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The purpose of our study was to evaluate the microsatellite instability (MSI) at selected loci with known involvement in the oncogenesis of chronic B-cell lymphocytic leukaemia (B-CLL). DNA from B cells (tumour cells) and from T cells (normal controls) of 27 samples of 26 patients with previously untreated B-CLL was extracted. Microsatellite instability in six microsatellite markers was tested using GeneScan Analysis Software. The rate of replication errors positive phenotype (RER+) was determined (MSI in more than 30% of examined loci). RER+ was found in four out of 27 patients (14.8%). A larger proportion of patients with stage C B-CLL exhibited RER+ than those with stage A or B (P < 0.05). A higher prevalence of RER+ was demonstrated in a subgroup of patients with additional malignancies (three out of eight patients) in comparison with patients with B-CLL alone (1/19) (P = 0.031). In conclusion, our study demonstrated that MSI might have a more prominent role in pathogenesis of B-CLL than reported to date. This may result from a selection of microsatellite markers adjacent to chromosomal loci, which are involved in B-cell malignancies, and using GeneScan Analysis Software, which is most modern and precise method of microsatellite analysis. [ABSTRACT FROM AUTHOR]

Formaldehyde (FA) is a widely produced industrial chemical. Sufficient evidence exists to consider FA as an animal carcinogen. In humans the evidence is not conclusive. DNA-protein crosslinks (DPC) may be one of the early lesions in the carcinogenesis process in cells following exposures to carcinogens. It has been shown in in vitro tests that FA can form DPC. We examined the amount of DPC formation in human white blood cells exposed to FA in vitro and in white blood cells taken from 12 workers exposed to FA and eight controls. We found a significant difference (P = 0.03) in the amount of DPC among exposed (mean ± SD 28 ± 5%, minimum 21%, maximum 38%) than among the unexposed controls (mean ± SD 22 ± 6%, minimum 16%, maximum 32%). Of the 12 exposed workers, four (33%) showed crosslink values above the upper range of controls. We also found a linear relationship between years of exposure and the amount of DPC. We conclude that our data indicate a possible mechanism of FA carcinogenicity in humans and that DPC can be used as a method for biological monitoring of exposure to FA. [ABSTRACT FROM PUBLISHER]

Introduction: Atopic dermatitis (AD) is a chronic inflammatory skin disease associated with cutaneous hyper-reactivity to environmental triggers, and is characterized by pruritic eczematous skin lesions. Recent insights into the pathophysiology of AD point to the important role that abnormal barrier permeability plays, which leads to compromised hydration, common micro-organismal colonization and immune dysregulation. Current treatment strategies for AD, such as topical corticosteroids and moisturizers, control the disease symptom's severity and duration. KAM-3008 is a novel, barrier-based, steroid-free, topical cream indicated for the relief of symptoms associated with AD. In addition to moisturizing and emollient ingredients, several herbal extracts have been incorporated into the formulation to enhance its anti-allergic and anti-inflammatory properties., Methods: In this study, the safety and efficacy of KAM-3008 body cream in relieving the symptoms of AD in adult subjects after 7, 14 and 21 days of treatment were investigated using the transepidermal water loss (TEWL), skin hydration and scoring atopic dermatitis (SCORAD) measurements., Results and Conclusions: Based on both the quantitative TEWL and qualitative SCORAD assessments, we found that KAM-3008 body cream is a safe, well-tolerated and efficacious stand-alone treatment for the relief of AD symptoms.

Development of multiple tumors of different histopathologic types may suggest a profound generalized genetic defect, such as malfunction of DNA mismatch repair (MMR) mechanism. Defects in this mechanism are best reflected in microsatellite instability (MSI). We aimed to determine the role of MSI in a group of patients with dual malignancies and compared the data with that of patients with a single malignancy. Fifty patients were enrolled in the study, of whom 16 patients developed both solid and hematologic nonfamilial malignancies, 18 patients developed a single matched hematologic malignancy, and 16 a single matched solid malignancy. Five microsatellite markers were replicated by polymerase chain reaction (PCR) after DNA extraction from paraffin-embedded tissue blocks and analyzed by the GeneScan Analysis Software. The MSI-high phenotype was defined as instability in at least 40% of the examined loci. A higher prevalence of MSI-high phenotype was found in patients with dual malignancies (31.3%) compared with patients with single hematologic (5.6%) or solid malignancy (6.3%) (P = 0.0498 and 0.07, respectively). In conclusion, defects in DNA MMR mechanism may have an important role in the development of multiple sporadic nonfamilial malignancies.

The role of microsatellite instability and defects in DNA mismatch repair mechanism in the pathogenesis of gastric lymphoma of mucosa-associated lymphoid tissue (MALT) type is still controversial, as both negative and positive findings have been reported. This may be explained mainly by arbitrary selection of the tested loci, the use of various techniques of microsatellite instability analysis and by different definitions of replication error positive phenotype. The aim of our study was to evaluate the instability at selected microsatellite markers using the GeneScan Analysis Software. DNA from paraffin-embedded tissue blocks of 13 previously untreated patients with localized gastric MALT lymphoma was extracted. Five microsatellite markers, which are located in hMSH2, hMLH1, P16, APC and MLL loci, were selected from the genetic database. We found genetic instability in tumors of 9/13 patients with gastric MALT lymphoma (69%). Seven of them had replication-error-positive phenotype (54%). Microsatellite instability was found in 39% of the samples in the MLL locus, 39% in the APC, 46% in the P16, 23% in the hMLH1 and none in the hMSH2. This study demonstrates that microsatellite instability has more prominent role in pathogenesis of gastric MALT lymphoma than reported to date. We suggest that microsatellite instability should be analyzed with markers adjacent to chromosomal loci that are involved in lymphomas. Our findings support the 'Real Common Target genes' theory of high rate of microsatellite instability in specific genes, which are associated with related tumors.

Uncontrolled inflammation is considered to exacerbate the neuronal loss that follows spinal cord trauma. However, controlled inflammation response appears to be beneficial. Skin-coincubated macrophages injected into contused spinal cord of rats resulted in improved motor recovery and reduced spinal cyst formation. The macrophages express elevated levels of cell-surface molecules CD80, CD86, CD54 and MHC-II, markers characteristic of antigen presenting cells (APCs). Additionally, skin-coincubation elevates secretion of interleukin-1 beta (IL-1 beta) and Brain-Derived Neurotrophic Factor (BDNF), and reduces secretion of tumor necrosis factor alpha (TNF-alpha). We propose that macrophages activated by skin-coincubation bolster neuroprotective immune activity in the spinal cord, making the environment less cytotoxic and less hostile to axonal regeneration.

Objective: Clonal B-lymphocytes of chronic lymphocytic leukemia (B-CLL) are characterized by decreased sensitivity to programmed cell death and, therefore, they accumulate in vivo. However, these malignant cells die rapidly in vitro. In the current study we concentrated on the contribution of autologous serum (AS) and lymphocyte subsets to the survival of the malignant cells in vitro., Methods: Mononuclear cells from the peripheral blood of 26 CLL patients and 24 controls were incubated overnight in the presence or absence of AS and heat-inactivated AS (HI-AS) or fetal calf serum (FCS). Also, isolated B cells were incubated at different concentrations in the presence of AS and/or isolated T cells. The level of apoptosis of CD19+ cells was measured by flow cytometry., Results: Spontaneous apoptosis of unfractionated B-CLL cells incubated with AS, FCS or without serum was significantly lower than the rate of B-cell death in the control group, in similar culture conditions. AS had an antiapoptotic effect on unfractionated B-CLL cells when compared with FCS. The rate of apoptosis of B-CLL cells was directly associated with stage. HI of AS had a variable effect, which was related to the stage of the disease. High concentrations of B cells and the addition of autologous T cells reduced the rate of apoptosis when incubated without serum. The antiapoptotic effect of T cells was most prominent in progressive stages., Conclusions: B-CLL cells exhibit decreased spontaneous apoptosis, which is partially prevented by humoral (AS) and cellular (T cells and B-CLL cells) factors. The equilibrium between apoptotic and antiapoptotic factors changes with disease progression.

Hepatocellular carcinoma (HCC) is a very common and highly malignant tumor, associated mainly with chronic viral hepatitis, cirrhosis of any cause, aflatoxin exposure and ethanol consumption. The aim of this study was to map genomic aberrations in HCC by a recently developed technique: comparative genomic hybridization (CGH). We applied CGH on 17 liver specimens, of which seven were HCCs. The rest were benign liver tumors, cirrhotic and normal livers, and other liver malignancies. Our study included mainly large tumors (mean size 10.5 cm) unrelated to viral hepatitis or cirrhosis. Our CGH analysis detected genomic imbalances in 42% of HCCs. The common aberrations included DNA gains of 1q, 9p, and 8q and DNA losses of 17p, 13q, 9q, 4q, and 11q. Also, we detected trisomies 8, 9, 18 and 21, which have not been reported previously. Gains and losses of DNA found in this study probably involve oncogenes and tumor suppressor genes that play a role in the puzzle of hepatocarcinogenesis. This study also suggests a possible link between the size of the tumor and the burden of genetic changes.

Polycythemia vera (PV) and essential thrombocytosis (ET) are clonal chronic myeloproliferative disorders originating from a multipotent stem cell. Bone marrow examinations reveal chromosomal abnormalities in 15-43% of PV patients and 5% of ET patients, but no specific recurring abnormality has been found to date. We aimed to find cytogenetic aberrations in PV and ET by comparative genomic hybridization (CGH), a relatively new molecular cytogenetic technique. In this study, CGH analysis was performed on peripheral blood leukocytes of 12 PV patients and 8 ET patients. One patient (8.3%) with PV had an abnormal karyotype with a deletion in 7q11.2 and one patient with ET (12.5%) had a gain in 18p. Peripheral blood analysis by CGH revealed a low frequency of cytogenetic abnormalities in PV and ET patients. However, using CGH we were able to detect two cytogenetic aberrations that were not reported previously in these disorders.

Aims: Leiomyosarcomas (LMS) are diverse tumours with different biological behaviour. To evaluate the biological nature of intraabdominal and retroperitoneal leiomyosarcomas we retrospectively examined the immunoreactivity of p53, bcl-2 and proliferative activity expressed as Ki-67-labelling index in 43 tumours., Methods: Immunohistochemical staining was performed using a peroxidase-streptavidin method on paraffin-embedded sections using specific anti- p53, anti- bcl-2 and anti Ki-67 monoclonal antibodies., Results: Of 43 tumours, seven were located in the stomach, 11 in the small or large bowel, 12 in the uterus, 11 in the retroperitoneum and two cases in the urinary bladder. Five-year disease-free survival was 46.5%. Twenty-three patients (53.4%) died of the disease. Positive immunoreactivity for p53 and bcl-2 was found in 18 (41.9%) and 26 patients (60.5%), respectively. Positive Ki-67 staining was observed in eight patients (18.6%). Proliferative indices were higher in LMS with high mitotic activity (P=0.004) and high grade (P=0.009). All Ki-67 positive LMS were intermediate or high-grade tumours. Ki-67-labelling index showed inverse relationship to bcl-2 expression. A trend towards higher survival and expression of bcl-2, p53 or Ki-67 was found., Conclusions: Our results demonstrate that p53 and bcl-2 are expressed in a substantial number of intraabdominal and retroperitoneal leiomyosarcomas. In our study, the expression of these biomarkers did not predict patient outcome. Higher Ki-67 labelling indices were found in more biologically aggressive leiomyosarcomas., (Copyright Harcourt Publishers Limited.)

Background and Objectives: p53, c-erbB-2, and tumor microvascular density have been shown to be potential prognostic tools in female breast cancer. Our objective was to assess the significance of these biomarkers as prognostic factors in infiltrating male breast cancer., Methods: A retrospective study of expression of p53, c-erbB-2, and tumor microvascular density was done on a group of 26 male breast cancer patients. Biotin-streptavidin immunohistochemical study with specific anti-p53, anti-c-erbB-2, and anti-CD34 antibodies was carried out on paraffin sections of breast carcinoma. The data of expression of the biomarkers were merged with clinicopathological data such as tumor grade, T class, TNM stage, estrogen receptor status, tumor recurrence, and patient survival., Results: p53 and c-erbB-2 were expressed in 46% and 39% of carcinomas, respectively. No correlation was found between positive immunoreactivity of p53, and tumor grade, size, T class, TNM stage, and survival. Nor was any relation found between tumor size, T class, TNM stage, survival, and c-erbB-2 overexpression. c-erbB-2 overexpression was significantly higher in high grade carcinomas. Estrogen receptor (ER) were positive in 21 out of 26 of tumors (81%). No trends were observed between estrogen receptor status and clinicopathological parameters or survival (data not shown). There was a positive correlation between mean microvascular density (MVD), advanced T class, and survival: higher MVD counts were found in patients with advanced tumors and in those who had tumor relapses or died of metastatic disease., Conclusions: This study suggests that tumor microvascular density may serve as a potential prognostic tool in male breast carcinoma., (Copyright 2000 Wiley-Liss, Inc.)

Ductal carcinoma in situ (DCIS) of the breast, a precursor lesion of invasive breast cancer, is a heterogeneous disease in terms of histomorphologic features and biologic behavior. Our aim was to assess the proliferative activity, expressed as topoisomerase IIalpha (Topo IIalpha) immunoreactivity and c-erbB-2 expression in relation to morphologic features and architectural pattern of DCIS. The study included 26 DCIS, which were reclassified according to the recommendations of Consensus Conference. Topo-IIalpha and c-erbB-2 immunoreactivity were detected on paraffin sections. Topo IIalpha was consistently negative in normal ductal epithelium. Topo IIalpha-labeling index (Topo IIalpha-LI) was 0.7+/-0.6% for grade I, 4.3+/-3.9% for grade II, and 13.4+/-8.9 for grade III lesions (P<.01). For mixed nuclear grade DCIS, Topo IIalpha-LI was 6.8+/-4.8. There was no difference in Topo IIalpha-LI between different architectural patterns in low- and intermediate-grade lesions. In high nuclear grade DCIS, there was a progressive increase in Topo IIalpha-LI from solid toward cribriform and comedo-type DCIS. Positive c-erbB-2 immunoreactivity was found in 46% of DCIS, being highest in DCIS with high nuclear grade (78%) and in lesions with extensive necrosis. Topo IIalpha-LI was significantly higher in c-erbB-2-positive lesions (Topo IIalpha-LI- 12.4+/-8.5) as compared with negative lesions (Topo IIalpha-LI- 3.9+/-4.5, P<. 0001). Overexpression of c-erbB-2 and Topo IIalpha is associated with poorly differentiated lesions. Proliferative activity in individual ducts of DCIS depended primarily on the nuclear grade and was independent of architectural patterns of individual ducts in architecturally heterogeneous lesions., (Copyright 2000 by W.B. Saunders Company.)

Background: Genetic aberrations observed in the large bowel during the neoplastic progression have a cumulative effect and are responsible for the propagation of the multistep malignant process. In the present study we evaluated the immunoreactivity of c-fos, ras, bcl-2 and p53 in aberrant crypt foci (ACF) and minute polyps of the large bowel obtained from patients with colorectal cancer., Methods: ACF and minute polyps were collected from macroscopically normal colonic mucosa. Protein immunoreactivity was detected on parafin sections utilizing the biotin-streptavidin method on 25 hyperplastic, 10 dysplastic ACF, 5 hyperplastic and 10 dysplastic adenomas., Results: 41% of the lesions displayed positive ras immunoreactivity. bcl-2 immunoreactivity was positive in six minute polyps of which five were neoplastic. fos immunoreactivity was detected in five ACF and seven minute polyps, mainly in dysplastic lesions. Two neoplastic polyps were positive for p53 immunoreactivity. Coexpression of two or more oncoproteins was found with increasing frequency in dysplastic versus hyperplastic lesions and in polypoid lesions versus ACF., Conclusion: Abnormal expression and coexpression in oncoproteins can be identified in the earliest stages of colorectal tumorigenesis and may contribute to the progression of selected lesions during ACF-adenoma-carcinoma sequence.

Aberrant crypt foci (ACF) are one of the earliest putative preneoplastic, and in some cases, neoplastic lesions in human colons. These microscopic lesions, identified on methylene blue-stained mucosa with a low-power-magnification microscope, are thought to be closely related to the earliest steps in multistage colonic tumorigenesis. We investigated the distribution pattern and histomorphological features of ACF in 74 patients with sporadic colorectal cancer. The distribution pattern shows a slightly higher prevalence with older age. The prevalence of the ACF in sigmoid colon was significantly higher in patients with colorectal cancer as compared with patients with benign colonic diseases. Also, significantly more ACF were detected in distal parts of the large bowel (descending, sigmoid colon, and rectum) than in proximal parts. Of 42 microdissected lesions, 12 were dysplastic and 30 were hyperplastic foci. The average size of dysplastic lesions was significantly larger than hyperplastic foci. More apoptotic bodies were found in dysplastic lesions. These lesions also showed an upward expansion of proliferative compartment and higher proliferation indices expressed as proliferating cell nuclear antigen-labeling index. Lymphoid follicles were frequently observed in the base of both hyperplastic and dysplastic foci (40% and 66.6%, respectively). The coincidence of lymphoid follicles was 2.5 to 8 times higher than expected. These features may be related to further progression of selected ACF during colorectal tumorigenesis.

Background: One of the first steps in multistage colonic carcinogenesis is increased cell proliferation and an upward shift of the proliferation zone of colonic crypts. In the present study, progression in cell kinetics was followed up at all sequential stages of colonic carcinogenesis, starting with aberrant crypt foci (ACF), the earliest putative preneoplastic lesions, hyperplastic and dysplastic polyps, and invasive carcinomas., Materials and Methods: Colonic tissue and tumor specimens were prospectively obtained from 65 patients treated at our hospital for adenocarcinoma or malignant polyps. For identification of ACFs, dissected mucosal strips obtained from patients with colorectal cancer were stained with 0.1% methylene blue and scanned under dissecting microscope. Paraffin-embedded ACFs and macroscopic lesions were serially sectioned, deparaffinized, and stained with a monoclonal antiproliferating cell nuclear antigen (PCNA) antibody. The PCNA-labelling index (PCNA-LI), expressed as a ratio of positively stained nuclei to total nuclei counted, was calculated separately for basal, middle, and upper colonic crypt compartments. A comparison of the PCNA-LI was made for each compartment in normal mucosa, and hyperplastic and dysplastic lesions., Results: A stepwise increase in the PCNA-LI was observed during neoplastic progression of colonic lesions. The two most important variables of increased cell proliferation, expressed as PCNA-LI per crypt compartment, were the presence of dysplasia and the size of dysplastic lesions., Conclusions: In colorectal carcinogenesis, hyperproliferation with upward expansion of proliferative compartment is a characteristic feature at all stages of malignant progression.

The interaction of KB-V1, a multidrug resistant (MDR) variant of the KB-3-1 human oral carcinoma, with human complement was investigated. KB-V1 cells were found to be more sensitive than KB-3-1 cells to complement-mediated lysis. Detailed analysis of the capacity of KB cells to activate human complement demonstrated that both C3b deposition and formation of the membrane attack complex (MAC) are higher on KB-V1 than on KB-3-1 cells. Furthermore, the MAC formed on KB-V1 cells, but not on KB-3-1 cells, was found to be resistant to trypsin treatment, i.e. more stably inserted into the plasma membrane. Immunofluorescence analysis by flow cytometry showed that KB-V1 cells express less decay-accelerating factor (DAF, CD55) than KB-3-1 cells. Two other complement regulatory proteins, membrane cofactor protein (MCP, CD46) and CD59 are expressed to a similar extent on both KB-V1 and KB-3-1 cells. Treatment of KB-V1 cells with neutralizing anti-P-glycoprotein (P-gp) monoclonal antibodies reduced their sensitivity to complement. In addition, KB-V1 revertants which cease to express P-gp become more resistant to complement. These results indicate that multiple factors, such as reduced expression of DAF, enhanced deposition of C3b and increased binding and stability of the MAC may contribute to the increased complement sensitivity of KB-V1 cells. It is suggested that P-gp is responsible for the complement-sensitive phenotype of KB-V1 cells.

Formaldehyde is classified as a probable human carcinogen. DNA-protein crosslinks (DPCs) and sister chromatid exchanges (SCEs) may represent early lesions in the carcinogenic process. The authors examined the DPCs and SCEs in peripheral-blood lymphocytes of 12 and 13 workers exposed to formaldehyde and eight and 20 unexposed workers, respectively. The amounts of DPCs and SCEs in the exposed and the unexposed differed significantly after adjustment for smoking. There was a linear relationship between years of exposure and the amounts of DPC and SCE. The authors conclude that the data indicate a possible mechanism of carcinogenicity of formaldehyde, and that formaldehyde is mutagenic to humans. These results support the use of DPCs as a biomarker of occupational exposure to formaldehyde and to detect high-risk populations for secondary prevention.

Supernatants were collected from suspensions of MOPC-315 tumor cells harvested from ascitic tumors and kept for 24 hours in culture medium and from cultures of an MOPC-315 tumor-cell kept for a long period of time in vitro. The MOPC-315 supernatants were tested for immunosuppression of mitogenic stimulation of BALB/c spleen cells by ConA or LPS, of allogeneic response of effector BALB/c spleen cells against target C57BL spleen cells, of generation of antibody response against SRBC and of induction of LAK activity. The immunosuppression was marked in all the test systems, was not related to secretion of either C-type particles or of anti-TNP antibodies and was also induced by MOPC-315 tumor cells kept in serum-free medium. It is suggested that release of immunosuppressive factor(s) by MOPC-315 tumor cells might play a role in the mechanism(s) of defence of the tumor against the host.

Leukoregulin (LR) is a lymphokine secreted by human natural killer (NK) cells. Its effect on the susceptibility of K562 human erythroleukemic cells to lysis by antibody and complement was examined. As reported here, treatment of K562 cells with LR for 60 min at 37 degrees C confers on them resistance to complement damage. The LR-induced state of complement resistance is transient and the cells recover within 4-6 h unless a second dose of LR is added. The protective action of LR was observed using both conventional 51Cr-release and trypan blue inclusion assays. The protein synthesis inhibitors puromycin and cycloheximide and the protein kinase inhibitors tamoxifen, polymyxin B and W-7, could each block this action of LR. Fewer membrane attack complexes were found, following complement activation, on LR-treated than control cells. These results suggest that LR increases the capacity of K562 cells to down-regulate complement activation or repair the complement damage, possibly by inducing synthesis of defense proteins and/or activation of protective protein kinases.

This investigation shows that the regulation of heme synthesis in the regenerating rat liver does not differ from the regulation in the normal liver. The heme saturation of tryptophan pyrrolase was found to be low, indicating a reduced concentration of heme in the regulatory heme pool of the regenerating rat liver. As expected, ALAS in the mitochondrial fraction was found to be elevated. It was also shown that ALAS in the regenerating rat liver can be induced by the porphyrinogenic drugs AIA and DDC and that heme reduces its activity. The decrease observed in the activity of cytosolic ALAS might be due to impaired synthesis of the enzyme but does not affect the regulation of the heme biosynthetic pathway.

L-Alanine: 4,5-dioxovalerate transaminase (ADT) was determined in liver homogenates of rats treated by either inducers of porphyrin synthesis or the repressor, hemin. ADT activity was not induced by the porphyrinogenic agents nor reduced by hemin, indicating that ADT probably has no regulatory role in the heme synthesis pathway. The same conclusion was drawn from similar experiments performed in monolayers of chick embryo liver cells.