Agnieszka Malinska

Peritoneal mesothelium promotes the progression of ovarian cancer cells in vitro and in a mice xenograft model in vivo

The role of mesothelial cells in the intraperitoneal spread of ovarian cancer is still elusive. In particular, it is unclear whether these cells constitute a passive barrier preventing cancer cell progression or perhaps act as an active promoter of this process. In this report we show that omental human peritoneal mesothelial cells (HPMCs) stimulate adhesion and proliferation of ovarian cancer cells (A2780, OVCAR-3, SKOV-3). The latter was associated with the paracrine activity of GRO-1, IL-6, and IL-8 released to the environment by HPMCs. Furthermore, the growth dynamics of ovarian cancer xenografts produced in response to i.p. injection of ovarian cancer cells together with HPMCs was remarkably greater than for implantation of cancer cells alone. A layer of peritoneal mesothelium was consistently present in close proximity to the tumor mass in every xenograft model. In conclusion, our results indicate that HPMCs play a supporting role in the intraperitoneal invasiveness of ovarian malignancy, whose effect may be attributed to their ability to stimulate adhesion and proliferation of cancer cells.

Predictive factors of late venous aortocoronary graft failure: ultrastructural studies.

Background Venous aortocoronary graft arterialization may precede a preterm occlusion in some coronary artery bypass grafting (CABG) patients. The aim of the present study was to identify ultrastructural variations in the saphenous vein wall that may have an impact on the development of venous graft disease in CABG patients. Methods The study involved 365 consecutive patients with a mean age of 62.9±9.4 years who underwent isolated CABG. The thickness and area of the whole venous wall, the tunica intima, the tunica media and the adventitia and the number and shape (length, thickness and length/thickness ratio) of the nuclei in the medial smooth muscle cells nuclei in the distal saphenous vein segments were evaluated by ultrastructural studies. Patients were followed up for 41 to 50 months (mean 45.1±5.1). Saphenous vein graft patency was assessed by follow-up coronary angiography. Logistic regression models were used to identify independent risk factors for late graft failure. Results In 71 patients significant lesions in the saphenous vein grafts were observed. The whole venous wall thickness (437.5 µm vs. 405.5 µm), tunica media thickness (257.2 µm vs. 211.5 µm), whole venous wall area (2.23 mm2 vs. 2.02 mm2) and tunica media area (1.09 mm2 vs. 0.93 mm2) were significantly larger for this group of patients than for those without graft disease. In the latter group more elongated smooth muscle cell nuclei (higher length/thickness ratio) were found in the tunica media of the saphenous vein segments. Thickening of the saphenous vein tunica media and chunky smooth muscle cell nuclei were identified as independent risk factors for graft disease development. Conclusions Saphenous vein tunica media hypertrophy (resulting in wall thickening) and chunky smooth muscle cell nuclei might predict the development of venous graft disease.

Endothelial integrity of radial artery grafts harvested by minimally invasive surgery — immunohistochemical studies of CD31 and endothelial nitric oxide synthase expressions: a randomized controlled trial

OBJECTIVE To compare the endothelial integrity of radial artery grafts harvested by minimally invasive surgery and arteries harvested conventionally for coronary artery bypass surgery (CABG) in 200 participants, who were assigned to interventions by using random allocation. METHODS An immunohistochemical procedure with monoclonal antibodies was employed to estimate CD31 antigen and endothelial nitric oxide synthase (eNOS) expressions - markers defining endothelial integrity. RESULTS The CD31 immunostaining revealed that the endothelial cell integrity of the minimally invasive harvested arteries was preserved in 76.1±7.4% of the circumference of luminal endothelium, which was similar to results obtained in conventionally harvested grafts (77.2±9.8%; not significant). On the other hand, eNOS immunostaining indicated that the endothelial integrity of the minimally invasive harvested grafts was preserved in 75.4±10.5% while in conventionally harvested grafts it was reduced to 42.4±14.5% of the total luminal endothelium circumference (P<0.05). CONCLUSIONS The endothelial integrity of radial artery grafts harvested by minimally invasive surgery is better preserved than in the grafts obtained by the conventional manner. This could play an important role in improving graft patency and might represent a preliminary condition of stable functioning in coronary arterial bypasses.

Preexisting High Expression of Matrix Metalloproteinase-2 in Tunica Media of Saphenous Vein Conduits Is Associated with Unfavorable Long-Term Outcomes after Coronary Artery Bypass Grafting

Introduction. Migration of the smooth muscle cells (SMCs) to the tunica media in the saphenous vein (SV) transplants is facilitated by matrix metalloproteinases (MMPs). The aim of this study was to identify any associations between expression of MMP-2 or endogenous tissue inhibitors (TIMP-2 and TIMP-3) in the SV segments and late failure of the SV grafts. Methods. Two hundred consecutive patients with a mean age of 63.1 ± 8.9 years who underwent primary isolated venous CABG were examined. Patients were retrospectively split into two subgroups, with the SV graft disease (SVGD (+); n = 47) or without it (SVGD (−); n = 153). In the SV segments, immunohistochemical analysis of the expression of the MMP-2, TIMP-2, and -3 was performed. Results. In the SVGD (+) patients, tissue expression of MMP-2 was stronger, whereas that of both TIMPs was weaker than in the SVGD (−) patients. In majority of the SV segments obtained from the SVGD (−) individuals, a balance in MMP and TIMP expressions was found, whereas an upregulation of MMP-2 expression was usually noted in the SVGD (+) subjects. Conclusion. The strong expression of MMP-2 accompanied by reduced immunostaining of both TIMPs is associated with the development of the SV graft disease and unfavorable CABG outcomes.

Histological evaluation of age-related variations in saphenous vein grafts used for coronary artery bypass grafting.

Introduction Venous coronary artery bypass grafts (CABG) might undergo a process of arterialization resulting in neointimal formation and medial hypertrophy. It is often followed by critical occlusion of the graft lumen. The aim of the study was to assess histological representative features of saphenous vein reconstruction in aging as well as to establish optimal patients’ age limits applicable for optimal selection of grafts. Material and methods One hundred and ten patients undergoing venous CABG were divided into 4 age subgroups: (A) 50 years and less, (B) 51-60 years, (C) 61-70 years and (D) > 70 year-old subjects. Distal venous graft segments were saved for an adequate morphometric assay which was followed by suitable statistical analysis. Results The entire venous wall thickness as well as its tunica media were found to become significantly thinner between subgroups A and D. The number of smooth muscle cell (SMC) nuclei within the tunica media did not differ between study subgroups. The majority of these nuclei in subgroup D were found, however, to be more elongated than in subgroup A (SMC length/width index in subgroup D was found to be significantly higher than in subgroup A). Conclusions Progressive, age-related thinning of the venous wall and tunica media as well as SMC nucleus elongation might suggest impairment of SMCs’ migration and proliferation rate. Consequently, individuals aged 70 years and over may benefit clinically more from venous CABG than younger patients due to the lower risk of arterialization and occlusion of the graft lumen in the future.

Mycophenolate mofetil (MMF) treatment efficacy in children with primary and secondary glomerulonephritis

Introduction The aim of our study was to analyse the efficacy and safety of mycophenolate mofetil (MMF) as part of the complex immunosuppressive therapy in children with different types of primary and secondary glomerulonephritis, who were not eligible for the standard treatment routine suggested by evidence-based guidelines. Material and methods The study group comprised 85 children with proteinuric glomerulopathies hospitalized between 2007 and 2010, who were non-responders to immunosuppressive therapy. The dose of MMF was established as 1 g/m2/24 h. Remission was defined as negative proteinuria in three consecutive urinalyses. Results The patients were divided into 4 groups: idiopathic nephrotic syndrome (n = 35), primary glomerulonephritis (n = 15), auto-antibody associated glomerulonephritis (n = 20) and lupus nephropathy (LN, n = 15). Ten patients from the first group (29%) and 5 patients each from the second and third group (34% and 25% respectively) did not respond to MMF therapy. On the other hand, all the children diagnosed with LN have reached clinical and biochemical remission. Conclusions Alternative rescue MMF therapy should always be taken into consideration in proteinuric patients who are non-responders to steroids, cyclosporine A and cyclophosphamide in whom the initial glomerular filtration rate is higher than 60 ml/min/1.73m2. It is recommended that MMF be administered as part of the standard treatment regimen in patients diagnosed with lupus nephropathy. In these groups of patients, the potent benefits of this therapy are higher than expected side-effects.

Nephrotic syndrome unfavorable course correlates with downregulation of podocyte vascular endothelial growth factor receptor (VEGFR)-2

Idiopathic nephrotic syndrome (INS) in children is most commonly caused by primary glomerulopathies. Morphological lesions observed in INS might be secondary to inflammatory factors of mainly extra-renal origin. The vascular endothelial growth factor (VEGF) family is regarded as playing a crucial role in this pathomechanism. The aim of the present work was to analyze the possible relation between VEGF-C and VEGF receptor (VEGFR)-2 expressions at electron microscopy level in different INS cases. The study group comprised 18 children with minimal change disease (MCD), 30 patients diagnosed with diffuse mesangial proliferation (DMP) and 11 subjects with focal segmental glomerulosclerosis (FSGS). An indirect immunohistochemical assay employing monoclonal anti-VEGF-C and anti-VEGFR-2 antibodies was applied in the study. The immunohistochemical expression of VEGF-C within podocyte cytoplasm was significantly increased in DMP subjects who were resistant to steroids and in all FSGS patients compared to MCD children and controls (p 〈 0.05). VEGF-C over-expression in these cases was followed by downregulation of VEGFR-2. Nephrotic syndrome progression correlates with the downregulation of podocyte VEGFR-2. For this reason, decreased VEGFR-2 expression in the podocyte processes of children with idiopathic nephrotic syndrome might be regarded as a potent factor of unfavorable prognosis.

Cytokeratin 8 in venous grafts: A factor of unfavorable long-term prognosis in coronary artery bypass grafting patients

BACKGROUND Smooth muscle cells, present in the saphenous vein (SV) tunica media, may contribute to late occlusion of venous aortocoronary grafts. The aim of present study was to evaluate expression of selected cytoskeletal proteins in tunica media of SV grafts obtained from patients undergoing coronary artery bypass grafting (CABG) and correlate procured results to late venous graft failure observed in these patients. METHODS The study involved 218 patients (mean age of 62.5 ± 8.7 years) who underwent primary isolated CABG with the use of at least one venous aortocoronary bypass graft. Expressions of alpha-smooth muscle actin, smooth muscle-myosin heavy chain, calponin and cytokeratin 8 in SV wall were estimated by means of immunohistochemistry. The primary clinical endpoint was defined as the presence of any coronary artery disease (CAD) progression symptom while angiographic one as significant stenosis in the venous graft. RESULTS Thirty-eight (18.1%) patients have reached the primary clinical endpoint. Freedom from clinical CAD deterioration was 0.95 ± 0.02, 0.87 ± 0.03 and 0.83 ± 0.04, for 12-, 24-,36-month follow-up, respectively. Forty-four study participants have reached the angiographic endpoint. Multivariate logistic regression analysis revealed an increased expression of cytokeratin 8 accompanied by calponin under expression in SV tunica media were independent risk factors for venous graft failure. CONCLUSIONS An increased expression of cytokeratin 8 and weak of calponin in tunica media of SV grafts might be useful markers of unfavorable long-term prognosis in CABG patients. In the future, assessment of their expression may enable to select the most appropriate candidates for SV grafts.

Homing of hemopoietic precursor cells to the fetal rat thymus: intercellular contact-controlled cell migration and development of the thymic microenvironment

Colonization of rat thymic anlage by the first wave of hemopoietic precursor cells (HPc) was investigated by means of transmission electron microscopy and immunocytochemistry. HPc began migration into the thymic anlage between 13 and 13.5 gestation days (GD), terminated colonization at about GD 16, and migrated sequentially through the two compartments of the thymic anlage under the control of typical populations of stromal cells. First, HPc migrated through the external compartment of the perithymic mesenchyme, tightly interconnected with fibroblasts. The type of junctions between the cells indicated that the fibroblasts played a role in the control of HPc trafficking and in their entrance to the epithelial compartment. The second stage of colonization was initiated by the entrance of HPc to the epithelial compartment and their interaction with thymic epithelial cells (TECs). Based on morphological criteria, two populations of HPc were distinguished that colonized the anlage at various stages of its development. The predominant population with ultrastructural traits common to thymocytes “homed” into the epithelial type primordium. A small number of HPc, identified by protein S-100 expression and by Birbeck’s granules as precursors of dendritic cells, colonized lymphoepithelial anlage in which subsets of cortical and medullary TECs could be distinguished. Thymocyte migration and their reciprocal interactions with cortical TECs differed from the trafficking of dendritic cells toward the medulla. The results demonstrated the influence of maturing thymocytes on the development of cortical epithelial cells and the dynamic organization of the medullary microenvironment with direct involvement of dendritic cells.

SOCS3 and SOCS5 mRNA expressions may predict initial steroid response in nephrotic syndrome children

Suppressors of Cytokine Signaling (SOCS) inhibit Signal Transducers and Activators of Transcription (STATs) phosphorylation by binding and inhibiting Janus Kinases (JaKs). The aim of the present study was to evaluate the influence of glucocorticosteroids on the JaK/STAT signaling pathway in the leukocytes of nephrotic syndrome (NS) patients. The study group was composed of 34 steroid sensitive NS (SSNS) children and 20 steroid resistant NS (SRNS) subjects. Gene expression was assessed by real-time PCR using pre-designed human JaK/STAT PCR array. Protein expression was evaluated using ELISA assay (plasma concentration) and immunofluorescence (in situ protein expression). In SSNS children, the initial increased expression of JaK1, JaK2, JaK3, STAT1, STAT2, STAT6, TYK2, SOCS1, SOCS2, SOCS3, SOCS4 and SOCS5 was reduced back to the control limits. Similarly, in SRNS patients the increased levels of almost all mRNA expressions for the abovementioned genes were decreased, with the exceptions of SOCS3 and SOCS5 expressions. These mRNA expressions were still significantly increased and correlated with early unfavorable course of nephrotic syndrome in children. Plasma levels of SOCS3, SOCS5, IL-6 and IL-20 were significantly increased in SRNS subjects after six weeks of steroids medication compared to SSNS and control participants. We conclude that SOCS3 and SOCS5 increased mRNA expressions might predict initial resistance to steroids in NS patients.