Modulation of anti-tumour immunity and the effect of bacterial endotoxin on the growth of different syngeneic tumours from small inocula in mice

Studies were undertaken to determine the influence of E. coli lipopolysaccharide (LPS) on the growth of various doses of two antigenically-distinct syngeneic murine fibrosarcomas designated H1 and H7. The 'weakly' antigenic H1 tumour injected subcutaneously (s.c.) along the abdominal wall was profoundly susceptible to the growth-potentiating effects of a single intraperitoneal (i.p.) injection of 2 micrograms LPS, administered concurrently. 'Sneaking through' effects in control mice were observed with doses of 10 and 100 H1 tumour cells. Rejection of medium-sized inocula 25 or 500 H1 tumour cells were abolished by the administration of LPS. In contrast, the 'strongly' antigenic H7 tumour did not exhibit the 'sneaking through' phenomenon and its growth was only temporarily affected by LPS. Studies were also performed to determine the effect of LPS on the kinetics of delayed-type hypersensitivity (DTH) induced by mitomycin C-treated (MCT) H1 or H7 tumour cells inoculated s.c. into the footpads of mice. The 'strongly' antigenic MCT H7 tumour cells induced consecutive waves of footpad swelling of diminishing intensity and corresponded to periods of anti-tumour resistance. The specific phase of MCT H7-induced footpad swelling, maximal at day 6, was delayed in its induction if LPS was administered concurrently with MCT H7 tumour cells. In contrast, the 'weakly' antigenic MCT H1 tumour cells induced only one specific phase of footpad swelling which was rapidly down-regulated. The induction of immunity by MCT H1 tumour cells was also delayed by the concomitant administration of LPS. Because the 'weakly' antigenic H1 tumour was unable to sustain consecutive waves of anti-tumour immunity, the delay in the expression of such immunity by LPS allowed the H1 tumour cells to multiply to eventually overwhelm a rapidly down-regulated immune response. In contrast, the incidence of tumours arising from the 'strongly' antigenic H7 tumour cells was not significantly affected in LPS-treated mice because the tumour cells which escaped the first encounter with delayed anti-tumour immunity, succumbed to subsequent waves of resistance in both normal and LPS-treated mice injected with fewer than 1 X 10(5) H7 tumour cells.     

Malignant myoepithelioma of the vulva resembling a rhabdoid tumour

AIMS: We report an example of malignant myoepithelioma of the vulva, which has not been hitherto described. We discuss the differential diagnosis and briefly review the literature. METHODS AND RESULTS: The lesion was found in an 81-year-old woman as an indolent 40 mm tumour. The neoplastic cells showed a myoid, spindled, epithelioid and plasmacytoid phenotype. Hyalinization of extracellular material and myxoid changes were present. There was a partly solid and microcystic pattern and a tight cohesiveness of cells was lacking. The circumscribed multinodular tumour somewhat resembled an extrarenal rhabdoid tumour, having large tumour cells with prominent nucleoli and large amounts of acidophilic cytoplasm. Immunohistochemically, the tumour cells were immunoreactive for cytokeratin, vimentin, muscle-specific actin, alpha-smooth muscle actin, and S100 protein, but not for desmin, epithelial membrane antigen, factor VIII-related antigen, CD34 and CD31. CONCLUSIONS: The histological and cytomorphological appearance of the tumour well as the immunohistochemical findings suggest the diagnosis of malignant myoepithelioma, possibly derived from minor vestibulary glands or ectopic breast tissue. Differential diagnoses are, in particular, extrarenal rhabdoid tumour and 'proximal type' epithelioid sarcoma. Differentiation is important, because the tumours show a different behaviour and prognosis.     

Sebaceous carcinoma of the parotid gland

Summary Sebaceous carcinoma of salivary gland origin is extremely rare and, because of its rarity, the clinicopathological characteristics and the histogenesis are not fully understood. We present a case of sebaceous carcinoma of the parotid gland which brings the total number of reported cases to 22.The tumor showed epithelial cell nests which were mainly composed of sebaceous cells with marked cellular atypia. In most of the nests, glandular spaces lined by ductal epithelium were present. Scattered mucous cells and flattened eosinophilic cells at the periphery of the nests were also seen. Ultrastructural and immunohistochemical observations of the tumour revealed coexistence of sebaceous and glandular differentiations in some tumour cells. Tumour cells with lipid granules often participated in the formation of glandular structures or exhibited intracytoplasmic lumina, and immunohistochemical localization of lactoferrin and secretory component, the functional markers of ductal epithelium of salivary gland, was demonstrated not only in duct-forming tumour cells but also in many sebaceous tumour cells.It seems likely that sebaceous carcinoma originates from pluripotential duct cells which can differentiate into sebaceous, ductal and mucous cells.     

A critical and comparative study of methods of isolating tumour cells from the blood

From the large number of techniques described for isolating tumour cells from the blood, five have been selected because they were those most widely used by previous workers and can be performed with a minimum of special apparatus. These techniques have been assessed for their efficiency in retaining tumour cells added to blood samples, in eliminating normal blood cells, and in preserving the morphology of the retained cells.     

An Experimental Study on the Anti-Ehrlich Ascites Carcinoma Effect of Purified Toad Venom Extract

The objective of this paper was to study the anti-Ehrlich ascites carcinoma effect of purified toad venom extract and its mechanism. Mouse model of Ehrlich ascites carcinoma was established with cisplatin as the control to observe the inhibitory effect of purified toad venom extract on malignant peritoneal effusion in mice. The results showed that compared with the control group, ascites volume, number of tumour cells and tumour cell viability decreased and ascites inhibition rate reached over 50% in each treatment group, and with the increase of the dose, incidence of ascites showed a downward trend. The number of tumour cells in ascites and tumour cell viability in the purified toad venom high-dose group were lower than those of the cisplatin group. Compared with the model group, survival time was prolonged in varying degrees in the purified toad venom groups and cisplatin group. The study concluded that purified extract of toad venom has an anti-Ehrlich ascites carcinoma effect.     

Ultrastructural classification of tumor cells (based on the example of human squamous cell lung cancer)

Ultrastructural classification of tumour cells of human neoplasms is suggested on the basis of ultrastructural examination of 110 lung squamous cell carcinomas. All tumour cells may be subdivided into two groups: the tumour cells of the first group show the ultrastructural organ- and (or) tissue specific features; the tumour cells of the second group do not exhibit such features. The tumour cells of the 1st group may be represented by one, two or several different types. The tumour cells of both groups, depending on the intracellular organelles development, may belong to one of the following three types: 1) rich in organelles, 2) with moderate number of organelles and 3) poor in organelles. All the tumours on the whole should be divided into three varieties: the first variety--the number of the 1st group cells dominate; the second variety--the number of cells of the 1st and the 2nd group is the same; the third variety--the 2nd group cells dominate. The structural-functional alterations of tumour cells (change of nuclei and other organelles) should be fixed in every case. The proposed classification reflects the diagnostic needs of electron microscopy.     

Modulation of anti-tumour immunity and the effect of bacterial endotoxin on the growth of different syngeneic tumours from small inocula in mice.

Studies were undertaken to determine the influence of E. coli lipopolysaccharide (LPS) on the growth of various doses of two antigenically-distinct syngeneic murine fibrosarcomas designated H1 and H7. The ''weakly'' antigenic H1 tumour injected subcutaneously (s.c.) along the abdominal wall was profoundly susceptible to the growth-potentiating effects of a single intraperitoneal (i.p.) injection of 2 micrograms LPS, administered concurrently. ''Sneaking through'' effects in control mice were observed with doses of 10 and 100 H1 tumour cells. Rejection of medium-sized inocula 25 or 500 H1 tumour cells were abolished by the administration of LPS. In contrast, the ''strongly'' antigenic H7 tumour did not exhibit the ''sneaking through'' phenomenon and its growth was only temporarily affected by LPS. Studies were also performed to determine the effect of LPS on the kinetics of delayed-type hypersensitivity (DTH) induced by mitomycin C-treated (MCT) H1 or H7 tumour cells inoculated s.c. into the footpads of mice. The ''strongly'' antigenic MCT H7 tumour cells induced consecutive waves of footpad swelling of diminishing intensity and corresponded to periods of anti-tumour resistance. The specific phase of MCT H7-induced footpad swelling, maximal at day 6, was delayed in its induction if LPS was administered concurrently with MCT H7 tumour cells. In contrast, the ''weakly'' antigenic MCT H1 tumour cells induced only one specific phase of footpad swelling which was rapidly down-regulated. The induction of immunity by MCT H1 tumour cells was also delayed by the concomitant administration of LPS. Because the ''weakly'' antigenic H1 tumour was unable to sustain consecutive waves of anti-tumour immunity, the delay in the expression of such immunity by LPS allowed the H1 tumour cells to multiply to eventually overwhelm a rapidly down-regulated immune response. In contrast, the incidence of tumours arising from the ''strongly'' antigenic H7 tumour cells was not significantly affected in LPS-treated mice because the tumour cells which escaped the first encounter with delayed anti-tumour immunity, succumbed to subsequent waves of resistance in both normal and LPS-treated mice injected with fewer than 1 X 10(5) H7 tumour cells.     

A poorly differentiated apudoma of the gallbladder]

Apudoma was found in the gall bladder removed in a 76-year-old woman because of the chronic calculous cholecystitis exacerbation. Carcinoid syndrome was absent clinically. Histologically, the tumour was a poorly differentiated carcinoid with areas of small cell and polymorphic carcinoma. Argyrophilic Pasquale reaction in the tumour cells was negative, electron microscopically small neurosecretory granules were found. Numerous ACTH-reactive cells and single serotonin-reactive cells were revealed in the tumour parenchyma by means of immunohistochemical PAP-method using antibodies against ACTH, serotonin, calcitonin, somatostatin, insulin, glucagon, P-substance. Focal hyperplasia and intestinal metaplasia of epithelium with the increase of the number of argyrophilic, ACTH-reactive cells were observed outside the tumour.     

Angiomyolipoma of the kidney. Immunoreactivity with HMB-45. Light- and electron-microscopic findings

Immunoreactivity with HMB-45 has recently been described in renal angiomyolipoma, a tumour of smooth muscle cells. HMB-45 is a monoclonal antibody that reacts specifically with melanosomes. In order to determine whether the tumour cells contain melanosomes and synthesize melanin, seven tumours were studied by light microscopy and immunohistochemically with the antibodies HMB-45, KP1 (CD68), PG-M1 (CD68), Ki-M1P, anti-lysozyme, anti-smooth-muscle actin, anti-vimentin, anti-S100 protein and KL1 (anti-keratin). Two tumours were also studied by electronmicroscopy and one by immuno-electronmicroscopy. Histochemical investigation for dopa oxidase was performed on cryostat sections. The tumours contained varying numbers of HMB-45-positive muscle cells. Reactivity was noted in lysosomal granules and rough endoplasmic reticulum. Typical premelanosomes were found in the tumour cells by electronmicroscopy. Groups of tumour cells stained for dopa oxidase. The tumour cells were not reactive for lysozyme, but reacted with KP1, PG-M1 and Ki-M1P. Immuno-electronmicrosopy showed that reactivity for KP1 was located within lysosomal granules. The findings show that the tumour cells of renal angiomyolipoma contain premelanosomes and that they are able to synthesize melanin, because they contain dopa oxidase. Immunoreactivity with KP1, PG-M1 and Ki-M1P can be attributed, in the absence of staining for lysozyme, to the large number of lysosomal granules. The tumour cells were not found to be related to macrophages or myeloid cells.     

Surface markers of small lymphocytes appearing in the mouse Ehrlich ascites tumour, host spleen and blood

Small lymphocytes sampled from intraperitoneally growing Ehrlich ascites tumour in CBA/H-T₆ mice as well as host spleen and blood at different days of tumour development were characterized radioautographically on the basis of two surface markers, IgM for B cells and θ antigen for T cells. A direct binding of ¹²⁵I-labelled anti-IgM detected natural surface IgM, while an indirect binding following a prior exposure to anti-θ antibody detected θ antigen. Cells remaining unlabelled with the latter procedure were considered to lack both markers (double negative). While the incidence of IgM+ve small lymphocytes within the tumour declined, their absolute numbers increased with tumour growth. Low levels of antiglobulin binding shown by these cells were considered to reflect low levels of maturation, because (1) our previous studies indicated that they were newly formed, and (2) the extent of antiglobulin binding by B lymphocytes in the marrow is known to increase with increasing post-mitotic age. The proportions and the absolute numbers of θ+ve as well as the double negative small lymphocytes increased within the growing tumours. Within the host spleen, the incidence of IgM+ve small lymphocytes remained unchanged but their absolute numbers increased because of splenomegaly. The degree of antiglobulin binding by these cells was comparable to that of the normal splenic population. The incidence of θ+ve cells dropped but their absolute numbers remained unchanged in the spleen during tumour growth. In contrast, the incidence as well as the absolute numbers of double negative cells increased markedly. This cell category increased also in the blood, possibly in transit to the tumour site and other lymphoid organs from the bone marrow, where they were most prevalent. Their bone marrow origin was further suggested by a preponderance of marrow derived small lymphocytes at the tumour site as well as in the host spleens found in our earlier studies. Double negative population in the spleen showed a paucity of C′3 and Fc receptors on the cell surface and included cells capable of producing B lymphoid colonies in vitro.     

Ultrastructure of tumour invasion and desmoplastic response of bronchogenic squamous cell carcinoma

Summary Using ultrastructural methods we studied the interaction of tumour cells and lung parenchyma in deep areas (i.e., more than about 3 mm from the tumour surface) of 50 bronchogenic squamous cell carcinomas. The tumour periphery, studied previously, had shown organized associations of tumour cells and lung epithelial cells and a surprising lack of invasion of non-epithelial tissue compartments. The deeper areas, where the tumour cells and the lung parenchyma had been in contact for longer periods, consisted of irregular groups of tumour cells and desmoplastic stroma which was very similar to granulation tissue. The deeper areas also contained many intact lung epithelial cells, arranged in compressed and distorted alveolar structures. Where non-neoplastic epithelial cells and tumour cells had direct contact, they formed common junctional complexes and basal laminae. In part of the tumours, the cells were largely devoid of a basal lamina. However, in most instances a continuous basal lamina surrounded every tumour cell group studied, even when these formed irregular strands or seemed to be completely isolated.     

Immunohistochemical expression of p53 proteins in Wilms' tumour: a possible association with the histological prognostic parameter of anaplasia

Wilms' tumour (nephroblastoma) has been associated with chromosomal abnormalities at the 11p13, 11p15 and 16q regions. A study into the possibility of mutations occurring within p53, the ubiquitous adult tumour suppressor gene, in Wilms' tumour was carried out. Thirty-eight ca ses were studied. Of these 36 were categorised into the favourable histology group and two into the unfavourable histology group based on the National Wilms' Tumour Study criteria. Archival formalin-fixed, paraffin-embedded tissue sections from each case were stained with a polyclonal (AB565:Chemicon) and a monoclonal (DO7:Dako) antibody raised against p53 protein using a peroxidase-labelled streptavidin biotin kit (Dako). 'Cure' (disease-free survival of 60 months or longer) was documente d in 39% of cases with favourable histology tumours. Eleven percent in this group succumbed to the disease. Both cases with unfavourable histology died. Four out of 36 (11%) tumours with favourable histology demonstrated weak to moderate staining with both AB565 and DO7 in more than 75% of tumour cells. In contrast, p53 protein expression in unfavourable histology tumours was significantly increased compared with the favourable histology group ( P  = 0.021) with both cases demons trating immunopositivity in >75% of tumour cells when stained with AB565 and DO7. The intensity of staining ranged from moderate to strong in both cases. It appears from this preliminary study that the immunohistochemical expression of p53 protein in Wilms' tumour, presumably a result of mutation in the p53 tumour suppressor gene, correlates with histological classification, histological categorisation being one of the useful features in the prognostic assessment of Wilms' tumours.     

Cellular fibroma of the ovary: description of a case

Cellular fibromas of the ovary are rare neoplasms belonging to the group of sex-cord stromal tumours. They have been described to show from 1 to 3 mitotic figures per 10 high power fields (HPF) and they generally behave in a benign fashion. Herein we describe the clinicopathological features of a case of ovarian cellular fibroma. The patient, a 22-year-old woman, presented with acute abdominal pain. Laparotomy revealed a large ovarian mass. Histologically the lesion was composed of spindle cells showing slight or moderate pleomorphism and 3 mitoses per 10 HPF. The spindle cells were immunoreactive for vimentin, smooth muscle actin and inhibin alpha-subunit. The differential diagnoses that we considered included the mitotically active leiomyoma because of the strong positivity for smooth muscle actin, but positive immunoreaction with anti alpha-inhibin antibody helped in confirming a sex-cord stromal tumour. Electron microscopy did not show any evidence of smooth muscle differentiation.     

Sclerosing stromal tumour of the ovary--a case report.

Sclerosing stromal tumour (SST) of the ovary is a rare, benign tumour of the ovary, distinct from thecoma-fibroma group of tumours because of predominant occurrence below 30 years of age, lack of hormonal manifestations and histologic heterogenity. A case of 17-year-old female patient is described in the present article. The differential diagnosis is also discussed.     

Automated screening for micrometastases in bone marrow smears

It is possible to detect micrometastases in primary breast cancer using immunocytochemical staining of bone marrow smears. However, using the light microscope the procedure is time-consuming and laborious because such cells occur rarely (less than 1 in 10,000). Using an image analysis system, the Leytas machine, and a specially prepared reproducible slide it has been possible to automate the technique. A 100% concordance was found between the machine and the light microscope in the identification of slides containing moderate to high numbers of tumour cells in bone marrow, and in those containing no tumour cells. However, in those slides containing low numbers of tumour cells (1-10 tumour cells/10(6) normal bone marrow cells) the sensitivity was decreased to 91%. In the presence of non-specific staining the false positive rate was increased from 0% to 22%. This method represents a potential improvement in the assessment of an important clinical staging procedure.     

Epithelioid and retiform hemangioendothelioma of the skull bone--report of four cases

Hemangioendothelioma (HE) is a borderline or intermediate type of vascular neoplasm. We report clinical and histopathological characteristics of four cases of HE arising from the skull bones because of its extreme rarity in this location. The age of the patients ranged from 6-45 years. Three patients presented with a painless swelling over the head and one case had sphenoid wing mass with dimness of vision and proptosis. Radiographic images showed a well-demarcated, osteolytic lesion in the skull bone in all the four, one case in addition had sclerotic edges and another had specks of calcification. Grossly, the tumour was very vascular with hemorrhagic areas. Histologically, three cases showed features of an epithelioid variant of HE, with short strands and solid nests of rounded to slightly spindled, eosinophilic endothelial cells, some of them having small intracellular vacuoles. The stroma was myxoid--hyalinised with focal mixed inflammatory infiltrate. One case had features of a 'retiform' histological variant composed of numerous elongated vessels lined by a single layer of hobnail endothelial cells, focal lymphocytic infiltrate and papillae with hyaline collagenous cores. The tumour cells in all the four were immuno-labelled by antibody to factor VIII-associated protein. The tumour cells lacked cytological atypia and mitosis was sparse. These features were important in prognostication as low-grade tumours can be cured by complete wide-resection.     

Tumour vascularity and proliferation: clear evidence of a close relationship.

Evaluation of various prognostic factors often reveals that some are closely related. In this issue of the Journal of Pathology, evidence is presented linking intratumoural microvessel density with tumour cell proliferation. This is expected, because an adequate blood vascular system is necessary for effective tumour cell proliferation. The blood vascular supply of a tumour is critical not only in providing tumour cells with nutrients, oxygen, and waste elimination, but also because activated endothelial cells release important paracrine growth factors for tumour cells and secrete collagenases, urokinases, and plasminogen activator. The latter allow capillary ingrowth and the spread of tumour cells into and through the adjacent fibrin-gel matrix, connective tissue stroma, and into the lymphatic and/or vascular spaces. Finally, an adequate vascular supply helps to 'switch off' apoptosis and prevent other forms of tumour necrosis, thus contributing to overall tumour growth and spread.     

The spectrum of micronodular thymic epithelial tumours with lymphoid B-cell hyperplasia

AIMS: A rare type of thymoma, micronodular thymoma with lymphoid B-cell hyperplasia, was recently reported by Suster and Moran. Thymic epithelial tumours with a similar pattern but with varied cytological features of the tumour cells are analysed. METHODS AND RESULTS: A total of 11 cases of thymic epithelial tumours characterized by micronodular proliferation of tumour cells separated by abundant lymphoid stroma with prominent germinal centres were reviewed clinicopathologically and examined immunohistochemically. The presence of Epstein-Barr virus (EBV) genome was also examined by in-situ hybridization. Based on the morphology of tumour epithelial cells, cases were subdivided into four groups: group 1 (two cases) having spindle epithelial cells; group 2 (two cases) showing an admixture of spindle and polygonal epithelial cells; group 3 (five cases) having polygonal epithelial cells, with mild to moderate cytological atypia in four cases, and group 4 (two cases) representing lymphoepithelioma-like carcinoma. The degree of cytological atypia and the number of tumour cells positive for MIB-1 and p53 gradually increased towards group 4. The abundant lymphoid stroma in all cases contained many CD20-positive B-cells and CD3 and CD45RO-positive T-cells. CD99-positive immature T-cells were present in all cases of groups 1 and 2 and in most cases of group 3, but not in both cases of group 4 tumours. IgG, IgM and IgD-positive plasma cells and lymphocytes were also present in all cases, more prominent in those of groups 3 and 4. The EBV genome was detected in only a few lymphocytes in five cases. CONCLUSIONS: The tumours in this series belong to a distinct category of thymic epithelial tumours and each of the above groups may constitute a spectrum in the continuum of cytological atypia. The aetiological relationship of EBV with these tumours could not be proved. The lymphoid B-cell hyperplasia may result from a host immune response and may suggest a favourable clinical course of this type of tumour.     

Elevated serum alpha-fetoprotein in a patient with undifferentiated carcinoma of the gall bladder.

An uncommon case of undifferentiated carcinoma of the gall bladder in a 65 year old Chinese man, who presented with an increased serum alpha-fetoprotein concentration, is reported. Histologically, the tumour had a primitive appearance and was composed of a pavement-like array of poorly differentiated columnar/polygonal cells. Alpha-fetoprotein was demonstrated in some of the tumour cells using an immunoperoxidase technique. Alpha-fetoprotein secretion in this instance may have occurred because the gall bladder and the liver are of similar embryological origin. Alpha-fetoprotein may also be related to the resurgent expression of oncofetal antigens. This tumour may represent another rare cause of increased serum alpha-fetoprotein concentrations.     

Spindle cell squamous carcinoma of the oral region

Summary Six cases of spindle cell squamous carcinoma (SCSC) of the oral cavity were studied clinicopathologically, immunohistochemically and ultrastructurally to summarize the clinicopathological features of this rare neoplasm and to discuss the debatable histogenesis of the sarcomatoid component and the differential diagnosis of SCSC. The mean age of the patients was 72 years and the female to male ratio was 1:2. Four of them had a history of irradiation for pre-existing squamous cell carcinoma. One patient died of SCSC. While clinical and histological prognostic factors of SCSC could not be determined, it was shown that radical surgery resulted in good prognosis. The epithelial nature of the sarcomatoid component of SCSC was clearly revealed by a combination of immunohistochemical staining for keratins and electron microscopic demonstration of tonofilament-like filaments and/or desmosome-like structures. Together with electron microscopic evaluation of the tumour cells, immunohistochemical characterization of tumour cells using antibodies to keratin, vimentin, glial fibrillary acidic protein and S-100 protein is very helpful in differentiating SCSC from true spindle cell sarcoma, melanoma and malignant myoepithelioma. In the immunohistochemical differential diagnosis of SCSC, it is important to remember that SCSC should not be ruled out of the differential diagnosis by a positive reaction for vimentin in sarcomatoid tumour cells. Absence of staining for keratin in the sarcomatoid tumour cells does not always exclude SCSC, because some SCSCs show immunoreactivity of keratin in their sarcomatoid components only with some anti-keratin antibodies. Different kinds of anti-keratin antibodies should be applied in the differential diagnosis of SCSC.