Quality of life in colostomy patients practicing colonic irrigation: An observational study.

AIMS OF THE STUDY: The presence of colostomy has a major impact on quality of life that could potentially be improved by performing colonic irrigation (CI), yet few studies have assessed the impact of this technique on quality of life. The aim of this study was to assess the quality of life between two groups of patients having a colostomy; those practicing CI vs those not practicing CI. PATIENTS AND METHODS: The French Federation of Ostomy (FFO) members were evaluated by a self-questionnaire assessing their experience of CI. Quality of life as assessed by the Stoma-QOL questionnaire was compared between patients practicing CI or not. RESULTS: In total 752 patients were eligible for the study. The median age was 75 years, and 47.26% were men. The median duration between stoma surgery and questionnaire completion was 12.3 years. Forty-one percent of the patients practiced CI. The median quality of life score was significantly higher for the patients practicing the CI: (69.26 vs 58.33, P<0.001). In multivariable analysis, the risk factors for not performing CI were age, obesity, the presence of colostomy for less than six years, and a non-oncologic indication for operation. CONCLUSIONS: CI appeared to improve the quality of life of patients with colostomy. This care is a therapeutic education issue and should be proposed to all patients. Supervision by the enterostomal therapy nurse is recommended especially for patients with a high risk of failure.

Chitosan-based hydrogels for developing a small-diameter vascular graft: in vitro and in vivo evaluation.

AIMS: Vascular grafts made of synthetic polymers perform poorly in small-diameter applications (cardiac and peripheral bypass). Chitosan is a biocompatible natural polymer that can provide a novel biological scaffold for tissue engineering development. The goal of this study was to demonstrate the biocompatibility of a novel chitosan preparation in vitro and in vivo, and to assess its potential as a scaffold for vascular applications. METHODS AND RESULTS: A series of experiments of increasing complexity, ranging from in vitro biocompatibility and hemocompatibility tests to in vivo studies in small and large animals (rats and sheep), was performed to provide a comprehensive analysis of chitosan hydrogels' biological properties. In vitro studies established that: (i) chitosan supported human endothelial progenitor cells adhesion, proliferation and resistance to physiological shear stress; (ii) chitosan did not activate platelets, the complement system, or the intrinsic coagulation pathway. In vivo results showed: (iii) no resorption of chitosan and no chronic inflammation at 60 days in a rat heterotopic implantation model (magnetic resonance imaging and histology); (iv) no flow obstruction (Doppler ultrasound) and no thrombus formation (histology and scanning electron microscopy) at 2 h after a carotid arteriotomy repair with chitosan patches in sheep. Finally, two chitosan tubes were implanted as carotid interposition grafts for 3 days in sheep showing that chitosan was strong enough to be sutured, to withstand arterial pressure, and no flow obstruction was observed through this short period. CONCLUSION: Chitosan-based hydrogels displayed promising in vitro biocompatibility and hemocompatibility properties as well as in vivo short-term performance.

Anatomical basis of the coordination between smooth and striated urethral and anal sphincters: loops of regulation between inferior hypogastric plexus and pudendal nerve. Immuno-histological study with 3D reconstruction.

PURPOSE: (1) Describe both nervous pathways to the sphincters, and highlight the anatomical support of their coordination. (2) Obtain a 3D representation of this complex innervation system. METHODS: A computer-assisted anatomical dissection technique was used. Serial histological sections were cut in the pelvis of four female human foetuses (aged 19-32 weeks of gestation). The sections were treated with conventional staining, and with seven different immunostainings. The sections were digitalized and, finally, a 3D representation was built from the corresponding images. RESULTS: Myelinated and sensory fibres were detected at the inferior hypogastric plexus (IHP) level. Our analysis showed that most of the afferent sensory fibres come from the urinary and anal sphincters through the anterior and posterior branches of the IHP respectively. A highly positive nitrergic (anti-NOS1) and sensitive (anti-CGRP) labelling was found in the external layer of the urethral sphincter. The 3D representation allowed describing the two components of the innervation system. A sensory-motor regulation loop was found for both sphincters. CONCLUSION: A 3D description of the components of both nervous pathways to the sphincters has been established. Our findings on the innervation of the sphincters tend to question the classical infra/supra levatorian muscle description. The coordinated work of the internal and external layers of the anal and urethral sphincter is probably mediated by multiple roles regulation.

Is low tie ligation truly reproducible in colorectal cancer surgery? Anatomical study of the inferior mesenteric artery division branches.

AIM: Curative surgery is the standard treatment for colorectal cancer. The ligation level of the inferior mesenteric artery (IMA) is still debated, as neither low tie (LT) nor high tie ligation (HT) has shown any benefit on the patients' overall survival. We examined whether LT is standardizable and easily reproducible from an anatomical point of view. METHOD: One hundred CT angiographies of healthy patients were analysed for the anatomy of the IMA and its division branches: left colic artery (LCA), sigmoid arteries trunk and superior rectal artery. Data analysed comprised angles between the IMA and the aorta, diameters of the IMA and its branches, repartition of the branches and distances between the origin of the branches and the origin of the IMA. RESULTS: IMA anatomy showed no variation. In contrast, its division branches showed important variability in terms of distance to the origin and repartition: in 19.9% of the patients, the IMA directly splits into three branches, and in 17.6% of the patients, the LCA originated at more than 5 cm from the origin of the IMA. These frequent variations led us to assume that the standardization of LT is very difficult in a context of neoplasm, where the quality of the lymphadenectomy is fundamental. CONCLUSION: The division branches of the IMA are extremely subject to interindividual variations, making it difficult if not impossible to reproduce identically a surgical procedure based on their anatomy. HT appears to us as the only relevant procedure for colorectal cancer.

Parastomal hernia. A study of the French federation of ostomy patients.

INTRODUCTION: The prevalence of parastomal hernia (PSH) varies considerably in the literature. This condition impacts negatively on quality of life. Yet there is no surgical consensus concerning treatment. The aim of the study was to determine treatment and recurrence rates of PSH in a large population of ostomy patients. PATIENTS AND METHODS: This retrospective study was carried out by a self-administered questionnaire in a population drawn at random from the database of the French federation of ostomy patients (FSF). RESULTS: Seven hundred and eighty-two patients were eligible for the study. The mean duration of follow-up was 10.5 years. PSH was reported by 202 patients (25.6%) and appeared on average 18 months after creation of the stoma. In multivariate analysis, ileostomy had a lower risk of developing PSH than did colostomy; age mote than 60 years and peristomal complications at the time of stoma creation increased the risk. Only 24% of patients with PSH were free from symptoms related to the hernia. The main complaints were pain (35%), difficulties in fitting a stomal appliance with leakage (28%); 114 patients (56%) underwent operative repair. The morbidity rate of reoperation was 33%, and 57 patients (52%) had recurrence of PSH within an average of 6 months. Transposition of the stoma to another location and the use of prosthetic mesh decreased recidivism AF recurrence? CONCLUSION: PSH aggravates the inherently diminished quality of life of stoma patients. There are many proposed surgical treatments but recurrence occurs in more than half of patients. Randomized trials on the treatment of PSH are nonexistent. The use of a prosthetic mesh may reduce the risk of recurrence. The prophylactic use of prophylactic mesh at the time of initial stoma formation is a strategy worthy of consideration.

Blood platelets and inflammation: their relationship with liver and digestive diseases.

An expansion of knowledge from basic and clinical research has highlighted the critical role of platelets in inflammation and tissue repair in addition to their established contribution to hemostasis. Activated platelets are a rich source of mediators participating to inflammation and tissue regeneration. Platelet-derived microparticles recapitulate essential platelet functions and their contribution to the pathogenesis of inflammatory diseases has been emphasized. Recent findings suggest that platelets are both friends and foes for the liver. Platelets are essential to liver regeneration, platelet-derived serotonin being critical. However platelets can also exacerbate liver damage, as in immune-mediated injury. The dual role of platelets has recently been exemplified in animal models of liver fibrosis. Platelets release profibrogenic mediators, such as CXC Chemokine Ligand 4, that is instrumental in the progression of liver fibrosis. On the other hand, thrombocytopenia aggravates liver fibrosis, an outcome linked to the downregulation of hepatic stellate cell collagen production by platelet derived hepatocyte growth factor. CD154, a key molecule in inflammation, is expressed by platelets and is a pathogenic mediator in inflammatory bowel disease. Here, we summarize some of the mechanisms linking platelets with inflammation and comment few recent articles indicating why platelets may prove to be important pathogenic mediators in liver and gastrointestinal diseases.

[Expression of the elastic fibers components during the fœtal liver development]. / Expression des composants de la matrice élastique au cours du développement hépatique fœtal.

Elastic fibers are composed of microfibrils containing fibrillin-1 and an elastic component, elastin. Microfibrils may not be associated with elastin. In the adult liver, fibrillin-1 and elastin are coexpressed within the stroma and portal tracts vessel walls. Fibrillin-1 is expressed alone around the bile ducts and within the Disse space. There is little work that has studied the elastic fiber organization during the fœtal liver development. Here, we studied the expression of fibrillin-1 and elastin by immunohistochemistry on 20 cases of fœtal liver. During the development of the portal tract, the two components are coexpressed on interstitial elastic fibers and within vessel walls. Fibrillin-1 is expressed alone around the bile structures during their maturation. Unlike adult liver, fibrillin-1 is expressed on thin and very irregular microfibrils within the Disse space. Our study shows that the elastic matrix development in the portal tract follows the development of the different structures, notably biliary structures. In the Disse space, microfibrils are not continuous. Their maturation may be in relation with the change of the hepatic blood flow after birth.

Celiac disease: from oral tolerance to intestinal inflammation, autoimmunity and lymphomagenesis.

Celiac disease is a multifactorial disorder and provides a privileged model to decipher how the interplay between environmental and genetic factors can alter mucosal tolerance to a food antigen, lead to chronic intestinal inflammation, and ultimately promote T-cell lymphomagenesis. Here we summarize how HLA-DQ2/8 molecules, the main genetic risk factor for this disease can orchestrate a CD4(+) T-cell adaptive immune response against gluten, and discuss recent data which shed light on the innate and adaptive immune stimuli that collaborate to induce a proinflammatory TH1 response, a massive expansion of intraepithelial lymphocytes, and a cytolytic attack of the epithelium. The intestinal immune response driven in genetically predisposed patients by chronic exposure to gluten emerges as the pathological counterpart of normal acute intestinal responses to intracellular pathogens.

Expression of thrombopoietin by umbilical vein endothelial cells.

Current data suggest that the primary source of thrombopoietin (TPO) is the liver. Extra-hepatic sites for TPO production have been demonstrated essentially through the study of the expression of TPO mRNA. In this work, we report that TPO is expressed at low levels by endothelial cells (EC) derived from the umbilical vein (HUVEC). Both TPO mRNA and the protein are expressed and the protein is functional as assessed by biological assay. Expression of TPO by HUVEC may be useful to study the regulation of the production of this cytokine and to understand the apparent specific interactions between mature megakaryocytes and EC in the bone marrow.

CD40-ligand stimulates myelopoiesis by regulating flt3-ligand and thrombopoietin production in bone marrow stromal cells.

CD40 ligand (CD40L)/CD40 interactions play a central role in T-cell-dependent B-cell activation as previously shown by in vitro studies, the phenotype of CD40L knockout mice and the defective expression of CD40L in patients who have X-linked immunodeficiency with hyper-IgM. The distribution of CD40 in cells other than of myeloid and lymphoid lineages has suggested additional functions for this receptor/ligand couple. Here we show that CD40L stimulates myelopoiesis with a noticeable effect on megakaryocytopoiesis in cocultures of hematopoietic progenitor cells and bone marrow stromal cells. These results suggest a mechanism by which T-cell or platelet-associated or soluble CD40L may regulate myelopoiesis. (Blood. 2000;95:3758-3764)

Differential effect of interferon alpha on chronic myelogenous leukaemia and normal haematopoietic progenitors in a stromal cell co-culture context: role of the flt3 ligand.

Interferon alpha (IFN-alpha) is used to treat chronic myelogenous leukaemia (CML) patients. However, its target(s) remain(s) unknown. One possibility is that there is a differing sensitivity of the leukaemic from the normal colony-forming cell (CFC) compartments to IFN-alpha. Co-cultures of progenitors with stromal cells provide a valuable tool to dissect direct and indirect activities of IFN-alpha. In this study, we have used endothelial cells (EC) as a source of stromal cells. In co-cultures of normal progenitors with EC, IFN-alpha increased the generation of clonogenic cells, mainly via an increased production of flt3 ligand (FL) by EC. In contrast, in co-cultures of CML progenitors with EC, IFN-alpha inhibited the generation of clonogenic cells, mainly by direct inhibition on the progenitors, the up-regulation of FL production by stromal cells being unable to compensate for the direct inhibitory effects of IFN-alpha. These data provide evidence for a differential effect of IFN-alpha on the growth of CML and normal CFC cells in a stromal context and suggest that an alteration in the response of CML progenitor cells to FL is important in the explanation of this differential effect.

Effect of antisense oligonucleotides on CD34+ cells from chronic myeloid leukemia.

Chronic myeloid leukemia (CML) is a hematopoietic stem cell disorder characterized by a specific hybrid gene BCR-ABL (formed as a result of t(9;22)). This leads to two possible mRNA usually present in leukemic cells, either B2A2 or B3A2. Targeting these mRNA by antisense oligonucleotides (AS) might offer the opportunity to decrease leukemic growth. We have tested the ability of AS to inhibit the in vitro proliferation of CD34 positive (CD34+) blood cells from 16 patients with newly diagnosed CML. CD34+ cells were isolated by an immunomagnetic technique and incubated for 16 to 18 hours with an 18 mer AS (0.25 mM). Sense oligonucleotides served as controls. The effects of AS were evaluated by clonogenic test (production of CFU-GM). Moreover, colonies were picked out and studied by RT-PCR to analyse the presence of BCR-ABL transcript. For nine patients with B3A2 transcript, the median inhibition of CFU-GM formation at day 14 was 64.0 +/- 11.2% (68.0 +/- 11.4% at day 21) and for the seven patients with a B2A2 transcript: 59.0 +/- 11.4% (72.5 +/- 12.0% at day 21). AS showed no effect on CD34+ cells from three normal volunteer donor cells. However, for every patient studied, colonies picked out remained BCR-ABL positive with the RT-PCR technique.

The rat Mist1 gene: structure and promoter characterization.

Transcription factors of the basic Helix-Loop-Helix (bHLH) protein family play key roles in several developmental processes. Mist1 belongs to this group of proteins and shares several properties with the other family members. For example, Mist1 is capable of dimerization with the ubiquitously expressed E2A bHLH proteins and exhibits a strong DNA-binding activity to the core E-box sequence. Using in-situ hybridization and Northern blot hybridization, Mist1 mRNA has been detected in a variety of embryonic and adult rodent tissues. To understand the molecular mechanisms involved in the expression of the gene, we have cloned the rat Mist1 gene and analyzed 2.5 kb of its 5' flanking region. The Mist1 gene spans over 5 kilobases and is composed of two exons separated by a unique intron. The entire coding region is localized in the second exon. Sequence analysis of the promoter region indicated an absence of TATA-box or CAAT-box sequence, but several consensus Sp1-binding sites were present near the transcription start site. Deletion analysis of the promoter region identified a 272 bp proximal fragment to be sufficient to drive expression of a reporter gene in NIH3T3 fibroblasts. Subsequent deletion of potential Sp1 sites results in a marked decrease in promoter activity. Electrophoretic mobility shift assays revealed that Sp1 binds to two different regions in the proximal promoter, a typical Sp1 site located at (-38; -33) and a G/C-rich region between (-67; -62). These data suggest that the basal expression of this TATA-less gene might be driven by general transcription factors, such as Sp1.

Increased liposome-mediated gene transfer into haematopoietic cells grown in adhesion to stromal or fibroblast cell line monolayers.

We investigated transfection rates of CD34+ haematopoietic progenitor cells (HPC) or haematopoietic cell lines (TF-1, KG1a and K562) using the LacZ gene as a reporter and cationic liposomes. The transfection efficiency of CD34+ haematopoietic progenitor cells (HPC) or TF-1, KG1a and K562 grown in suspension is very low (average percentage of 0.013 for HPC and 0.03 for cell lines). Adhesion of HPC or cell lines to plates by immunological or physical methods significantly enhances transfection efficiency; however, the percentage of transfected cells still remained low. We found that adhesion of TF-1, KG1a and K562 HC to MS-5 stroma cells or NIH-3T3 fibroblast cells increased transfection efficiency. Under these conditions transfection is achieved in 11.2-25% (mean 18.30%) for the cell lines and 13.6% (range 8.2-24.2%) for CD34+ HPC. These results indicate that liposome-mediated transfection of HC is significantly increased when cells are grown in adherence to stroma or fibroblast monolayers.

Expression of Flt3-ligand by the endothelial cell.

Flt3-ligand (FL) is a cytokine that is of paramount importance in the proliferation of primitive hematopoietic progenitors. In this study, we show that endothelial cells (EC) produce large amounts of soluble FL and express a membrane-bound form of the molecule. Bone marrow microvascular EC also produce FL, suggesting that EC are an important source of FL in the bone marrow. High concentrations of FL in EC supernatants contrast with its undetectable levels in long-term bone marrow cultures. A single mRNA for FL is detected, suggesting that soluble FL derives from the membrane-bound species by proteolytic release. FL mRNA is stable with a half-life of about 3 h. II-1alpha increases FL mRNA levels and membrane and soluble FL expression. Glucocorticoids, known inhibitors for many hematopoietic growth factors do not down-regulate the expression of FL. On the contrary, GC increase the expression of both species of FL. The neutralization of FL in cocultures EC/ hematopoietic progenitors results in an acceleration of the maturation of the progenitors. IFN-alpha, MIP-1 alpha and TGF-beta stimulate production of membrane-bound and soluble FL. This stimulation is essential to explain their modulatory effect on the generation of clonogenic cells in cocultures EC/hematopoietic progenitors. Leukemia (2000) 14, 153-162.

Expansion and differentiation of haemopoietic progenitor cells on endothelialized hydroxyapatite under static conditions.

We have analysed the potential of an endothelialized hydroxyapatite matrix (HAP), a synthetic bone substitute, as a cellularized support for the expansion of haemopoietic progenitor cells. Scanning electron microscopy (SEM) showed that the endothelial cells (EC) tended to form a monolayer fitting closely to the matrix, and that the progenitors adhered to the EC layer. Endothelialized HAP supported the proliferation and differentiation of the progenitors with the addition of the exogenous cytokines IL-1, and IL-3. The expanded cell population essentially belonged to the myeloid and monocytic lineages, with a smaller percentage of the megakaryocyte lineage. In comparative experiments CD34+ progenitors were expanded on endothelialized tissue culture flasks, and a significant higher viability of the expanded cells was found with the endothelialized HAP. A high percentage (approx. 40%) of mature granulocytes was generated in accordance with the presence of differentiating cytokines such as G-CSF and GM-CSF at high concentrations in the coculture medium. Other cytokines that could be detected were IL-6, M-CSF, SCF, flt3-ligand. More than 50% of the expanded cell population was able to phagocytose bacteria and to generate an oxydative burst. These data indicate that cellularized HAP may be a useful matrix for stromal cell-based expansion systems.

Leukaemia inhibitory factor expression is inhibited by glucocorticoids through post-transcriptional mechanisms.

Leukaemia inhibitory factor (LIF) is a pleiotropic cytokine which is involved in the regulation of the immune response and haematopoiesis. The authors investigated the regulation of the expression of LIF by glucocorticosteroids (GC). Endothelial cells (EC) constitutively produce LIF and this production is enhanced by interleukin 1 (IL-1). GC were found to inhibit the basal production of LIF by EC and to suppress its IL-1-induced augmentation. Whether corticosteroids suppress LIF production by blocking transcription of LIF mRNA, or by blocking LIF synthesis at a post-transcriptional level was examined. Northern blot hybridization analysis demonstrated that GC act mainly by decreasing the LIF mRNA level. In the presence of translation inhibitors a superinduction of LIF mRNA was observed. Dexamethasone (DEX) at a concentration of 1 microM was responsible for a rapid increase in the degradation rate of LIF mRNA which resulted in reducing its level by more than 50% within 2 h, whereas the transcription rate of LIF gene was not significantly altered in these conditions. These results demonstrated that GC inhibit LIF mRNA expression mainly by increasing the turnover rate of the LIF mRNA. The early LIF mRNA destabilizing activity of GC was translation dependent as shown by experiments with protein translation inhibitors. The results indicate that corticosteroids are inhibitors of LIF expression and that this inhibition mainly occurs through post-transcriptional mechanisms.

Endothelial cell support of hematopoiesis is differentially altered by IL-1 and glucocorticoids.

We investigated the ability of endothelial cells (EC) to support hematopoiesis in contact and non-contact cocultures with isolated CD34+ or CD34/CD38low cells. In the absence of exogenous cytokines, umbilical vein EC (HUVEC) efficiently support proliferation of hematopoietic cells and generation of colony-forming cells (CFC). Cytokines (IL-6, LIF, G-CSF, GM-CSF, M-CSF, but not IL-1, IL-3, IL-7) were detected in HUVEC coculture supernatants. Neutralization of these cytokines profoundly inhibited the ability of EC supernatants to support the differentiation of hematopoietic progenitors and led to an accumulation of immature cells. Contact cocultures were significantly more efficient than non-contact cocultures. The expanded cell population essentially belonged to the myeloid and monocytic lineages. Contact cocultures generated cells expressing the CD61 or CD41 antigens. Interleukin-1alpha (IL-1alpha) augmented EC capacity to support hematopoiesis, this property resulting from the upregulation of cytokine expression. Glucocorticoids (GC) reduced this capacity by downregulating the biosynthesis of cytokines by EC and not by a direct effect on the progenitor cells. EC from the bone marrow microvasculature (BMEC) support the proliferation and the differentiation of hematopoietic progenitors. Synergistic increase in progenitor cells expansion and generation of CFC occurred when EC cocultures were added with exogenous cytokines. Supernatants of IL-1alpha-stimulated EC potentiated the effects of an association of IL-1, IL-3, IL-6, LIF, SCF, Flt3-ligand, TPO, G-CSF, GM-CSF, M-CSF and IL-11 on the proliferation of hematopoietic progenitors suggesting that EC may produce other soluble growth factors potentiating the action of the above set of cytokines.

Ex vivo cytokine expansion of peripheral blood Ph-negative cells in chronic myeloid leukaemia.

Hematopoietic progenitors can be expanded ex vivo in the presence of various cytokine combinations. Since normal early progenitor or stem cells persist in the blood and bone marrow of patients with Philadelphia chromosome [Ph]-positive chronic myeloid leukaemia (CML), the selection of normal (Ph-negative) progenitor cells from CML patients would be of considerable clinical value for ex vivo purging and autologous transplantation. To obtain these cells, CD34-positive (progenitor) cells from the peripheral blood (PB) of CML patients were either pretreated or not with 5-fluorouracil (5FU) and then grown in suspension culture for 7 days with a combination of cytokines. We compared different combinations of cytokines containing interleukin-1 alpha (IL1), interleukin-3 (IL3), stem cell factor (SCF), leukemia inhibitor factor (LIF), Flt3-ligand (FLT3L), and thrombopoietin (TPO). 5FU decreased cell proliferation in the liquid culture but concurrently increased the expansion of CFU-GM. While the addition of cytokines such as FLT3L and TPO improved CFU-GM expansion. FISH and RT-PCR analysis showed that this method significantly favored a higher frequency of Ph-negative cells after expansion in liquid culture. Therefore ex vivo expansion of putatively normal hematopoietic progenitor cells from cytapheresis is feasible in CML.