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Biomedical subjects

L Chyczewski

Publications and source records attributed to L Chyczewski.

At least 19 recordsLinked to original sources
PubMed2006

Acute lymphoblastic leukaemia cells express CCR7 but not higher amounts of IL-10 after CD40 ligation.

OBJECTIVE: Production of cytokines that support T-cell activation and proliferation and migration to lymph nodes is one of the most important terms of cancer vaccine development. In previous studies we and others used CD40 ligation to obtain higher expression of co-stimulatory and adhesion molecules on leukaemic cells from children with acute lymphoblastic leukaemia (ALL). This time we assess the cytokine and chemokine gene expression profile in CD40-stimulated ALL cells. MATERIAL AND METHODS: Malignant cells from 25 children with BCP-ALL were stimulated (or not) with huCD40LT and rIL-4 for 96 h. Eleven different molecule, cytokine and chemokine mRNAs levels (CCR7, IL-23, TGF-beta-IP, IFN-gamma, IL-10, CD1a, CD40, CD54, CD80, CD83, CD86) were determined using the real-time PCR technique with TaqMan chemistry using ready-to-use low-density arrays for gene expression by Applied Biosystems. RESULTS: 1) Increases in mRNA levels for CD40, CD54 and CD80 after CD40L and IL-4 stimulation were observed, 2) CCR7 mRNA expression was higher after CD40 ligation than before the culture (p = 0.002), 3) IL-10 mRNA expression was higher after the culture with medium than before the culture (p = 0.01). CONCLUSIONS: The results show that leukaemia-derived dendritic cells obtained with CD40 ligation express CCR7 - chemokine is involved in migration to lymph nodes and does not produce higher amounts of IL-10, a potent immunosuppressive cytokine. Our preclinical findings could be used in the design of immunotherapy trials for the treatment of children with ALL.

CD40 Antigens↗
PubMed2006

The -675 4G/5G plasminogen activator inhibitor-1 promoter polymorphism in house dust mite-sensitive allergic asthma patients.

BACKGROUND: Plasminogen activator inhibitor (PAI)-1 plays an important role in inflammation and tissue remodeling. Recently, the -675 4G/5G PAI-1 polymorphism has been linked with asthma. OBJECTIVE: This study was undertaken to evaluate associations of the -675 4G/5G PAI-1 polymorphism with functional and immunologic parameters of newly diagnosed house dust mite-sensitive allergic asthmatics (HDM-AAs). METHODS: This study was performed in 127 HDM-AAs, who responded with at least 20% fall of forced expiratory volume during the first second (FEV(1)) to a bronchial challenge with Dermatophagoides pteronyssinus allergen and during the follow up observation fulfilled GINA criteria for mild-moderate asthma. About 89 healthy control nonatopic subjects (HCs) were used as controls. RESULTS: The frequency of 4G allele was greater in HDM-AAs (0.69; 95% CI: 0.62-0.76) than in HCs (0.55; 95% CI: 0.48-0.62; P = 0.0034). The PAI-1 polymorphism was associated with an increased risk of HDM-AA; adjusted for sex and age odds ratio was 2.62; (95% CI: 1.16-5.92) for 4G/5G genotype and 3.48 (95% CI: 1.54-7.89) for 4G/4G genotype compared with 5G/5G genotype. Total serum immunoglobulin E (tsIgE) level in 4G/4G homozygotes (557 +/- 343 kU/l) was significantly greater than in 5G/5G homozygotes (241 +/- 288 kU/l; P < 0.001). Both nonspecific and allergen-specific bronchial reactivities were greater in 4G/4G homozygotes than in 5G/5G homozygotes. 4G/4G genotype was associated with significantly higher morning plasma PAI-1 concentration in HDM-AAs and HCs. Morning plasma PAI-1 concentration correlated significantly with log(PC20) (r = -0.39; P = 0.0001) and with log(tsIgE) (r = 0.247; P = 0.0117). CONCLUSION: These results support the hypothesis linking the 4G/4G PAI-1 genotype with an increased risk of allergic asthma, bronchial hyperreactivity, and increased tsIgE levels.

Adult↗
PubMed2006

Low expression of costimulatory molecules and mRNA for cytokines are important mechanisms of immunosuppression in acute lymphoblastic leukemia in children?

UNLABELLED: Mechanisms leading blasts of acute lymphoblastic leukemia to escape from immune surveillance are still unknown. Only few reports showed that ALL cells are inefficient antigen presenting cells. The aim of the study was to assess expression of critical costimulatory/adhesion molecules and mRNA for main pro- and anti-inflammatory cytokines in ALL cells. Children with B-cell precursor ALL (n=20) were prospectively enrolled into the study. Expression of costimulatory/adhesion molecules (CD1a, CD11c, CD40, CD54, CD80, CD83, CD86, CD123, HLA class I and II) was assessed by flow cytometry and mRNA for cytokines (IFN-gamma, IL-10, IL-4, TGF-beta) - with real-time PCR. RESULTS: 1) high expression was observed for HLA I and II class, moderate for CD40, CD83, CD86 and low or no expression for CD80, CD54, CD1a, CD11c and CD123; 2) we found expression of mRNA for IFN-gamma, IL-10, IL-4 and TGF-beta in blasts cells (but not in all specimens). We noted relatively lower expression of all assessed cytokines comparing to T-cells obtained from healthy donors but interestingly expression for IL-10 was higher in normal B-cells than in blast cells, and IFN-gamma and IL-4 were not found in normal B-cells. In summary we suggest that ALL-blasts present low expression of costimulatory/adhesion molecules and mRNA for cytokines and this probably contribute to the absence of host T- cells stimulation to immune response.

Antigens, CD↗
PubMed2005

Use of synaptophysin immunohistochemistry in intestinal motility disorders.

The purpose of this study was to describe synaptophysin (SY) immunoreactivity in colonic specimens from patients with Hirschsprung's disease (HD), chronic constipation (CC), or anal atresia (AA). This membrane protein is specific for the synaptic vesicles in the central and peripheral nervous system and responsible for neurotransmission. Biopsy specimens of the intestinal wall were obtained from 18 patients (age range, 2 days to 7 years). Immunohistochemistry was performed using rabbit anti-human antibodies specific for synaptophysin (DAKO). In the ganglionic colon of HD patients and others the immunoreactivity of SY-positive synapses was abundantly present in the smooth muscle layers. Distinct immunoreactivity showed ganglion cells and nerve fibers inside circular and longitudinal muscle layers. In some non-HD patients' colonic specimens SY-positive synapses were present in the muscularis mucosae. In the aganglionic colonic segment of HD-patients no immunoreactivity of synapses and ganglions was seen. In the transition zone, where ganglion cells appeared sporadically, synapses were very rarely present. In two patients from the CC group the amount of visualized synapses was clearly smaller and the concentration of ganglion cells within ganglions in these cases was much lower than usual (but still within normal ranges). In the AA group in the distal part of the atretic rectum (at the place where the fistula was cut) SY-positive synapses were present in smooth muscle layers and small dysplastic ganglions were seen in the submucosal and muscular region, but not in large numbers. These patients had a normal distribution of ganglion cells and synapses at the place of colostomy. Synaptophysin immunohistochemistry is an indirect labeling method with a high detection rate for intestinal ganglion cells by demonstrating their synapses. Changed intestinal distributions of SY-positive synaptic vesicles usually accompany colonic ganglion cell disorders. The pattern of SY-positive synapses distribution in circular and longitudinal colonic muscles and intermuscular ganglions can reflect functional disturbances of large bowel motility and could be helpful in the description of the innervation status of colonic specimens in HD patients.

Anus, Imperforate↗
PubMed2005

HBV-DNA and sFas, sFasL concentrations in serum of healthy HBsAg carriers.

PURPOSE: Increased HBV-DNA concentration is a prognostic factor of disease progression in chronic hepatitis B patients. Moreover, active hepatic inflammation during HBV replication influences apoptosis intensification. The aim of this study was to estimate occurrence of HBV replication among carriers of HBsAg. Furthermore, we analysed the correlation between HBV replication and HBeAg or anti-HBe presence as well as known apoptosis indicators--sFas and sFasL concentration. MATERIAL AND METHODS: The study included 34 HBV infected patients, aged 20-43 yrs defined as HBsAg healthy carriers. HBV-DNA was extracted from patients' serum using two different DNA isolation kits: the QIAamp DNA Mini Kit (QIAGEN Ltd, USA) and the Gene Elute Mammalian Genomic DNA Miniprep Kit (Sigma, USA). HBV-DNA concentration in serum was measured by RT-PCR based on TaqMan Universal Master Mix (Applied Biosystems). The detection limit of this system was as few as 10 HBV-DNA copies/mL of serum. HBV-DNA concentration was calculated from a linear standard curve obtained between 10 and 10(8) DNA copies/reaction. HBeAg and anti-HBe in serum were detected by MEIA method (ABBOTT, Germany). The concentration of sFas and sFasL in serum was-estimated by ELISA method (Bender MedSystems, Austria). RESULTS: HBV active replication was detected in 79% HBsAg carriers. The HBV-DNA levels exceeding 10(5) copies/mL were observed in 64% patients. Among HBsAg carriers presenting HBeAg, HBV replication occurred more often and was more intensify than in HBsAg carriers presenting anti-HBe antibodies. The sFasL occurrence in serum of 56% HBsAg carriers shows an active apoptosis, independent from ALT and AST activity within normal ranges.

Adult↗
PubMed2004

Clinical usefulness of K-RAS mutation detection in colorectal cancer and in surgical margins of the colon.

The incidence of K-RAS gene mutations in tumour and surgical margins was investigated in 63 patients with adenocarcinomas of varied clinical stage and histological grade. Point mutations of codon 12 K-RAS gene were detected, using the PCR-RFPL technique in cancer tissue in 23 patients (36.5%) and in colon margin mucosa in 1 patient (3.7%), out of 27 examined subjects. No significant correlations were found between the mutations and clinical features. Tumours, located in the left colon, and mucinous neoplasms displayed a higher incidence of mutations. No correlation was observed with either Dukes or TMN clinical advancement.

Colon↗
PubMed2004

Presence of MHC-I in rat glioma cells expressing antisense IGF-I-receptor RNA.

The supposed immunogenic character of glioma cells transfected with antisense IGF-I-Receptor (IGF-I-R) expression vector was tested for the presence of MHC-I currently present in cells of IGF-I antisense type. C6 rat glioma cell line was comparatively transfected in vitro with IGF I antisense (pMT-Anti-IGF I) or IGF I Receptor antisense (pMT-Anti-IGF I R) expression vectors. The wild and transfected cells were examined for the presence of IGF-I and MHC-I molecules. Using RT PCR technique, the transfected "antisens" cells showed total inhibition of IGF-I. The both transfected cultures of IGF-I and of IGF-I-R type were positively stained for MHC-I. Moreover "antisense IGF-I-R" cells as compared to "IGF-I antisense" cells showed slightly higher expression of MHC-I. The transfected cells showed also the feature of apoptosis in 60% of cells. The immunogenicity of IGF-I-R antisense glioma cells is related to MHC-I presence; therefore both approaches of antisense IGF-I and of antisense IGF-I-R could be use in paralel for cellular therapy of glioblastoma.

Animals↗
PubMed2004

Presences of human papillomavirus DNA (HPV) and immunohistochemical p53 overexpression in papillomas of oral cavity.

HPV belongs to a family of tumorigenic viruses and induces cutaneous and mucosal proliferation of epithelial cells (papillomas, condylomas, warts). The abnormality accumulation of p53 protein appears to be a common step in the development of many human cancers and has been frequently reports in human cancers including head and neck. The aim of this study was to examine HPV infection incidents and p53 alteration in oral papillomas. HPV was detected by PCR, p53 accumulation by immunohistochemical detection. The overexpression of p53 was revealed in 20 cases (55%) out of 36 papillomas. We found p53 overexpression in 8 out of 13 HPV positive papillomas (61.5%) and in 12 out of 23 HPV negative papillomas (52%). Our findings indicate that HPV infection and/or changes in p53 protein coexist in oral cavity papillomas.

DNA, Viral↗
PubMed2004

mRNA for chosen pro- and anti-inflammatory cytokines in T-lymphocytes in paediatric leukemias and lymphomas--a preliminary report.

We assessed mRNA for chosen pro- and anti-inflammatory cytokines in T-lymphocytes of peripheral blood in paediatric patients with leukemias and lymphomas. Levels of four different cytokine mRNAs (IFN-gamma, IL-10, IL-4, TGF-beta) were determined by the real-time PCR technique. In the whole examined group, at the time of diagnosis, we noted lower amounts of mRNA for TGF-beta1, comparing to respective values in the control patients. In the ALL group, we observed the following: 1) at the time of diagnosis: lower amounts of mRNA for IL-4 and for TGF-beta1, comparing to respective values in the control group; 2) lower amounts of mRNA for IL-10 after remission induction, comparing to the time of diagnosis. In our opinion, "immunedysregulation" in lymphoproliferative diseases in children is not caused by IFN-gamma deficiency. The deficit of anti-inflammatory cytokines, i.e., IL-4, TGF-beta, with higher amounts of IL-10, suggests their role in cancer development.

CD4-Positive T-Lymphocytes↗
PubMed2004

Attempts to detect Helicobacter pylori in atherosclerotic plaques.

Cardiovascular and cerebrovascular diseases are regarded to be the main causes of mortality in developed countries, atherosclerosis being at their pathological base. During the recent years, attention was paid to the role of bacterial infections, including Helicobacter pylori, in the process of atherogenesis and coronary heart disease development. The aim of the study was an evaluation of H. pylori presence--by means of PCR technique--in atherosclerotic changes, obtained by endarterectomy, performed during coronary artery bypass grafting (CABG). In the analysed group of patients, the following risk factors were found: hyperlipidaemia, smoking, hypertension, obesity, diabetes mellitus, cardiac infarction. No DNA of the bacteria was traced in any of the patients.

Adult↗
PubMed2003

Therapy increases poly-ADP-ribose and p53-Ser392-P levels in recurrent squamous cell lung cancer.

p53 protein is a critical regulator of the cell cycle and apoptosis and its levels and functions change in response to many stimuli. To assess whether the cytotoxic drugs induce DNA changes, affect phosphorylation and stability of p53 protein, we determined poly-ADP-ribose levels, the expression of p53 protein and its carboxyl-terminal Ser-392 phosphate levels in fiberoptic bronchoscopy biopsy samples taken from patients suffering from recurrent squamous cell lung cancer before and after radiotherapy and chemotherapy. All 14 patients included in this study were in IA-IIIA clinical stage prior to surgery. Radiation/chemotherapy decreased G2/M cell numbers but increased S-phase cells by almost 50% compared to ploidy status before therapy, while median p53 expression was doubled (109% increase). p53 phosphorylated on Ser-392 was also increased by approximately 70% in patients treated with radiotherapy and with chemotherapy and correlated with elevated poly-ADP-ribose levels. Our data suggest that apart from changes in p53 quantity, posttranslational phosphorylation/dephosphorylation-mediated alterations may play an important role in neoplastic cell proliferation as well as in antiproliferative activity of drugs inducing DNA damage and apoptosis.

Aged↗
PubMed2003

IGF-I triple helix gene therapy of rat and human gliomas.

PURPOSE: IGF-I anti-gene technology was applied in treatment of rat and human gliomas using IGF-I triple helix approach. MATERIAL AND METHODS: CNS-1 rat glioma cell and primary human glioblastoma cell lines established from surgically removed glioblastomas multiforme were transfected in vitro with IGF-I antisense (pMT-Anti-IGF-I) or IGF-I triple helix (pMT-AG-TH) expression vectors. The transfected cells were examined for immunogenicity (immunocytochemistry and flow cytometry analysis) and apoptosis phenomena (electron microscopy). 3 x 10(6) transfected cells were inoculated subcutaneously either into transgenic Lewis rats or in patients with glioblastoma. The peripheral blood lymphocytes (PBL) derived from "vaccinated" patients were immunophenotyped for the set of CD antigens (CD4, CD8 etc). RESULTS: Using immunocytochemistry and Northern blot techniques, the transfected "antisense" and "triple-helix" cells showed total inhibition of IGF. Transfected cultures were positively stained either for both MHC-I and B7 antigens--60% of cloned lines, or for MHC-I only--40% of cloned lines. Moreover "triple helix" cells as compared to "antisense" cells showed slightly higher expression of MHC-I or B7. Transfected cells also showed the feature of apoptosis in 60%-70% of cells. In in vivo experiments with rats bearing tumors, the injection of "triple helix" cells expressing both MHC-I and B7 interrupted tumor growth in 80% of cases. In contrast, transfected cells expressing only MHC-I stopped development in 30% of tumors. In five patients with surgically resected glioblastoma who were inoculated with "triple helix" cells, PBL showed an increased percentage of CD4 + CD25+ and CD8 + CD11b-cells, following two vaccinations. CONCLUSIONS: The anti-tumor effectiveness of IGF-I anti-gene technology may be related to both MHC-I and B7 expression in cells used for therapy. The IGF-I antigene therapy of human glioblastoma multiforme increases immune response of treated patients.

Animals↗
PubMed2002

Suitability of selected markers for identification of elements of the Intestinal Nervous System (INS).

The possibility of identifying and characterising elements of the enteric nervous system based on their contents of cathepsin D, chromogranin A, neuronal specific enolase and S-100 protein was studied in colorectal specimens (operative full-thickness, seromuscular and mucosomuscular biopsies) obtained from 15 children, aged 2 days to 10 years. Nine patients suffered from Hirschsprung's disease, and two from chronic constipation. Four neonates with imperforate anus or meconium ileus composed the control group. All markers were identified immunohistochemically by antibodies against human antigens with appropriate detection methods. Chromogranin A staining was not always adequate to identify all neuronal cell bodies and other nervous elements. However, it proved superior to the other methods in the depiction of neuroendocrine cells in the intestinal mucosa. Cathepsin D antibodies stained normal and abnormal neural cells with different intensity; nerve fibres were not stained. This marker did not allow an unequivocal differentiation of ganglion cells from macrophages within the submucosa; the latter exhibited exceptionally strong marking and in some cases represented the predominant elements in this area. Neuronal specific enolase was distinctly expressed in nerve cells and fibres of the intestinal wall. Atrophic and hypoplastic features could be identified, suggesting that this method may give some insight into functional aspects. Continuous connections between ganglions were also observed. S-100 protein antibodies resulted in a negative image of unstained ganglion cells surrounded by extensively marked Schwann cells and neural fibres. With respect to clinical application, all these markers may provide supplementary information for the differential diagnosis of intestinal motility disorders.

Biomarkers↗
PubMed2002

Preliminary evaluation of mast cells and angiogenesis processes in experimental fibrosarcoma.

The aim of the study was the evaluation of angiogenesis processes in fibrosarcoma induced by 3-methylcholanthrene (3-Mc) in reference to the number of mast cells (MCs). 76 male Wistar rats were divided into 4 groups: two experimental (E) groups--after injection of 0.2 mg 3-Mc dissolved in olive oil (0.25ml), and 2 control (C) groups. In E1 group, 52 rats were killed after development of the fibrosarcoma; E(2)--10 rats were killed before development of the tumor; C(1)-8 rats received 0.25ml olive oil; C(2)--8 rats received no treatment. Tissue material was fixed in buffered formalin or Carnoy's and Bouin's fluid. Paraffin sections were stained with H+E and Azan methods, and with alcian blue-saphranine and toluidine blue. Immunohistochemical reactions detecting tryptase in MCs were also performed. Angiogenic objects (microvessels and single endothelial cells) were recognized using antibodies against factor VIII (vWF), P selectin (CD-62P), and CD-90. We found a distinct relationship between intensification of neoangiogenesis at the tumor periphery and increased number of MCs.

Animals↗
PubMed2002

Angiogenesis and Bcl-2 protein expression in patients with endometrial carcinoma.

Considering the particular importance of angiogenesis and tumor suppressor genes expression in solid tumors, angiogenesis and Bcl-2 protein expression were evaluated in order to specify their role in the biology of endometrial carcinoma. Clinical material comprised 66 patients (postmenopausal, aged 52 to 76 years) with endometrial adenocarcinoma. For evaluation of angiogenesis immunohistochemical method was applied using DAKO EPOS Anti-Human Von Willebrand Factor/HRP antibodies. Morphometric method was applied to count angiogenic points (microvessels + single endothelial cells), using a light microscope with morphometric appliance. Angiogenic points density (APD) was defined as the density of AP per square mm. Immunohistochemical staining for Bcl-2 cytosomic protein expression was performed using MoAb124 (dilution 1:80, Dako A/S, Denmark) monoclonal antibodies. The percentage of 10% positive cells was considered as Bcl-2 positive tissue expression. Positive cytoplasmic reaction of Bcl-2 in 51.3% of patients with Stage I endometrial cancer, and in 23.8% and 0% of patients with II and III FIGO stage, respectively, was observed. No relationship between Bcl-2 tissue cytoplasmatic expression and tumor grade was found. However, an inverse correlation between cytoplasmatic expression of Bcl-2 and FIGO stage was observed. The APD (angiogenic points density) was increasing with the clinical (FIGO) stage of endometrial cancer, but it was not observed in the case of tumor histologic grade. Bcl-2 expression and angiogenesis may be a useful parameter in evaluation of the biology of endometrial adenocarcinoma as the study conducted showed the influence of Bcl-2 protein expression upon angiogenesis.

Adenocarcinoma↗