HERG1 gene expression as a specific tumor marker in colorectal tissues.

INTRODUCTION: Colorectal carcinomas exhibit a frequent recurrence after curative surgery, which may partially be due to histopathologically inconspicuous minimal residual disease. Reliable markers for tumor cells in colorectal tissue are still missing. Therefore, in this study we compared the predictive value of the putative tumor markers carcinoembryonic antigen (CEA), cytokeratin-19 (CK19) and cytokeratin-20 (CK20) to that of a novel marker, the human ether-a-go-go-related gene (HERG1) K(+) channel, a suggested regulator of tumor cell proliferation. MATERIALS AND METHODS: Using RT-PCR we studied HERG, CEA, CK19 and CK20 expression in colorectal carcinomas and non-carcinoma controls. HERG1 immunhistochemistry was performed in a total of 66 specimens, in colorectal carcinoma (n = 23), in matched histopathologically negative samples (n = 23) taken near the excision site from the same tumor patients and in healthy control biopsies (n = 20). In order to verify the relevance of HERG1 for tumor proliferation we studied the effect of HERG1 inhibition in the Colo-205 colon cancer carcinoma cell line using the MTT-assay. RESULTS: HERG1 was expressed in all tumor samples regardless of their stage and in adenomas larger than 0.4 cm, but absent in small adenomas, sigmadiverticulitis specimen and healthy histopathologically negative samples, except for one which developed a tumor recurrence. In contrast, CEA, CK19 and CK20 were absent in some tumors. The selective HERG1 inhibitor E-4031 dose-dependently impaired tumor growth in the proliferation assays. DISCUSSION: Our data indicate that HERG1, but not CEA, CK19 or CK20, is a highly sensitive and reliable tumor biomarker that may constitute a novel molecular target for tumor treatment.

Immunocytological analysis of leukocyte subpopulations in urine specimens before and after prostatic massage.

OBJECTIVE: To evaluate the presence of leukocyte subpopulations in urine after prostatic massage (VB 3) in symptomatic patients with > or =10 leukocytes/high power field (magnification x1,000) in expressed prostatic secretions, and who were classified as suffering from chronic bacterial prostatitis or inflammatory chronic pelvic pain syndrome. METHODS: 115 consecutive patients were investigated. Granulocytes in centrifuged midstream urine (VB 2) and VB 3 were counted after Papanicolaou stain. Macrophages, B and T lymphocytes were analyzed after immunocytological staining with monoclonal antibodies according to the alkaline phosphatase anti-alkaline phosphatase method. The counts were quantified as the number of cells per view field at a magnification of x400. In all patients, acute or chronic urethritis had been excluded before enrollment in the study. 16 men without signs or symptoms of urogenital inflammation served as controls. RESULTS: Of the 115 patients, 101 men demonstrated > or =10 leukocytes/view field in VB 3. In comparison to VB 2, the leukocyte subpopulations in VB 3 demonstrated an increase in granulocytes (9.2-fold), macrophages (7.6-fold), T lymphocytes (7.6-fold), and B lymphocytes (4-fold). The increase was statistically significant (p<0.001 each). The proportion of these cells in VB 3 was 81.6, 11.1, 5.5, and 1.8%, respectively. As compared to controls, all leukocyte subsets in VB 3 were significantly elevated (p>0.001 each). CONCLUSION: Elevated numbers of leukocytes in VB 3 are indicative of prostatitis provided that urethral inflammation and leukocyturia in VB 2 are excluded. Granulocytes are the predominant cell type of inflammation. The increase in macrophages, T and B lymphocytes in prostatic secretions indicate the participation of both the cellular and humoral immune system in the inflammatory process.

[Bone marrow granulomatosis in Q-fever]. / Knochenmarksgranulomatose bei Q-Fieber.

BACKGROUND: Petersdorf and Beeson defined Fever of Unknown Origin (FUO) as an illness characterized by rectal temperature exceeding 38.3 degrees C on at least 3 occasions, evolving during at least 3 weeks, with no diagnosis reached after 1 week of in-patient investigation. A quarter of FUO cases is caused by infectious diseases, most often hidden abscesses, subacute endocarditis and tuberculosis. CASE REPORT: In a 29-year-old patient with undulating fever of 3 months duration solely the demonstration of bone marrow fibrin ring granulomas led to the diagnosis of protracted Q-fever. The diagnosis later has been proved by elevated Coxiella burnetii antibody titers. CONCLUSION: The case report underlines the diagnostic value of bone marrow biopsy in the evaluation of fever of unknown origin.

Nephron-sparing surgery for renal cell carcinoma--is tumor size a suitable parameter for indication?

OBJECTIVES: Because patients with small renal cell carcinomas (RCC) are being treated by nephron-sparing surgery with increased frequency, a generally accepted parameter giving additional information as to which kind of tumor is suitable for this treatment is urgently required. METHODS: In a retrospective analysis of 245 patients who underwent radical nephrectomy for RCC, we investigated whether tumor size could provide the necessary information. We analyzed the association of tumor size with pTNM classification, grade, and the findings of the flow cytometric analysis of the DNA content analyzing the DNA index (DI). RESULTS: Stage pT1 was found in 23 patients (9.4%), pT2 in 100 (40.8%), pT3 in 109 (44.5%), and pT4 in 13 patients (5.3%). Grade 1 was found in 87 patients (35.5%), grade 2 in 120 (49.0%), and grade 3 in 38 patients (15.5%). A low DI was found in 71%, a moderately increased DI in 20%, and a high DI in 9%. Lymph node metastases were detected in 14% and -distant metastases in 22%. Closer examination of the tumors less than 2.5 cm (n = 23) revealed a significantly lower incidence (P <0.001) of infiltration of the renal capsule (n = 0) than in the rest of the group. Positive lymph nodes or distant metastases could not be found in this subgroup. A multifocal appearance of RCC was detected in only 2 (8.7%) of the 23 patients; it was detected in 67 (30.2%) of the 222 patients in the rest of the group. None of the 23 patients had grade 3 tumors (P <0.05). Fifty-two percent of the tumors were grade 1 and 48% grade 2. None of the 23 had a high DI; a moderately increased DI was found in 1 patient and a low DI in 22 of the 23 patients. Detailed examination of the tumors between 2.5 and 4 cm (n = 29) revealed an infiltration of the renal capsule in 11 (38%); lymph node metastases were found in 2 (6.9%) and metastases in 4 (13.8%). A multifocal appearance was found in 4 (13.9%) of the 29 patients; grade 3 tumors were detected in 3 (10.3%) of 29 patients, grade 2 tumors in 12 (41.4%), and grade 1 tumors in 14 (48.3%). In this subgroup, a high DI was found in 14% (not significant). The examination of tumors larger than 4 cm in size revealed worse results in the pTNM classification, grade, and flow cytometric results. CONCLUSIONS: Only tumors smaller than 2.5 cm should be considered suitable for nephron-sparing surgery in patients eligible for elective surgery. In patients in whom nephron-sparing surgery is imperative, even tumors between 2.5 and 4 cm appear to be suitable. In patients requiring extensive resection, however, the risk of local recurrence seems to be higher because of the higher incidence of multifocality.

Expression of p16 and lack of pRB in primary small cell lung cancer.

The retinoblastoma protein (pRB), p16, and cyclin D1 are major components of the RB pathway, which controls the G1 checkpoint of the cell cycle. Proper regulation of this pathway is crucial for normal cell proliferation. Abnormal forms of these proteins have been found in various types of malignant tumours. In the present report, immunohistochemical techniques were applied to study the expression of pRB, p16, and cyclin D1 in 161 samples of primary small cell lung cancer (SCLC) and 20 samples of non-small cell lung cancer (NSCLC). While pRB and cyclin D1 staining was negative in 161 specimens of SCLC, expression of p16 was observed in 153 samples. In contrast to SCLC, 16 out of 20 NSCLC cases exhibited pRB expression and 15 showed cyclin D1 expression, but only very weak p16 staining was found in five samples. These observations could provide additional criteria for the distinction between SCLC and NSCLC. Furthermore, these findings, based on primary tissues, implicate different mechanisms in the tumourigenesis of SCLC and NSCLC.

Immunoselection in vivo: independent loss of MHC class I and melanocyte differentiation antigen expression in metastatic melanoma.

Peptides derived from melanocyte differentiation antigens have been identified as targets for MHC class I-restricted cytolytic T lymphocytes (CTLs) in human melanoma Regression of antigen-expressing tumors as well as selection of antigen-loss variants in the presence of antigen-specific CTLs have previously been reported. In the present study, we determined the expression of the melanocyte differentiation antigens Melan A/MART-1 and tyrosinase by mRNA analysis and by immunohistochemical staining with the monoclonal antibodies (MAbs) A103 and T311. Co-expression of Melan A/MART-1 and tyrosinase was detected by both methods in 18/20 melanomas tested. However, immunohistochemistry provided additional information on intensity and microheterogeneity of antigen expression that cannot be detected by mRNA analysis as a molecular basis for the escape from CTL recognition of antigen-negative tumor cells. Comparative analysis of repeated biopsies of metastatic lesions in 5 HLA-A2+ patients showed a gradual loss of Melan A/MART-1 expression in 4/5 and of tyrosinase in 2/5 samples in association with tumor progression. However, 3 of these patients had growing antigen-positive tumors in the presence of antigen-specific CTLs. This led us to assess the expression of MHC class I, the essential restriction element for CTL recognition, and of HLA-A2. We found an unexpectedly high frequency of MHC class I-negative tumors (9/20). Loss of MHC class I expression was detected in 3/5 progressive tumors and isolated loss of HLA-A2 in 1/5 tumors. Our results suggest that strategies enhancing the expression of MHC class I and tumor-associated antigens need to be considered in attempts at making vaccination more effective.

Applied Sciences: Absence of CD44-standard in human neuroblastoma correlates with histological dedifferentiation, N-myc amplification and reduced survival probability.

Expression of CD44 was examined by immunohistochemistry in 205 primary neuroblastomas together with histological grading according to the Shimada classification at the time of diagnosis. In addition, Southern blot analysis to determine N-myc gene amplification was carried out in the same tissue. When compared with clinical data such as stage, age and event-free survival probability it was found that CD44 expression characterizes well differentiated tumours and thus correlates with prognosis (event-free survival probability in CD44 positive patients 0.6 (n = 129) vs. 0.0 (n = 21) in CD44 negative patients). In tumours with N-myc = 1, CD44 positivity was found in 91% of patients as compared to 57% of patients with N-myc > 1 in tumours. All tumours of 13 patients with metastatic stage 4s (with good prognosis) showed CD44s expression. Thus, detection of CD44s expression might serve as a prognostic indicator which can be rapidly detected at diagnosis.

Intermediate filaments in rat pancreatic acinar tumors, human ductal carcinomas, and other gastrointestinal malignancies.

BACKGROUND/AIMS: Keratin is a member of the intermediate filament family in epithelial cells. Two-dimensional gel electrophoresis of different epithelial cells has shown 20 different keratin polypeptides. Therefore, mapping of the keratin polypeptides can be used to define a specific tissue. METHODS: Cytokeratin expression was investigated by using monoclonal antibodies in human surgical specimens and autopsy material of pancreatic, gastric, liver, and colon carcinomas and cholangiocarcinomas, and their metastasis to lymph nodes and liver was examined. In addition, rat acinar cell carcinomas were used to compare cytokeratin expression in ductal vs. acinar cell pancreatic carcinomas. RESULTS: Human pancreatic ductal carcinomas expressed keratins 7, 8, 18, and 19, whereas the majority of rat acinar carcinomas did not express keratins typical for ducts in rat pancreas. The keratin patterns of gastric and colon carcinomas were identical with keratins 8, 18, and 19. In contrast, hepatocellular carcinomas expressed the same keratin pattern as pancreatic acinar carcinomas with keratins 8 and 18, whereas cholangiocarcinomas expressed keratin 7, 8, 18, and 19, similar to pancreatic ductal carcinomas. Metastasis of pancreatic ductal and colon carcinomas retained their keratin patterns. CONCLUSIONS: Keratin polypeptide typing of unknown malignant cells can be a useful tool for cell identification.

Primary leiomyosarcoma of the spinal leptomeninges.

A case of leiomyosarcoma of the spinal leptomeninges is presented, with clinical, radiological, light microscopic and immunohistochemical data. The probable origin of the tumor from a pluripotent mesenchymal cell is discussed.

[Three cases of a fulminant course of idiopathic pulmonary fibrosis (Hamman-Rich syndrome)]. / Drei Fälle von foudroyant verlaufender idiopathischer Lungenfibrose (Hamman-Rich-Syndrom).

The Hamman-Rich syndrome is defined as an acute pulmonary disease of unclear aetiology that takes a rapid and prognostically unfavourable, usually fatal course. We report on three patients admitted to the intensive-care ward during a period of 6 months in a state of mandatory artificial respiration, each patient dying within 3 weeks after admission. Basing on clinical and histological criteria these patients were diagnosed as suffering from Hamman-Rich syndrome. At the time of the clinically identifiable onset of the disease all the patients had fever (> 39 X), leucocytosis (> 20 x 10(3)/microliters) and dyspnoea. These signs and symptoms were at first, in conjunction with the radiological identification of diffuse pulmonary infiltrations, misdiagnosed as pneumonia. The patients had to be artificially respirated after a short time because of the foundroyant course of the disease. Despite optimised respiratory parameters it was already initially apparent that there was a severe disturbance of the gas exchange function (paO2/FiO2 < 150) and high respiratory pressures (> 40 mmHg). Polymorphonuclear neutrophilics dominated in the bronchoalveolar lavage. Lung biopsy showed marked fibrosing that was a decisive factor in diagnosing. An infectious agent as triggering cause of the disease could not be identified in any of the patients. Treatment was effected with antibiotics, steroids and cyclophosphamide. The patients died after 14, 17 and 21 days, respectively, from intractable respiratory insufficiency with increasing loss of compliance of the lungs (compliance of lungs and thorax < 20 ml/mmHg).(ABSTRACT TRUNCATED AT 250 WORDS)

Monoclonal antibodies against luminal membranes of renal proximal tubules which are kidney-specific.

After immunization with porcine brush-border membrane proteins, 11 monoclonal antibodies were generated which react with proximal tubules. Their antigenic polypeptides were characterized with respect to apparent molecular weight, histochemical localization in porcine and human kidney, and tissue distribution in pig. In porcine kidney, six antibodies bind selectively to the proximal tubule whereas the others also react with other nephron segments. With the exception of one antibody which reacts with the luminal and the basolateral membrane of the porcine proximal tubule, the other antibodies specific for the proximal tubule only stain the brush-border membrane. Four of them react along the entire length of the porcine proximal tubule, whereas one (R1A2) binds to the S3-segment in pig and to the entire length of the proximal tubule in man. This indicates that segment-specific expression may be species-dependent. Testing the antibodies in 21 different extrarenal tissues it was found that three of the antibodies, specific for the brush-border membrane in renal proximal tubules, only react in kidney. Two of these are specific for pig kidney whereas one also reacts with human kidney. This antibody (N4A4) is directed against a polypeptide with an apparent molecular weight of 400,000. Electron microscopic immunohistochemistry showed that N4A4 binds to the intervillus region of the brush-border membrane and to subapical vesicles.

Expression of cell adhesion molecules alpha-2, alpha-5 and alpha-6 integrin, E-cadherin, N-CAM and CD-44 in renal cell carcinomas. An immunohistochemical study.

Renal cell carcinoma is known to metastasize early independent of tumour grade. Invasion of the renal vein plays an important role in the prognosis. Cell adhesion molecules have been investigated, including the expression of alpha-2, alpha-5, and alpha-6 integrin, E-cadherin, neural-cell adhesion molecule and CD-44 in 34 renal cell carcinomas, using the alkaline phosphatase-anti-alkaline phosphatase technique. Our results indicate a differential expression of these cell adhesion molecules (alpha-2, alpha-5 and E-cadherin) depending on histological type and tumour grade.

Translocation (5;22) in an Askin's tumor.

A boy with a brain metastasis of an Askin's tumor was investigated. Cytogenetic studies revealed a near-diploid karyotype with monosomies of chromosomes 5 and 22 in the presence of a derivative chromosome der(5)t(5;22)(q35;q11). In particular, no involvement of chromosome 11 was seen.

Cytokeratin expression in mucinous sweat gland carcinomas: an immunohistochemical analysis of four cases.

Four mucinous sweat gland carcinomas were examined for the distribution of cytokeratin (CK) polypeptides using immunohistochemical techniques on paraffin-embedded sections. All the tumour specimens reacted with monoclonal antibodies to CK 7, CK 8, CK 18 and CK 19. Antibodies to CK 1, CK 1/2/10/14, CK 1/5/10/11, CK 13, CK 14 and CK 20 did not stain any of the carcinomas. The results add additional support to the notion that mucinous sweat gland carcinoma represents a tumour histogenetically related to the eccrine secretory coil. Furthermore, the absence of CK 20 might significantly contribute to the differentiation of this tumour from cutaneous metastases from gastrointestinal carcinomas.

Distribution of cytokeratin polypeptides in syringomas. An immunohistochemical study on paraffin-embedded material.

The distribution of cytokeratin (CK) polypeptides expressed in syringomas (12 cases) was compared with that in normal eccrine sweat ducts using immunohistochemical techniques on paraffin-embedded tissue. Intradermal and intraepidermal segments of the eccrine duct showed reactivity with an antibody to CK1/5/10/11 in all cell layers, whereas CK19 expression was restricted to the luminal cell layer. CK14 was expressed in all cells of the eccrine duct except for the peripheral cells of the intraepidermal duct. Expression of CK5/6 was seen in the basal cells of the dermal duct and of the lower intraepidermal duct (sweat duct ridge) exclusively. Reactivity with an antibody to CK1 was found in the intermediate cells of the uppermost part of the eccrine dermal duct. In addition, this antibody gave a strong staining of the peripheral cells of the intraepidermal duct, leaving basal cells of the sweat duct ridge and luminal cells unstained. In syringoma, CK distribution was essentially comparable with that found in the uppermost part of the dermal duct and in the sweat duct ridge. Namely, ductal luminal cells expressed CK1/5/10/11, CK19, and variably CK14. Intermediate cells of ductal structures and solid nests were homogeneously stained by antibodies to CK1 and CK1/5/10/11, whereas CK14 was expressed heterogeneously. The basal or outermost layer of ductal structures and solid nests was reactive with antibodies to CK1/5/10/11, CK5/6, and CK14. With regard to CK expression, the results indicate that syringoma represents a tumor differentiating toward both the uppermost part of the dermal duct and the lower intraepidermal duct (sweat duct ridge) of the eccrine sweat gland.

Eccrine syringofibroadenoma: a case report with analysis of cytokeratin expression.

A 56-year-old man presented with a 30-year history of a slowly enlarging lesion on the sole of his right foot. A biopsy showed an anastomosing network of small cuboidal cells with the formation of occasional sweat ductal lumina and a marked fibrovascular stroma. The histological findings were interpreted as consistent with the diagnosis of an eccrine syringofibroadenoma. Using immunohistochemistry all the tumour cells were positively stained by the pan-cytokeratin antibody Lu-5 and an antibody to the cytokeratins 1/5/10/11. In addition the luminal ductal cells expressed cytokeratin 19 and CEA. Tumour cells were negative for cytokeratins 1, 7, 8, 13 and 18 and did not express vimentin and GCDFP-15. The results indicate that the eccrine syringofibroadenoma is differentiated towards the dermal eccrine duct.

A monoclonal antibody that recognizes a formalin-resistant epitope on the p 24 core protein of HIV-1.

A murine monoclonal antibody (called H-11) that binds to the p 24 core protein of HIV-1 was characterized by radioimmuno-precipitation, immunofluorescence, western blot assays, immunocytochemistry and immunohistochemistry. This antibody was found to be especially suited for demonstrating the presence of HIV-1 in formalin-fixed, paraffin-embedded tissues.

[Eccrine poroma. A clinico-pathologic and immunohistologic study with special reference to tumor cell differentiation]. / Das ekkrine Porom. Eine klinisch-pathologische und immunhistologische Studie unter besonderer Berücksichtigung der Tumorzelldifferenzierung.

In this study 15 eccrine poromas were analysed clinically, histologically and immunohistologically. They were all solitary lesions, showing a predilection for the head and neck. In none of the tumours was diagnosis possible on the basis of clinical examination. Histomorphologically, eccrine poromas were characterized by aggregations of neoplastic cells continuous with the epidermis. The neoplasms consisted of two cell types, poroid and cuticular. Poroid cells predominated, while cuticular cells were only found in small foci, sometimes showing tubular differentiation. Immunohistologically, most of the tumour cells showed a cytokeratin pattern (CK1, 5, 10, 11+, CK1-19+) favouring differentiation toward the abluminal cell of the dermal eccrine duct rather than toward the abluminal cell of the intraepidermal segment of the eccrine duct. Only a small proportion of cells revealed the immunohistological features of the abluminal cell of the intraepidermal duct (CK1+, CK1, 5, 10, 11+, CK1-19+). In addition, cuticular cells showed differentiation toward the luminal cell of the eccrine duct (CK19+, CK1, 5, 10, 11+, CK1-19+). Simple-type cytokeratins such as CK7 and CK18 were not expressed. In conclusion, our findings favour the hypothesis that ascribes the origin of eccrine poroma to a pluripotential stem cell of the transitional zone between the dermal and the intraepidermal segments of the eccrine duct.

Unexpected immunoreactivities of intermediate filament antibodies in human brain and brain tumors.

Immunoreactivities of 35 different monoclonal antibodies (MAbs) that detect intermediate filaments were studied systematically on serial cryostat sections of 14 well-defined human gliomas (five astrocytomas, three oligodendrogliomas, six glioblastomas) and on normal brain. Glial fibrillary acidic protein (GFAP), vimentin, desmin, neurofilaments, and broad-specificity keratin MAbs, as well as MAbs that recognize several or only single keratin polypeptides, were used. Unexpected reactivities were surprisingly frequent. As these may lead to diagnostic confusion and misinterpretation on this material, the authors investigated these phenomena more thoroughly. Four major sources of artifactual staining were found: 1) positive staining attributable to the rabbit gamma G immunoglobulins used in the alkaline phosphatase anti-alkaline phosphatase technique; 2) certain desmin and keratin MAbs cross-reacted with astrocytic glia and with other brain-specific epitopes; 3) technical difficulties; 4) some MAbs directed against neurofilaments and keratins showed unexpected reactivities only on individual anaplastic gliomas. The implications of these findings for intermediate filament typing of neuropathologic material are discussed.

Cytokeratin expression and vimentin content in large cell anaplastic lymphomas and other non-Hodgkin's lymphomas.

The immunophenotypes of 74 malignant lymphomas (9 Hodgkin's disease, 19 low-grade B-cell, 20 high-grade B-cell, 8 T-cell, and 18 large cell anaplastic lymphomas [LCAL]) have been characterized with antibodies against leucocyte differentiation antigens, keratin, and vimentin. All the non-LCAL were CD45 positive and keratin negative. The LCALs had a more varied immunophenotype, with CD45 present only in 11 of 18 cases and keratin present in 5 of 18 of these rare lymphomas. The lymphoid origin of these latter cases was proven by gene rearrangement studies. All LCALs were CD30+, and, where tested, vimentin positive. Of four different vimentin monoclonal antibodies tested, V9 and MVI stained the highest number of lymphomas. Positive staining of tumor cells was seen in 61 of 71 cases. Vimentin-negative cases included Burkitt's as well as some follicular lymphomas.