A comparative study of angiogenic and cytokine responses after laparoscopic cholecystectomy performed with standard- and low-pressure pneumoperitoneum

Background  Surgical procedures enhance production of pro- and anti-inflammatory cytokines and angiogenic factors that play a pivotal role in the immunological response to surgical trauma and take part in the pathogenesis of tumor growth and adhesions formation. The purpose of the study was to access the influence of low-pressure CO₂ pneumoperitoneum on the inflammatory and angiogenic responses during the postoperative period after laparoscopy. Methods  The study group consisted of 40 patients, operated on due to cholelithiasis using standard-pressure (n = 20) and low-pressure (n = 20) CO₂ pneumoperitoneum. Serum concentration of interleukin (IL)-6, IL-8, IL-10, vascular endothelial growth factor (VEGF)-A, and endostatin were measured before and at 6, 24, and 48 h after surgery with commercially available enzyme-linked immunosorbent assay (ELISA). Results  Concentrations of IL-6 increased significantly after the operations in both groups. No differences were observed between the groups in regards to IL-6, IL-8, and IL-10 levels. Concentrations of VEGF-A measured at 6 and 48 h were significantly lower in patients who underwent laparoscopies performed with low-pressure pneumoperitoneum. No significant variations were observed in endostatin serum concentration. Concentrations of the studied parameters were not influenced by duration of surgery or by age, gender, or body mass index (BMI) of the patients. Conclusions  The results obtained in our study do not show any significant differences between studied operative procedures with regards to systemic inflammatory response. Changes in the concentrations of VEGF-A and endostatin observed in the studied population may suggest this technique is more favorable with regards to angiogenesis process intensity, along with all its consequences and implications.     

Stress urinary incontinence in women

The aim of the study was the investigation of the biochemical condition of elements likely to directly participate in active closing of the urethral lumen. We estimated glycogenolysis in urinary bladder, perivesical connective tissue and levator ani muscle (LAM) samples obtained intraoperatively from 80 stress incontinent women. Glycogen content as well as activities of active and total glycogen phosphorylase and acid exo-1,4-alpha-glucosidase were measured. Material from the urinary bladder and perivesical connective tissue was insignificantly altered, and glycogen contents in the bladder (2.03±1.38 g/100 g wet tissue) were considered to be normal. In the LAM glycogenolysis was much more activated than in other tissues (p<0.001 by Fischer's exact test). Of LAM specimens 78% (22/28) revealed imbalanced biochemistry of glycogen with activation of hydrolytic decomposition. We conclude that stress urinary incontinence in women is frequently associated with metabolic alterations in the periurethral striated fibres. This study indirectly supports our recent hypothesis on the pathogenesis of the disease in terms of muscle fibre type transitions.     

Characterization of human hepatocytes isolated from non-transplantable livers

INTRODUCTION: The successful use of hepatocytes depends on a reliable demonstration of the functional and morphological integrity of isolated cells. Herein we investigated whether the isolation and cryopreservation of primary human hepatocytes can compromise cell viability and liver-specific characteristics. MATERIAL/METHODS: Hepatocytes were isolated from encapsulated human liver segments by a modified 2-step perfusion technique. Isolated cells were Percoll-purified, cryopreserved, and stored in liquid nitrogen for 1-12 months. For rapid assessment of fresh and cryopreserve/thawed hepatocyte yield and viability, the cells were stained with trypan blue or labeled with fluorochromes. For immunocytochemical analysis, the cells were labeled with monoclonal antibodies for the presence of the following antigens and chemokines: CD3, CD45Ro, CD45Ra, CD34, CD68, CD90, CD95, CD20, HLA-DR, Ki67, PCNA, Bcl-2, p53, CXCR3, CXCR4, and SDF-1. The cells were tested for several specific functions, such as ureagenesis, energy status, MTT activity, lactate dehydrogenase leakage, and total CYP450 content. RESULTS: Assessment of both freshly isolated (Percoll-purified) and cryopreserved/thawed hepatocytes revealed a low constitutive level of contamination by non-parenchymal cells compared with crude (unpurified) preparations and tissue sections. All viable hepatocytes showed intact morphology and retained CYP450 protein, energy status, and urea synthesis. CONCLUSIONS: Modifications in hepatocyte preparations, such as depletion of dead, damaged, and nonparenchymal cells, improves cell purity, which can be adapted to further evaluation of hepatocyte immunogenicity. These data illustrate the importance and feasibility of human hepatocyte banking.     

Hepatitis C virus (HCV) genotype 1 NS5A resistance-associated variants are associated with advanced liver fibrosis independently of HCV-transmission clusters

Objectives

The aim of the study was to characterize the differences in the frequencies of NS3 and NS5A resistance-associated variants (RAVs) among Polish therapy-naive genotype 1 (G1) hepatitis C virus (HCV)-monoinfected and human immunodeficiency virus (HIV)/HCV-coinfected patients including clustering patterns and association of RAV frequency with liver fibrosis.

Strong association of HLA-Cw6 allele with juvenile psoriasis in Polish patients

Association of psoriasis vulgaris with HLA-C is not equally strong in different human populations. It has not yet been studied in Polish patients at DNA level, but only by serology that is inadequate for HLA-C. Therefore, we examined the distribution of HLA-C alleles by means of low resolution PCR-SSP in 102 Polish psoriatics and 123 healthy controls. We have found significantly higher representation of HLA-Cw*06 (odds ratio, 18.73; P(cor)<0.001) and significantly lower representation of HLA-Cw*07 (odds ratio, 0.41; P(cor)<0.038) in patients than in controls. Association of HLA-Cw*06 with psoriasis was even stronger in early age at onset (0-20 years) group: odds ratio, 77.71; P(cor)<0.001. Therefore, our population seems to belong to those with strong association of psoriasis with HLA-Cw*06.     

Fibroblasts from recurrent fibrotic overgrowths reveal high rate of proliferation in vitro - findings from the study of hereditary and idiopathic gingival fibromatosis

ABSTRACT

Purpose: Investigate the content of fibrotic fibrils in gingival tissue and the proliferation of fibroblasts collected from recurrent and non-recurrent hereditary gingival fibromatosis (HGF) and idiopathic gingival fibromatosis (IGF).

Methods: Gingival biopsies were collected from HGF (n = 3) and IGF (n = 3) donors with recurrent and non-recurrent gingival overgrowths and from a control group (Ctrl, n = 3). Hematoxylin staining was performed to evaluate the histomorphology of gingival tissue. Heidenhain’s AZAN trichrome staining served for visualization of fibrotic fibrils in gingiva. Quantitative analysis of the content of fibrotic fibrils in gingival tissue was performed using a polarized light microscope. Proliferation was evaluated at 24 h, 48 h, and 72 h in fibroblast cultures using a cell proliferation ELISA assay based on 5-bromo-2?-deoxyuridine (BrdU).

Results: Numerous blood vessels and fibroblasts were observed in recurrent overgrowths, whereas moderate blood vessels and moderate to scanty fibroblasts were detected in non-recurrent overgrowths. Heidenhain’s staining revealed numerous collagen fibers in both recurrent and non-recurrent overgrowths. Quantitative analysis in a polarizing microscope showed significant accumulation of fibrotic fibrils exclusively in the overgrowths with the recurrence. In all time-points, increased proliferation of cells from all recurrent overgrowths was observed, but not from overgrowths which do not reoccur.

Conclusions: The study revealed that recurrent gingival overgrowths consist of highly fibrotic and dense connective tissue with numerous blood vessels and abundant fibroblasts. We also demonstrated that unlike fibroblasts derived from overgrowths, which did not present recurrence, fibroblasts derived from highly fibrotic and recurrent overgrowths maintain high rate of proliferation in vitro.     

Effects of Dietary Oat Beta-Glucans on Colon Apoptosis and Autophagy through TLRs and Dectin-1 Signaling Pathways—Crohn’s Disease Model Study

Background: Crohn’s disease (CD) is characterized by chronic inflammation of the gastrointestinal tract with alternating periods of exacerbation and remission. The aim of this study was to determine the time-dependent effects of dietary oat beta-glucans on colon apoptosis and autophagy in the CD rat model. Methods: A total of 150 Sprague–Dawley rats were divided into two main groups: healthy control (H) and a TNBS (2,4,6-trinitrobenzosulfonic acid)-induced colitis (C) group, both including subgroups fed with feed without beta-glucans (βG−) or feed supplemented with low- (βGl) or high-molar-mass oat beta-glucans (βGh) for 3, 7, or 21 days. The expression of autophagy (LC3B) and apoptosis (Caspase-3) markers, as well as Toll-like (TLRs) and Dectin-1 receptors, in the colon epithelial cells, was determined using immunohistochemistry and Western blot. Results: The results showed that in rats with colitis, after 3 days of induction of inflammation, the expression of Caspase-3 and LC3B in intestinal epithelial cells did not change, while that of TLR 4 and Dectin-1 decreased. Beta-glucan supplementation caused an increase in the expression of TLR 5 and Dectin-1 with no changes in the expression of Caspase-3 and LC3B. After 7 days, a high expression of Caspase-3 was observed in the colitis-induced animals without any changes in the expression of LC3B and TLRs, and simultaneously, a decrease in Dectin-1 expression was observed. The consumption of feed with βGl or βGh resulted in a decrease in Caspase-3 expression and an increase in TLR 5 expression in the CβGl group, with no change in the expression of LC3B and TLR 4. After 21 days, the expression of Caspase-3 and TLRs was not changed by colitis, while that of LC3B and Dectin-1 was decreased. Feed supplementation with βGh resulted in an increase in the expression of both Caspase-3 and LC3B, while the consumption of feed with βGh and βGl increased Dectin-1 expression. However, regardless of the type of nutritional intervention, the expression of TLRs did not change after 21 days. Conclusions: Dietary intake of βGl and βGh significantly reduced colitis by time-dependent modification of autophagy and apoptosis, with βGI exhibiting a stronger effect on apoptosis and βGh on autophagy. The mechanism of this action may be based on the activation of TLRs and Dectin-1 receptor and depends on the period of exacerbation or remission of CD.