiectasia. This study aims to demonstrate the efficacy and tolerability of the hyfrecator as a treatment for radiation induced telangiectasia. Patients with radiation induced telangiectasia of breast or chest wall were prospectively identified from the breast cancer follow up clinic and offered treatment with the hyfrecator (sessions at 8 weekly intervals). Pre- and post-treatment photographs were obtained in a standardized manner and two blinded physician observers evaluated response. A linear analogue scale (LAS) was used by the patients to evaluate treatment response and any discomfort. At the end of treatment, patients completed a quality of life questionnaire. Of 16 patients enrolled, 15 completed the study. Treatment benefited all patients with severe or marked telangiectasia. Complete disappearance of telangiectasia was achieved in the majority (88%) of patients by the end of treatment. A median of six sessions (range 3-9) was required. All but one (93%) considered the treatment worthwhile. The majority (69%) judged the treatment to be painless or only mildly painful. 73% reported an improvement in self-confidence. The treatment was well tolerated by all the patients. All patients showed a remarkable clearance of vessels with a high degree of satisfaction with the results. Treatment with the hyfrecator is very effective for radiation induced telangiectasia. Three to four sessions achieve a substantial objective and subjective reduction in telangiectasia with a concomitant improvement in quality of life. It is a cost effective, ambulant out patient procedure requiring no local anaesthesia.

Morphological evidence of an altered bone marrow microenvironment in patients with acute nonlymphoblastic leukemia and myelodysplastic disorders.

Type IV nuclear bodies are classified as true intranuclear inclusions that ultrastructurally contain numerous densely packed 20- to 30-nm osmiophilic granules surrounded by a microfibrillar cortex. In the present study, we have found statistically significant ultrastructural differences in the frequency and size of type IV nuclear bodies in the in vitro bone marrow fibroblastic cells (FC) derived from eight nonleukemic subjects and 13 patients with acute nonlymphoblastic leukemia (ANLL) and myelodysplastic disorders (MDD). Patients with ANLL, MDD, and myelofibrosis, as a group, had four times as many type IV nuclear bodies as nonleukemic subjects. The mean frequency of type IV nuclear bodies for patients with ANLL and MDD was 2.68% +/- 3.27% as compared with 0.63% +/- 1.06% for the nonleukemic subjects (p less than 0.05). The mean maximum type IV nuclear body area of the ANLL and chronic myelomonocytic leukemia (CMML) patients as a group was 2.08 +/- 1.10 micron2, compared with a mean maximum area of 0.93 +/- 0.10 micron2 from nonleukemic subjects (p less than 0.05). The FC were otherwise morphologically indistinguishable and displayed the typical ultrastructural features of fibroblasts. These findings have provided the first morphologic evidence that supports the concept of an altered bone marrow microenvironment in patients with ANLL and MDD. Since type IV nuclear bodies are found in high frequency in virally infected tissues, our quantitative ultrastructural findings raise the possibility of a local viral infection that affects the bone marrow microenvironment of patients with ANLL and some MDD disorders.

The specific NOS2 inhibitor, 1400W, sensitizes HepG2 cells to genotoxic, oxidative, xenobiotic, and endoplasmic reticulum stresses.

We tested the hypothesis that the constitutive activity of the inducible form of nitric oxide synthase (NOS2) serves to protect cells against numerous endogenous stresses. To accomplish this, we treated HepG2 cell lines that were individually transfected with 13 different promoter/response element (RE) chloramphenicol acetyl transferase (CAT) reporter constructs, with a highly selective NOS2 inhibitor, 1400W [N-(3-(aminomethyl)benzyl) acetamidine)]. HepG2 cells were incubated for 6 h with 0, 1, 10, 50, 100, and 200 microM 1400W, and the activation of the promoter/RE CAT reporter constructs was simultaneously determined. The highest fold inductions occurred at 200 microM 1400W, a concentration that had no effect on overall cell viability, as determined by the MTT assay. Twelve of the 13 promoter/RE CAT reporter constructs were significantly activated by 200 microM 1400W. These results indicate the extensive protective role of constitutive NOS2 against genotoxic, oxidative, and endoplasmic reticulum stresses. The mechanism of this protection may involve the complexing of iron by nitric oxide (NO) to reduce hydroxyl radical formation, NO inhibition of electron transport and the generation of reactive oxygen species within mitochondria, NO inhibition of cyclooxygenase, lipoxygenase, and cytochrome P450 enzyme activity, and the scavenging of superoxide anions by NO to form peroxynitrite.

Reactive nitrogen species in colon carcinogenesis.

The role of reactive nitrogen species (RNS) in colon carcinogenesis is multifactorial and affects diverse processes, such as proliferation, apoptosis, differentiation, tumorigenesis, and metastases. This review describes the stages in colon carcinogenesis where nitric oxide (NO) and inducible NO synthase (NOS2) may influence the progression of a normal mucosa to overt metastatic cancer. Overexpression of NOS2 and an increase in the generation of NO and other RNS may lead to apoptosis resistance, DNA damage, mutation, up-regulation of COX-2, increased proliferation, an increase in oxidative stress and an increase in tumor vascularity and metastatic potential. Therefore, future goals are to establish mechanistically based biomarkers to assess individuals at risk for colon cancer and to implement chemopreventive and dietary strategies that reduce colon cancer risk. An understanding of NO signaling pathways in colon epithelial cells should provide the basis for novel biomarker development. Colon cancer prevention may be achieved effectively by chemically interfering with key components of the NO signaling pathways, changing dietary habits to reduce fat and increase antioxidant-containing vegetables, and dietary supplementation to increase DNA repair.

Myopathy associated with linear scleroderma. A histochemical and electron microscopic study.

Pathologic findings in biopsied rectus femoris muscle underlying an area of linear scleroderma are described. The affected muscle was weak and atrophic. On electromyography it showed motor unit potentials of decreased amplitude and duration. Light microscopic changes were minimal. There was atrophy of some histochemical type I fibers. More prominent changes were found at the ultrastructural level, where many of the capillary basal laminae were thickened and reduplicated. Most striking was the presence of two types of electron-dense, rounded inclusions within the mitochondria, one ranging from 29 to 47nm in diameter and the other from 54 to 131 nm in diameter.

New ultrastructural observations: parallel tubular arrays in human T gamma lymphoid cells.

T gamma cells are E-rosetting cells bearing Fc receptors for IgG (E+, Fc gamma + cells). Third population (non-T, non-B) lymphoid cells are also Fc gamma + cells and contain unique inclusions called parallel tubular arrays (PTA). Although T gamma cells and third population lymphoid cells should belong to a similar population of cells, previous ultrastructural studies on purified T gamma cells have failed to reveal the presence of PTA. In this study, we have unequivocally demonstrated PTA in the majority of T gamma cells using simple rosetting techniques. A total of 76 EA hu-rosettes and 108 EA ox-rosettes prepared from an E+ enriched fraction (using sheep erythrocytes as marker particles) were directly examined by electron microscopy. PTA were found in 87% of the EA hu-rosettes and 82% of the EA ox-rosettes. Ammonium chloride, commonly used in other laboratories to lyse erythrocytes during the purification procedure was found to cause a marked decrease in the number of ultrastructurally distinct PTA profiles. In contrast, hypotonic lysis had no effect on cellular ultrastructure. This study showed for the first time that T gamma cells are ultrastructurally similar to other Fc gamma + lymphoid cells and contain PTA as a distinct marker. The significance of our findings to the basic function of this E+ Fc gamma + lymphoid population is discussed.

Signet-ring cell lymphoma of T-cell origin. An immunocytochemical and ultrastructural study relating giant vacuole formation to cytoplasmic sequestration of surface membrane.

In contrast to previous accounts of signet-ring lymphoma as a B-cell neoplasm, we report a case of signet-ring, large-cell lymphoma of T-cell lineage. Immunologic and ultrastructural studies were performed on a subcutaneous mass noted initially, as well as on an enlarged lymph node that developed later, in a 69-year-old man. Immunologic assessment indicated strong expression of T-helper antigen (Leu 3a + b), universal T-antigens (Leu 1, 5), and Ia. There was an absence of T-suppressor/cytotoxic antigen (Leu 2a), universal T-antigens (Leu 4, 9), and immunoglobulin light and heavy chains. Collectively, these findings indicate a mature T-cell lymphoma of T-helper type in an activated (Ia+) state. In contrast to previous reports of T-cell and Ia occurring solely as surface antigens, we demonstrated pools of cytoplasmic Leu 1, 3, 5 and Ia that displaced the nucleus. The ultrastructure of the giant cytoplasmic vacuoles was identical to the microvesicle-containing vacuoles reported in signet-ring cell lymphomas of B-cell lineage. In our case of T-cell lineage, we found substantial evidence of endocytosis by the neoplastic cells and numerous giant multivesicular bodies. The pools of cytoplasmic T and Ia antigens may result from abnormal internalization of surface T-antigens or the sequestration of T-antigen-containing Golgi-derived vesicles. Our combined immunologic and ultrastructural findings suggest that aberrant membrane recycling may be the common denominator of signet-ring formation in both B- and T-cell signet-ring lymphomas.

Eccrine and squamous differentiation in Merkel cell carcinoma. An immunohistochemical study.

Of the 42 Merkel cell carcinomas that we studied, two showed numerous tubular structures within sheets and nests of small cells. The small cells stained for both neuron-specific enolase and keratin. The keratin decorated a dot-like paranuclear structure. The ducts stained positively for carcinoembryonic antigen (CEA) and CF-1 (cystic fibrosis-1, a monoclonal antibody that only stains eccrine duct and acrosyringium). Electron microscopy performed on one case showed cytoplasmic dense-core neurosecretory granules and intercellular lumina lined by cells containing microvilli. These ultrastructural and immunohistochemical features support the concept of eccrine differentiation in these tumors. A third case contained foci of typical keratinizing squamous cell carcinoma admixed with sheets of small cells. The immunohistochemical and ultrastructural characteristics of this tumor were essentially similar to those of a conventional Merkel cell carcinoma. Our findings suggest that Merkel cell carcinomas, similar to neuroendocrine tumors from other anatomic sites arise from a primitive totipotential stem cell that has the capacity to differentiate along different cell lines.

Clinical and electron microscopic observations of the conjunctiva of adult patients with Bitot's spots.

Three adult patients with Bitot's spots of the conjunctiva were observed in the American Southwest; all were in good health without obvious cause for vitamin A deficiency. Serum vitamin A levels were low normal in two; the third patient was on replacement vitamin A therapy with resultant high serum levels. Serum beta-carotene levels were low in two and normal in one. Schirmer test without anesthesia was low normal in two and with topical anesthesia was abnormal in all. After 8 weeks of oral vitamin A therapy, the Schirmer test and Bitot's spots showed little response. Electron microscopy of Bitot's spots showed changes characteristic of keratinizing squamous epithelium: absence of goblet cells, increased tonofibrils, flattening of intermediate cells, loss of superficial cell nuclei, and a keratin layer. Light microscopy of the inferior cul-de-sac conjunctiva showed increased surface goblet cells in two and absence of such cells in the third; by electron microscopy the substructure of the majority of the goblet cell mucin granules had a reticulated appearance in which an electron-dense fibrillar network was present within the granule matrix. The non-mucin-containing epithelial cells of the inferior cul-de-sac for the most part appeared normal.

Inhibitory effect of selenomethionine on the growth of three selected human tumor cell lines.

Selenium supplementation has been shown for many years to work as an anticarcinogenic agent both in epidemiology and in in vitro studies. Selenium supplementation has recently been shown to decrease total cancer incidence. However, the mechanism of action of selenium as an anticarcinogenic agent has yet to be elucidated. Selenomethionine was the predominant form of selenium in the dietary supplement in the study by Clark et al. (Clark, L.C., Combs, G.F., Turnbull, W.B., Slate, E.H., Chalker, D.K., Chow, J., Davis, L.S., Glover, R.A., Graham, G.F., Gross, E.G., Krongrad, A., Lesher, J.L., Park, H.K., Sanders, B.B., Smith, C.L., Taylor, J.R. and The Nutritional Prevention of Cancer Study Group (1996) Effects of selenium supplementation for cancer prevention in patients with carcinoma of the skin: a randomized controlled trial. J. Am. Med. Assoc., 276 (24), 1957-1963) and therefore we evaluated the growth inhibitory effects of selenomethionine against human tumor cells. Selenomethionine was tested against each of three human tumor cell lines (MCF-7/S breast carcinoma, DU-145 prostate cancer cells and UACC-375 melanoma) and against normal human diploid fibroblasts. All cell lines demonstrated a dose-dependent manner of growth inhibition by selenomethionine. Selenomethionine inhibited the growth of all of the human tumor cell lines in the micromolar (microM) range (ranging from 45 to 130 microM) while growth inhibition of normal diploid fibroblasts required 1 mM selenomethionine, approximately 1000-fold higher than for the cancer cell lines. In short, normal diploid fibroblasts were less sensitive than the cancer cell lines to the growth inhibitory effects of selenomethionine. Furthermore, we show that selenomethionine administration to these cancer cell lines results in apoptotic cell death and aberrant mitoses. These results demonstrate the differential sensitivity of tumor cells and normal cells to selenomethionine.

Induction of oxidative stress and apoptosis in myeloma cells by the aziridine-containing agent imexon.

Imexon is an iminopyrrolidone derivative that has selective antitumor activity in multiple myeloma. The exact mechanism of imexon action is unknown. In human 8226 myeloma cells, the cytotoxicity of imexon was schedule-dependent, and long exposures (> or = 48 hr) to low concentrations of imexon were most effective at inducing cytotoxicity. Our data suggest that imexon does not affect DNA, but it can alkylate thiols by binding to the sulfhydryl group. We have also demonstrated by HPLC studies that in human 8226 myeloma cells, imexon depletes cellular stores of cysteine and glutathione. Oxidative stress in 8226 cells exposed to imexon was detected by immunohistochemical staining with a monoclonal antibody to 8-hydroxydeoxyguanosine (8-OHdG), followed by confocal microscopy. These images showed increased levels of 8-OHdG in the cytoplasm of cells treated with different concentrations of imexon at 8, 16, and 48 hr. Interestingly, 8-OHdG staining was not observed in the nuclei of imexon-treated cells, in contrast to the diffuse staining seen with t-butyl hydroperoxide. Myeloma cells exposed to imexon showed classic morphologic features of apoptosis upon electron microscopy, and increased levels of phosphatidylserine exposure, detected as Annexin-V binding, on the cell surface. To prevent depletion of thiols, 8226 myeloma cells exposed to imexon were treated with N-acetylcysteine (NAC). Simultaneous, as well as sequential, treatment with NAC before imexon exposure resulted in protection of myeloma cells against imexon-induced cytotoxicity. Conversely, the glutathione synthesis inhibitor buthionine sulfoximine increased imexon cytotoxicity. These data suggest that imexon perturbs cellular thiols and induces oxidative stress leading to apoptosis in human myeloma cells.

Histochemical and ultrastructural features of an unusual pulmonary carcinosarcoma.

An autopsy case is presented in which a pulmonary carcinosarcoma filled the left chest of a 61 year old man. The extensive pleural involvement that this neoplasm exhibited has not been reported previously. By light microscopy the neoplasm initially was considered a mesothelioma because of the pattern of glands and undifferentiated sarcomatous stroma. However, by electron microscopy the sarcomatous component was found to show rhabdomyoblastic differentiation. Neither histochemical stains nor electron microscopy supported a mesothelial origin for the glandular component. Differential diagnostic considerations of pleuropulmonary neoplasms showing rhabdomyosarcomatous differentiation are discussed. This case illustrates the importance of detailed study in order to characterize and properly classify these neoplasms.