Cell Membrane-Derived Microvesicles in Systemic Inflammatory Response.

Human body reacts to physical, chemical and biological insults with a complex inflammatory reaction. Crucial components and executors of this response are endothelial cells, platelets, white blood cells, plasmatic coagulation system, and complement. Endothelial injury and inflammation are associated with elevated blood levels of cell membrane-derived microvesicles. Increased concentrations of microvesicles were found in several inflammatory reactions and diseases including acute coronary syndromes, stroke, vasculitis, venous thromboembolism, multiple sclerosis, rheumatoid arthritis, systemic lupus erythematosus, anti-phospholipid antibody syndrome, inflammatory bowel disease, thrombotic thrombocytopenic purpura, viral myocarditis, sepsis, disseminated intravascular coagulation, polytrauma, and burns. Microvesicles can modulate a variety of cellular processes, thereby having an impact on pathogenesis of diseases associated with inflammation. Microvesicles are important mediators and potential biomarkers of systemic inflammation. Measurement of inflammatory cell-derived microvesicles may be utilized in diagnostic algorithms and used for detection and determination of severity in diseases associated with inflammatory responses, as well as for prediction of their outcome. This review focuses on the mechanisms of release of microvesicles in diseases associated with systemic inflammation and their potential role in the regulation of cellular and humoral interactions.

Single-agent cytarabine is insufficient for the treatment of human mantle cell lymphoma in mouse xenograft model.

Mantle cell lymphoma is an aggressive type of B-cell non-Hodgkin lymphoma with adverse prognosis. It was demonstrated that alternation of CHOP and DHAP chemotherapy improved outcome of mantle cell lymphoma patients. However, which components of DHAP, cisplatin, cytarabine, or both, were responsible for the improved outcome remained unclear. To answer this question, antitumor efficacies of equally toxic doses of cytarabine, cisplatin, and three different combinations were compared in vivo using mouse xenograft models of mantle cell lymphoma. We demonstrated that cisplatin, alone or with cytarabine, is significantly superior to single-agent cytarabine in both eliminating lymphoma cells and suppressing their proliferation rate.

Comparison of Plasma Osteopontin Levels between Patients with Borderline Ovarian Tumours and Serous Ovarian Carcinoma.

Osteopontin (OPN) is a novel biomarker of various cancers including ovarian carcinoma. OPN is a promising adjunct to a major biomarker of ovarian cancer, CA125, in diagnosis, differential diagnosis and prognosis. The aim of our study was to measure the plasma level of OPN and CA125 in patients with borderline ovarian tumours (BOTs), serous ovarian carcinoma, and controls to determine its potential role in the differential diagnosis between serous ovarian carcinoma and BOT. The plasma samples of 66 women were analysed using Luminex technology, designed to simultaneously measure multiple specific protein targets. The mean OPN plasma level for the control group was 23.3 ng/ml; for BOT 26.3 ng/ml; and for patients with serous ovarian carcinoma 59.5 ng/ml. Specifically, there was a significant difference between the OPN levels in patients with ovarian carcinoma and BOT (P < 0.001) as well as controls (P < 0.001). There was no difference between the mean levels of OPN in patients with BOT and the control group (P = 0.286). Using the receiver operating characteristic (ROC), we determined the utility of OPN and CA125 to differentiate between BOT and serous ovarian carcinoma. The area under the ROC curve (AUC) for OPN was 0.793 (95% confidence interval (CI) 0.669-0.917, P < 0.001) and for CA125 0.766 (95% CI 0.626-0.907, P = 0.002). Based on our data, we suggest that OPN can be used as a possible differential diagnostic biomarker to distinguish between malignant serous ovarian carcinoma and BOT.

In vivo growth of mantle cell lymphoma xenografts in immunodeficient mice is positively regulated by VEGF and associated with significant up-regulation of CD31/PECAM1.

Mantle cell lymphoma (MCL) is an aggressive lymphoma subtype with dismal prognosis. New treatments are needed to improve outcome of relapsed/ refractory disease. Recently, several drugs targeting at least partially the process of angiogenesis have been successfully tested in the therapy of MCL. Molecular mechanisms that regulate MCL-induced angiogenesis and that might represent potential new druggable targets remain, however, incompletely understood. We established two mouse models of human MCL by subcutaneous xenotransplantation of JEKO-1 and HBL-2 cell lines into immunodeficient mice. Histological analyses of xenografts confirmed their neovascularization. The growth of xenografts was significantly suppressed by single-agent therapy with bevacizumab, monoclonal antibody targeting vascular endothelial growth factor (VEGF). Subsequently, we analysed expression of 94 angiogenesis related genes in ex vivo isolated JEKO-1 and HBL-2 cells compared to in vitro growing cells using TaqMan low-density arrays. The most up-regulated genes in both JEKO-1 and HBL-2 xenografts were genes encoding platelet/endothelial cell-adhesion molecule (CD31/PECAM1), VEGF receptor 1 (FLT1), hepatocyte growth factor (HGF), angiogenin (ANG) and transcription factor PROX1. The most downregulated genes in both JEKO-1 and HBL-2 xenografts were midkine (MDK) and ephrine B2 (EPHB2). In summary, our results demonstrate an important role of angiogenesis in the biology of MCL and provide preclinical evidence of potent anti-MCL activity of bevacizumab. In addition, gene expression profiling of 94 angiogenesis-related targets revealed several in vivo up-regulated and down-regulated transcripts. The most differentially expressed target in both MCL tumours was CD31/PECAM1. Whether any of these molecules might represent a potential druggable target in MCL patients remains to be elucidated.

Multi-level disruption of the extrinsic apoptotic pathway mediates resistance of leukemia cells to TNF-related apoptosis-inducing ligand (TRAIL).

Disruption of apoptotic pathways belongs to commonly reported molecular mechanisms that underlie cancer drug resistance. Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL, Apo2L) is a cytokine of the TNF family with selective anti-tumor activity and minimal toxicity toward healthy tissues. Primary leukemia cells are, however, largely intrinsically resistant to TRAIL-induced apoptosis. In this study we analyzed molecular differences between TRAIL-resistant K562 cell line and TRAIL-sensitive K562 clones. We demonstrate that TRAIL-sensitive K562 cells differ from the TRAIL-resistant cell line by cell surface downregulation of TRAIL decoy receptor 1, upregulation of both TRAIL death receptors, enhanced assembly and improved functioning of the death-inducing signaling complex, and increased cytoplasmic protein expression of CASP8 and key proapoptotic BCL2 members BID, BIM, BAD and BAK. The molecular basis of the intrinsic leukemia cell TRAIL resistance thus appears a consequence of the multi-level disruption of the extrinsic apoptotic pathway. The results of this study also suggest that the leukemia TRAIL-resistance is functional, leaving a possibility of overcoming the resistance by preexposure of the leukemia cells to potent TRAIL sensitizers, e.g. BH3-mimetics.

Humoral immune response to keyhole limpet haemocyanin, the protein carrier in cancer vaccines.

Keyhole limpet haemocyanin (KLH) appears to be a promising protein carrier for tumor antigens in numerous cancer vaccine candidates. The humoral immune response to KLH was characterized at the single-cell level with ELISPOT combined with separations of cell populations according to their expression of homing receptors (HRs). The analysis of HR expressions is expected to reveal the targeting of the immune response in the body. Eight orally primed and four nonprimed volunteers received KLH-vaccine subcutaneously. Circulating KLH-specific plasmablasts were found in all volunteers, 60 KLH-specific plasmablasts/10(6) PBMC in the nonprimed and 136/10(6) in the primed group. The proportion of L-selectin(+) plasmablasts proved high and integrin α(4)ß(7) (+) low. KLH serving as protein carrier in several vaccines, the homing profile of KLH-specific response may be applicable to the cancer antigen parts in the same vaccines. The present data reflect a systemic homing profile, which appears advantageous for the targeting of immune response to cancer vaccines.

Clinical and molecular characterization of a family with a dominant renin gene mutation and response to treatment with fludrocortisone.

BACKGROUND: A family was identified with autosomal dominant inheritance of anemia, polyuria, hyperuricemia, and chronic kidney disease. Mutational analysis revealed a novel heterozygous mutation c.58T > C resulting in the amino acid substitution of cysteine for arginine in the preprorenin signal sequence (p.cys20Arg) occurring in all affected members. METHODS: Effects of the identified mutation were characterized using in vitro and in vivo studies. Affected individuals were clinically characterized before and after administration of fludrocortisone. RESULTS: The mutation affects endoplasmic reticulum co-translational translocation and posttranslational processing, resulting in massive accumulation of non-glycosylated preprorenin in the cytoplasm. This affects expression of intra-renal RAS components and leads to ultrastructural damage of the kidney. Affected individuals suffered from anemia, hyperuricemia, decreased urinary concentrating ability, and progressive chronic kidney disease. Treatment with fludrocortisone in an affected 10-year-old child resulted in an increase in blood pressure and estimated glomerular filtration rate. CONCLUSIONS: A novel REN gene mutation resulted in an alteration in the amino acid sequence of the renin signal sequence and caused childhood anemia, polyuria, and kidney disease. Treatment with fludrocortisone improved renal function in an affected child. Nephrologists should consider REN mutational analysis in families with autosomal dominant inheritance of chronic kidney disease, especially if they suffer from anemia, hyperuricemia, and polyuria in childhood.

[Measurement of bone mineral density by heel ultrasound and forearm DXA in clinical practice]. / Hodnocení kostní minerální denzity ultrazvukem patní kosti a duální rentgenovou absorpciometrií predloktí v klinické praxi.

OBJECTIVE: The comparation of two possibilities of bone mineral density measurement--ultrasound and dual energy x-ray absorptiometry. DESIGN: Open nonrandomized observation study. SETTING: Department of Obstetrics and Gynecology, 1st Faculty of Medicine Charles University and General Faculty Hospital Prague. METHODS: We examined 190 women with mean age 56 years by both methods--Lunar PIXI (dual energy x-ray absorptiometry) on forearm and CUBA Clinical (broadband ultrasound attenuation) on heel. We took personal history for menopause status, hormone replacement therapy, smoking, sport activity and age. RESULTS: The incidence of T-scores was the same for both methods, there were differences in Z-scores. In both methods we have seen the same tendencies of interaction with risk factors. Bone mineral density (BMD) respective T-score significantly decreased with age. There were no significant connections between BMD and body mass index (lineary regression test), hormone replacement therapy (paired t-test), smoking and physical exercise (Mann-Whitney U test). T-score was significantly (p < 0.003) lower in women with history of fracture (Mann-Whitney U test). CONCLUSION: In spite of totally different principles of measurement both methods are able to screen BMD.

The role of apoptosis in cancer development and treatment: focusing on the development and treatment of hematologic malignancies.

Apoptosis is a normal aspect of human physiology ensuring tissue homeostasis. Evasion of endogenous cell death processes, including apoptosis, represents one of the characteristics of cancer. Defects in the physiological mechanisms of apoptosis contribute to the pathological cell expansion and to the development and progression of cancer. Resistance of malignant cells to cancer therapeutic agents may be, in some cases, caused by dysregulation of apoptotic pathways, e.g. BCL2 or IAP overexpression. The understanding of the physiological mechanisms that control apoptosis and the elucidation of apoptotic defects in cancer cells may lead to the development of targeted cancer therapies. Apoptotic pathways, molecules involved in the cross-talk between individual apoptosis pathways and promising new anti-cancer agents, which trigger directly or indirectly apoptosis of hematologic cancer cells, are reviewed in this article.

The biological environment of Hodgkin's lymphoma and the role of the chemokine CCL17/TARC.

Hodgkin's lymphoma is a lymphoproliferative disease, which differs in its morphology and therapeutic response from other lymphomas. Neoplastic cells represent only a minor cell population of the tumour, while the major part of the tumour is formed by inflammatory cells. It results from the production of cytokines and chemokines both by neoplastic cells and by inflammatory cells. An important prognostic marker in Hodgkin's lymphoma appears to be the chemokine (C-C motif) ligand 17 (CCL17), also known as thymus and activation-related chemokine (TARC). This chemokine is expressed by many cell types and tissues, and in the case of Hodgkin lymphoma, also by Reed-Sternberg cells. CCL17/TARC binds to chemokine receptors CCR4 and CCR8 and displays chemotactic activity for T lymphocytes and some other leukocytes. The understanding of biological pathways in Hodgkin's lymphoma could be important for monitoring of disease activity and for the development of future targeted therapy.

Cell death signalling pathways in the pathogenesis and therapy of haematologic malignancies: overview of therapeutic approaches.

Malignant diseases, including haematologic malignancies, are associated with defects in the cell death mechanism. These defects are not only important for the growth advantage of the malignant clone, but when understood can be used for specific therapeutic targeting of malignant cells while sparing normal cells. The promising groups of agents that trigger, directly or indirectly, apoptosis of haematologic cancer cells are reviewed in this article. Some of the agents have recently been approved for therapy, some are under the clinical evaluation in various phases of clinical trials and some are tested under the experimental laboratory conditions.

Cell death signalling pathways in the pathogenesis and therapy of haematologic malignancies: overview of apoptotic pathways.

Apoptosis, a Greek descriptive term for falling leaves or petals, plays an important role in the progression of many diseases. Apoptosis is essential for the development and survival of multi-cellular organisms. Malignant diseases, including haematologic malignancies, are associated with defects in the cell death mechanism. These defects are not only important for the growth advantage of malignant clones, but when understood can be used for specific therapeutic targeting of malignant cells while sparing normal cells. The cellular and molecular mechanisms of apoptosis have been extensively demonstrated and are reviewed in this article. In this part of the review we focus on basic details of the apoptosis pathways, key players of the receptor-mediated apoptosis, and molecules involved in the cross-talk between individual apoptosis pathways and apoptosis regulation.

Alterations of uromodulin biology: a common denominator of the genetically heterogeneous FJHN/MCKD syndrome.

Autosomal dominant hyperuricemia, gout, renal cysts, and progressive renal insufficiency are hallmarks of a disease complex comprising familial juvenile hyperuricemic nephropathy and medullary cystic kidney diseases type 1 and type 2. In some families the disease is associated with mutations of the gene coding for uromodulin, but the link between the genetic heterogeneity and mechanism(s) leading to the common phenotype symptoms is not clear. In 19 families, we investigated relevant biochemical parameters, performed linkage analysis to known disease loci, sequenced uromodulin gene, expressed and characterized mutant uromodulin proteins, and performed immunohistochemical and electronoptical investigation in kidney tissues. We proved genetic heterogeneity of the disease. Uromodulin mutations were identified in six families. Expressed, mutant proteins showed distinct glycosylation patterns, impaired intracellular trafficking, and decreased ability to be exposed on the plasma membrane, which corresponded with the observations in the patient's kidney tissue. We found a reduction in urinary uromodulin excretion as a common feature shared by almost all of the families. This was associated with case-specific differences in the uromodulin immunohistochemical staining patterns in kidney. Our results suggest that various genetic defects interfere with uromodulin biology, which could lead to the development of the common disease phenotype. 'Uromodulin-associated kidney diseases' may be thus a more appropriate term for this syndrome.

[Effect of early onset of transdermal and oral estrogen therapy on the lipid profile: a prospective study with cross-over design]. / Vliv casne zahájené transdermální a perorální estrogenní substitucní terapie na lipidový profil--prospektivní studie s cross-over designem.

OBJECTIVE: The aim of study was to evaluate changes of lipid profil during different types of estrogen replacement therapy (ERT). DESIGN: Prospective randomized study. SETTING: 1st Faculty of Medicine and General Faculty Hospital Prague. METHODS: Two routes of administration were used for 12 weeks: oral estradiol 2 mg/day and transdermal estradiol 50 microg/day (7-day pathes). Forty five healthy women with average age 49 +/- 6 years were randomised into prospective cross-over designed study. Forty one women finished the study and were analysed. Blood collections were performed from veins on the beginning of study and during last week of each therapeutic interventions. Statistical results have counted by paired t-test. RESULTS: Total cholesterol levels were not changed. Triglycerides grew up from 1,39 +/- 0,9 mmol/l to 1.61 +/- 0.8 mmol/l (p = 0,004) after oral ERT. This results showed significant (p = 0.0001) differences between oral and transdermal application because of nonsignificant (p = 0.187) lowering trends after transdermal ERT. The elevation of HDL levels after oral ERT (from 1.85 +/- 0.39 mmol/l to 2.09 +/- 0.42 mmol/l, p = 0.0001) was significantly (p = 0.009) more favourable than after transdermal ERT (to 1.96 +/- 0.42 mmol/l, p = 0.029). Changes of LDL levels are also more favourable (p = 0.0001). LDL levels decreased after oral ET from 3.06 +/- 0.97 mmol/l to 2.52 +/- 0.71 mmol/l in comparison with the nonsignificant decline on 3.0 +/- 1.0 mmol/l after transdemal ERT. CONCLUSION: All data from lipid metabolism had more favourable changes after oral ET with the exception of triglycerides. Knowledge of the patient's lipid profile and it's changes after each type of estrogen applications enable doctors individualisation of hormone replacement therapy from this point of view.

[Pregnancy and labor after combined pancreas-kidney transplantation in Czech Republic]. / Tehotenství a porod po kombinované transplantaci ledviny a pankreatu v Ceské republice.

OBJECTIVE: To inform about the first own experiences and to present opinion on leading pregnancy and delivery after a combined pancreas and kidney transplantation. DESIGN: Case report and review article. SETTING: Department of Obstetric and Gynecology, 1st Medical Faculty of the Charles University and General Faculty Hospital, Prague. RESULTS AND CONCLUSIONS: Pregnancies and deliveries after the transplantation of solid organs are not common. Mostly there are experiences with women after kidney transplantation, smaller or no experiences are with women after transplantation of other solid organs. About 25 pancreas transplantation per year are performed in the Czech republic. Two women after the combined kidney and pancreas transplantation were the first in Czech republic to get pregnant spontaneously and delivered by using a chronic immunosupressive. therapy (Prograf, Imuran, Prednison) in 2002 and 2003. These single pregnancies were led as a high-risk pregnancy in Regional Perinatology Center in collaboration with Transplant and Diabetic Center. Both pregnancies were termined from the obstetrical indication before the term by cesarean section. Both children were healthy. The pregnancy of both patients has not affected the function of the transplanted organs and development of both children has been normal.

[Reconstruction procedures following pelvic exenterations]. / Rekonstrukcní výkony po exenteraci pánve.

OBJECTIVE: Review of reconstruction procedures following pelvic exenterations. DESIGN: Review article. SETTING: Department of Obstetrics and Gynecology, Department of Urology, 1st Department of Surgery, Faculty Teaching Hospital and 1st Medical Faculty of the Charles University, Prague. METHODS: Review and critical assessment of published data. CONCLUSIONS: Reconstruction procedures are important part of pelvic exenterations. The procedures are crucial for following quality of life. Currently the most frequently used techniques for isolated pelvic floor support are omental flaps (carpets), for combined reconstruction of pelvic floor and vagina TRAM (transverse rectus abdominis musculocutaneus flap). Reconstructions prolong operation time; however they are accompanied with low morbidity and some techniques decrease total morbidity of exenterative procedure. Total and posterior exenterations require sigmoideostomy in vast majority of cases. Low rectal anastomosis might be used in cases of supralevator procedures. They cause high morbidity especially in patients following radiotherapy. In these patients temporary diverting colostomy is being recommended. A bowel segment is usually used for urinary diversion following total or anterior exenteration. Golden standard remain the incontinent ureteroenterostomies using ileum or colon transversum. Currently continent diversions are considered more often due to encouraging results and good quality of life. Heterotopic diversions, with continent conduit and cutaneous stoma, are frequently used. Risk of serious complications, especially fistulas and stoma stenosis, after all types of diversions is possible to reduce by using appropriate bowel segment not handicapped by previous radiotherapy.

[Abdominal radical trachelectomy--technique and experience]. / Abdominailni radikailni trachelektomie--technika a zkusenosti.

OBJECTIVE: Discussion of current experiences with abdominal radical trachelectomy in the treatment of early stages of cervical cancer in fertile women. DESIGN: Case-reports. SETTING: Department of Obstetrics and Gynecology, General Faculty Hospital and 1st Medical Faculty, Charles University, Prague. METHODS: Presentation of 4 cases of abdominal radical trachelectomy and pelvic lymphadenectomy. Discussion with published data. RESULTS: Three cases of open abdominal and one case of laparoscopic abdominal radical trachelectomies together with pelvic lymphadenectomies are presented. All procedures were indicated for cervical cancer stages IA2-IB1. Frozen section of pelvic nodes and a slice of upper margin of cervix revealed no metastasis or infiltration. In total 22-43 pelvic nodes were removed, being negative in all cases. Operative time ranged between 148 and 270 min. in laparotomy and 250 min. in laparoscopy. Blood loss reached 350-3500 ml. There were no intraoperative complications, postoperatively one case of bladder atony was treated by suprapubic drainage for 30 days, one case of ileus was managed pharmacologically. Vaginal suture healed properly in all cases. No complications occurred within limited follow-up of 1-5 months. CONCLUSION: Abdominal radical trachelectomy with pelvic lymphadenectomy is a rational alternative in the treatment of stages IA2-IIA cervical cancer in women of fertile age. Standard radicality in parametria resection and easy incorporation into armamentarium of oncogynecological centers are main advantages of such approach. Laparotomy can be avoided using laparoscopy.