[Treatment by stem cell therapy of erectile dysfunction of diabetic origin: State of the art]. / Traitement par les cellules souches de la dysfonction érectile d'origine diabétique : état des lieux.

PURPOSE: Review of various publications on stem cell therapy to treat erectile dysfunction of diabetic origin. MATERIAL AND METHODS: Bibliographic search in PUBMED performed using the keywords cell therapy strain/erectile dysfunction associated with diabetes. Among the 51 articles obtained from the PUBMED research, we selected 16 articles for their specificity of studying erectile dysfunction (DE) related to diabetes. RESULTS: Different types of stem cells have been studied: adipose derived mesenchymal stem cells/bone marrow derived mesenchymal stem cells as well as progenitor endothelial cells. The experimental protocols are quite similar from one study to the next with nevertheless some specifications concerning the studied cells and the monitoring of the latter. Intracavernous pressure (ICP) measured after the injection of stem cells into the corpus cavernosum was always significantly higher than the control populations. The addition of certain growth factors to stem cells by gene transfection improve the efficacy of the cells. No ideal tracking markers of the cells have been identified. CONCLUSION: The positive effect of the injection of stem cells on the ICP belongs to the cellular trans-differentiation effect but especially to the paracrine effects which have not yet been completely elucidated.

Chimerism analysis following nonmyeloablative stem cell transplantation using a new cell subset separation method: Robosep.

Chimerism analysis has become an important tool to manage patients in the peri-transplant period of allogenic stem cell transplantation. During this period, cells of donor and host origin can coexist and increasing proportion of cells of host origin is considered as a recurrence of the underlying disease. We currently performed chimerism analysis on separate peripheral blood cell subsets, lymphocytes and granulocytes. To improve our isolation method, a new automated device from Stem Cell Technology Roboseptrade mark was tested and compared to our manual separation technique. The results obtained on T cell purification showed an improvement of the purity (98.42% with Robosep vs. 92.42% with the manual technique Rosettesep) and of the recovery (63.43% with Robosep and 38% with Rosettesep). The results were significantly improved on patient samples with less than 10% CD3 positive cells (purity: 90% vs. 44.44%; recovery: 73.79% vs. 43.98%). Granulocytes separation was based on CD15 expression. The results showed an improvement of the purity with Robosep (96.90% vs. 86.20% with the manual technique Polymorphprep) but the recovery was impaired (35.2% vs. 52.30%). Using a myeloid (CD66/CD33) cocktail, recovery was improved with the Robosep device (64.04% with the myeloid cocktail vs. 22.4% with the CD15 cocktail). Our data demonstrated that Robosep allowed a performant cell purification in the early period post-transplantation even for populations representing less than 10% of the peripheral blood cells.

[Persistent thrombocytopenia in a child: morphological examination of blood platelets established the diagnosis of Wiskott-Aldrich syndrome]. / Thrombopénie persistante chez un enfant: l'examen morphologique des plaquettes a conduit au diagnostic de syndrome de Wiskott-Aldrich.

Thrombocytopenia frequently occurs in laboratory practice. The present work illustrates, through the presentation of a case report of Wiskott-Aldrich syndrome, the difficulties encountered to identify and characterize thrombocytopenia. The clinicobiological validation of a low platelet count involves both the biologist, who must assume the validation of numeration while mentioning the morphological characteristics of the platelets and other blood cells, as well as the physician who has to interpret these data according to the clinical context.

Viral-specific T-cell transfer from HSCT donor for the treatment of viral infections or diseases after HSCT.

Allogeneic hematopoietic stem cell transplantation (HSCT) is a curative option for treatment of some malignant and non-malignant hematological diseases. However, post-HSCT patients are severely immunocompromised and susceptible to viral infections, which are a major cause of morbidity and mortality. Although antiviral agents are now available for most types of viral infections, they are not devoid of side effects and their efficacy is limited when there is no concomitant antiviral immune reconstitution. In recent decades, adoptive transfer of viral-specific T cells (VSTs) became an alternative treatment for viral infection after HSCT. However, two major issues are concerned in VST transfer: the risk of GVHD and antiviral efficacy. We report an exhaustive review of the published studies that focus on prophylactic and/or curative therapy by donor VST transfer for post-HSCT common viral infections. A low incidence of GVHD and a good antiviral efficacy was observed after adoptive transfer of VSTs from HSCT donor. Viral-specific T-cell transfer is a promising approach for a broad clinical application. Nevertheless, a randomized controlled study in a large cohort of patients comparing antiviral treatment alone to antiviral treatment combined with VSTs is still needed to demonstrate efficacy and safety.

Introduction to endothelial cell biology.

Vascular endothelial cells form a monocellular layer on blood vessel walls with an estimated mass of 1.5 kg. One of the roles of endothelial cells is to control the hemodynamics through various metabolic activities affecting homeostasis, vascular tonus, blood fluidity, coagulating properties and blood cell adhesion. In other respects thousands of studies have underlined the crucial role of local blood flow conditions on their properties. However, the hemodynamic forces are different according to the anatomical site and to the type of blood vessels (arteries, veins, venules, ...). In microcirculation, the endothelial cells in the venules are particularly active and constitute the physiological site of liquid exchange (permeability) and above all cellular transit. During critical ischemia, the post-capillary venules are deeply involved. In other respects the properties of endothelial cells may be impaired in many diseases as atherosclerosis, hypertension, inflammation and metabolic diseases.

Mechanobiology of mesenchymal stem cells: Which interest for cell-based treatment?

Thanks to their immune properties, the mesenchymal stem cells (MSC) are a promising source for cell therapy. Current clinical trials show that MSC administrated to patients can treat different diseases (graft-versus-host disease (GVHD), liver cirrhosis, systemic lupus, erythematosus, rheumatoid arthritis, type I diabetes…). In this case, the most common mode of cell administration is the intravenous injection, and the hemodynamic environment of cells induced by blood circulation could interfere on their behavior during the migration and homing towards the injured site. After a brief review of the mechanobiology concept, this paper will help in understanding how the mechanical environment could interact with MSC behavior once they are injected to patient in cell-based treatment.

Cell and tissue engineering and clinical applications: an overview.

Most human tissues do not regenerate spontaneously; this is why cell therapies and tissue engineering are promising alternatives. The principle is simple: cells are collected in a patient and introduced in the damaged tissue or in a tridimentional porous support and harvested in a bioreactor in which the physico-chemical and mechanical parameters are controlled. Once the tissues (or the cells) are mature they may be implanted. In parallel, the development of biotherapies with stem cells is a field of research in turmoil given the hopes for clinical applications that it brings up. Embryonic stem cells are potentially more interesting since they are totipotent, but they can only be obtained at the very early stages of the embryo. The potential of adult stem cells is limited but isolating them induces no ethical problem and it has been known for more than 40 years that bone marrow does possess the regenerating functions of blood cells. Finally, the properties of foetal stem cells (blood cells from the umbilical cord) are forerunners of the haematopoietic system but the ability of these cells to participate to the formation of other tissues is more problematic. Another field for therapeutic research is that of dendritic cells, antigen presenting cells. Their efficiency in cell therapy relies on the initiation of specific immune responses. They represent a promising tool in the development of a protective immune response against antigens which the host is usually unable to generate an efficient response (melanomas, breast against cancer, prostate cancer, ..). Finally, gene therapy, has been nourishing high hopes but few clinical applications can be envisaged in the short term, although potential applications are multiple (haemophilia, myopathies, ..). A large number of clinical areas stand as candidates for clinical applications: leukaemia and cancers, cardiac insufficiency and vascular diseases, cartilage and bone repair, ligaments and tendons, liver diseases, ophthalmology, diabetes, neurological diseases (Parkinson, Huntington disease, ..), .. Various aspects of this new regenerative therapeutic medicine are developed in this work.

Stem cells and vascular regenerative medicine: A mini review.

Most human tissues do not regenerate spontaneously, which is why "cell therapy" are promising alternative treatments. The Principe is simple: patients' or donors' cells are collected and introduced into the injured tissues or organs directly or in a porous 3D material, with or without modification of their properties. This concept of regenerative medicine is an emerging field which can be defined as "the way to improve health and quality of life by restoring, maintaining, or enhancing tissue and organ functions".There is an extraordinarily wide range of opportunities for clinical applications: artheropathies, diabetes, cartilage defects, bone repair, burns, livers or bladder regeneration, organs reconstruction (lung, heart, liver ...) neurodegenerative disorders, sepsis ...  Different stem cells (SC) with different potential can be used and characterised (totipotent, mesenchymal of different origins, especially those present in tissues...). Today it is undeniable that cells like bone marrow, adipose tissue or Wharton Jelly stem cells, are of potential interest for clinical applications because they are easily separated and prepared and no ethical problems are involved in their use.In this paper some potential clinical applications in the vascular field are considered: peripheral arteriopathy in diabetic patients, cardiac insufficiency, traitment of erectile dysfunction, or organ regeneration with liver as example. But the regeneration of tissue or organ is and will remain a challenge for the future development of cell therapy. Many problems remain to be solved that could lead to the development of innovative strategies to facilitate cell differentiation, increase the yield of cells and ensure a standardised product, overcome the risks of teratogenic effects and/or immune reactions, enable grafting via direct cell or biotissue transplantation and avoid legal issues involved in national regulations.

Osteopetrotic mouse stroma with thrombopoietin, c-kit ligand, and flk-2 ligand supports long-term mobilized CD34+ hematopoiesis in vitro.

OP-9 cells are stromal cells derived from macrophage colony-stimulating factor (M-CSF)-deficient osteopetrotic mice. To evaluate the OP-9 capability to sustain long-term hematopoiesis, we reported the expansion of granulocyte colony-stimulating factor (G-CSF)-mobilized human peripheral blood (PB) CD34(+) cells in co-culture with murine OP-9 and MS-5 stromal cells, either transfected with various combinations of adenovectors (Ad) expressing c-kit ligand (KL) (either soluble or transmembrane form), thrombopoietin (TPO), flt-3/flk2 ligand (FL), and granulocyte-macrophage (GM)-CSF or with weekly addition of these cytokines. Expression of TPO as well as association of TPO, FL, and KL increased progenitor cell and week-6 cobblestone area forming cell (CAFC) production in all stromal co-cultures. Similar progenitor expansion was obtained by weekly addition of soluble cytokine. Five weeks of co-culture with OP9 and TPO, FL + KL resulted in the greatest expansion of progenitor cells and week-6 CAFC as measured by secondary assay on MS-5. In contrast to MS-5 and TPO or TPO + FL + KL cultures where hematopoiesis declined by week 4, progenitor as well as week-6 CAFC expansion continued for over 3 months in TPO + FL + KL OP9 cocultures. This was associated with decrease of CD14(+) macrophage production. The addition of human macrophage (M)-CSF or CD14(+) cells to the co-culture decrease progenitor and stem cell expansion; however, murine M-CSF to OP-9 co-cultures did not decrease progenitor expansion. High levels of stromal-derived factor-1 (SDF-1) production by MS-5 and low or absent production by OP-9 may account for stem cell adhesion and CAFC formation in the former cultures and the predominance of stem and progenitor cells in the nonadherent fraction in the latter cultures.

Stem Cells and Regenerative Medicine: Myth or Reality of the 21th Century.

Since the 1960s and the therapeutic use of hematopoietic stem cells of bone marrow origin, there has been an increasing interest in the study of undifferentiated progenitors that have the ability to proliferate and differentiate into various tissues. Stem cells (SC) with different potency can be isolated and characterised. Despite the promise of embryonic stem cells, in many cases, adult or even fetal stem cells provide a more interesting approach for clinical applications. It is undeniable that mesenchymal stem cells (MSC) from bone marrow, adipose tissue, or Wharton's Jelly are of potential interest for clinical applications in regenerative medicine because they are easily available without ethical problems for their uses. During the last 10 years, these multipotent cells have generated considerable interest and have particularly been shown to escape to allogeneic immune response and be capable of immunomodulatory activity. These properties may be of a great interest for regenerative medicine. Different clinical applications are under study (cardiac insufficiency, atherosclerosis, stroke, bone and cartilage deterioration, diabetes, urology, liver, ophthalmology, and organ's reconstruction). This review focuses mainly on tissue and organ regeneration using SC and in particular MSC.

Stem cells and applications: a survey.

Since the 1960s and the therapeutic use of hematopoietic stem cells of bone marrow origin, there has been increasing interest in the study of undifferentiated progenitors that have ability to proliferate and differentiate in different tissues. Different stem cells (SC) with different potential can be isolated and characterised. Despite the promise of embryonic stem cells, in many cases, adult stem cells provide a more interesting approach to clinical applications. It is undeniable that mesenchymal stem cells (MSC) from bone marrow, adipose tissue or MSC of Wharton Jelly, which have limited potential, are of interest for clinical applications in regenerative medicine because they are easily separated and prepared and no ethical problems are involved in their use.During the last 10 years, these multipotent cells have generated considerable interest and in particular have been shown to escape allogeneic immune response and be capable of immunomodulatory activity. These properties may be of a great interest for regenerative medicine. Different clinical applications are under study (cardiac insufficiency, atherosclerosis, stroke, bone, cartilage, diabetes, ophthalmology, urology, liver, organ's reconstruction…).

In vivo platelet kinetics in 31 diabetic patients. Correlation with the degree of vascular impairment.

In vivo platelet survival was estimated, in 31 diabetic patients, using 51Cr-labelled autologous platelets. A mathematical analysis attempted to measure the "potential" life span (senescence process) and the degree of a superimposed aleatory destruction (consumption process). The potential platelet life span in diabetics did not differ from normal values. However, excessive consumption was observed in one third of the studied cases, without correlation with the age of the patients, the clinical duration of diabetes, and the degree of vascular impairment. Thus, platelet kinetic studies did not provide presently useful indications, in a particular patient, regarding the prognosis of the vascular disease, and the justification of anti-aggregant therapy.

Complete recovery from Cryptosporidium parvum infection with gastroenteritis and sclerosing cholangitis after successful bone marrow transplantation in two brothers with X-linked hyper-IgM syndrome.

We describe two brothers who suffered from hyper-IgM syndrome (HIGM1) with similar clinical features: recurrent infections, especially cryptosporidium gastroenteritis with cholangitis. Their activated T cells did not express CD40L. Nucleotide sequencing revealed a mutation in both boys with respect to intron 4 and exon 5 boundaries of the CD40L gene in Xq26. They underwent successful bone marrow transplantation (BMT) from HLA-geno-identical siblings. The Cryptosporidium infection and cholangitis resolved thereafter. At 6 months after BMT, expression of CD40L on activated T lymphocytes was normal. After 1 year, both boys are well, and immune reconstitution has improved. Based on these two successful experiences, BMT with a genoidentical sibling seems a reasonable therapeutic approach for HIGM1, if Cryptosporidium infection occurs.

Multicenter randomized evaluation of a nutritional education software in obese patients.

OBJECTIVE: To study the efficacy of the nutritional education software, Nutri-Expert, in the management of obese adult patients. MATERIAL AND METHODS: Two groups of obese patients were followed up over one year in a randomized study: the first group received close traditional management (seven nutritional visits over the year, with physicians and dietitians conjointly) and the second one also used at home by Minitel the Nutri-Expert system. 557 patients were enrolled in the study by 16 French centers of diabetology and nutrition. Body mass index (BMI), tests of dietetic knowledge, dietary records and centralized biological measurements were assessed at inclusion, 6 and 12 months. 341 patients were evaluable at the end of the year. RESULTS: The group using Nutri-Expert scored significantly better in the tests of dietetic knowledge than the control group. For all patients, nutritional education led to a significant improvement in BMI, dietary records and biological measurements, without significant difference between the two groups. Five years after the end of the study, the weight of 148 patients was recorded; mean BMI was significantly lower than the initial value but there was no significant difference between the two groups. CONCLUSION: In the management of obese patients, Nutri-Expert system has a role to play in reinforcing nutritional knowledge; if regular follow-up is not possible, or if a large series of obese patients is to be treated, Nutri-Expert could partly replace traditional management, for example between visits.

CD34+ cells homing: quantitative expression of adhesion molecules and adhesion of CD34+ cells to endothelial cells exposed to shear stress.

To investigate the function of the main adhesion receptors (CD62L, CD49d, CD49e, CD11b and CD18) on CD34+ cells during homing, their expression was quantified by flow cytometry using calibration beads. CD34+ cells were isolated from bone-marrow (BM), cord blood (CB) or peripheral blood (PB) from patients with myeloma. As this process might mimic the mature leukocyte migration, we also observed the effect of exposing endothelial cells to shear stress (7 dyn/cm(2)) on the adhesion of CB CD34+ cells. The proportion of CD34+/CD62L+ cells was greater in PB than in BM (p<0.05). Likewise, we found a significantly greater expression of CD62L receptor on PB cells compared to BM cells (p<0.05) and on BM cells compared to CB cells (p<0.05). The proportions of CD34+/CD49d+ cells and CD34+/CD49e+ cells were significantly higher in the BM and CB than in PB. However, no significant difference in CD49d or CD49e antigen densities was observed. The beta_2 integrins (CD11b and CD18) receptors are also implicated in CD34+ cells homing to BM. No significant variation in CD34+/CD11b+ and CD34+/CD18+ cells frequency was noted. However quantitative analysis revealed that CD18 was more strongly expressed on BM cells than on PB and CB cells. The adhesion assay showed that fluid flow may favour a firm adhesion of CB CD34+ cells to endothelial cells whereas static conditions just allowed CD34+ cells sedimentation. In conclusion, quantitative expression of the main receptors on CD34+ cells indicates that the three main sources of CD34+ cells currently used for transplantation have neither the same phenotype nor the same number of antigenic sites for a receptor. So, we hypothesize that migrational capacity of these cells might be different. Moreover, it seems that shear stress could favor adhesion of CD34+ cells to endothelial cells.

In vitro study of intracellular IL-1beta production and beta1 integrins expression in stimulated chondrocytes--effect of rhein.

The purpose of the present study was to investigate the intracellular IL-1beta production and beta1 integrins (alpha4/beta1 and alpha5/beta1) expression on chondrocytes. Chondroytes monolayer (human chondrosarcoma cell line HEM-C55) were incubated for 12, 24 and 48 hours in the presence of Tumor Necrosis Factor-alpha (TNF-alpha, Sigma, France) or recombinant human IL-1alpha (rh-IL1alpha, Becton Dickinson, France). After direct immunolabelling, cells were either analyzed on FACScan flow cytometer (Becton Dickinson, France), or observed under an epi-fluorescence inverted microscope equipped with the CellScan EPR optical scanning acquisition system (IPLab-Scanalytics, USA). We found that the IL-1beta mean fluorescence intensity in flow cytometry and in 3D microscopy was increased in the presence of TNF-alpha or rh-IL-1alpha, and alpha4/beta1 or alpha5/beta1 expression was higher on stimulated cells than on control cells. On the other hand, we have evaluated the in vitro effects of rhein (10(-5) M, Negma, France), an active metabolite of diacerein, on the intracellular IL-1beta and beta1 integrins expressed by stimulated or no-stimulated chondrocytes. The results indicated that rhein leads to a reduction of IL-1beta synthesis whereas a weak decrease of beta1 integrins receptors expression is observed. From this study, it seems that rhein partially reduce cytokine-induced intracellular IL-1beta production, and it has a weak action on alpha4/beta1 or alpha5/beta1 receptors.

[Quantitative expression of the adhesion receptors VLA-4, VLA-5, L-selectin, MAC-1, and ICAM-1 on the surface of CD34+ cells]. / Expression quantitative des récepteurs d'adhérence VLA-4, VLA-5, L-sélectine, MAC-1, et ICAM-1 à la surface des cellules CD34+.

The aim of this work was to quantify by flow cytometry the main adhesion receptors on CD34+ cells. These cells were isolated from bone marrow (BM) or mobilized peripheral blood (PB). The proportions of CD34+/CD49d+ and CD34+/CD49e+ are weaker on PB cells, without quantitative expression variation. This phenotypic variation may induce CD34+ cells exist from BM into circulation, promoting the mobilization. The homing to the BM implicate the CD62L receptor, which expression was found more frequently and stronger on PB cells than on BM. The CD11b, CD18 and CD54 receptors are implicated in CD34+ cells adhesion to BM micro-environment. No significant variation in CD34+/CD11b+ and CD34+/CD18+ cells frequency was noted. Moreover, CD54 receptor was more frequently expressed on PB cells. Quantitative analysis revealed that CD18 was more strongly expressed on BM than on PB cells. This quantitative variation could promote progenitor adhesion by interacting with stromal cells. Finally, quantitative expression of the main receptors on CD34+ cells provides an original option for studying CD34+ cells during the mobilization, the homing or the adhesion to BM micro-environment.

From hemorheology to vascular mechanobiology: An overview.

Almost all of the cells of the human body are subjected to mechanical stresses. In endothelial cells, mechanical stresses can vary from some milli-Pascal (shear stress) to one ore more Pascal (hydrostatic pressure). Now it is know that mechanical stresses have a decisive part cellular physiology. However, if the main biological effects of mechanical stress are well related, the mechanisms allowed the relation between mechanical stress to physiological phenomenon remain nearly unknown (mechanotransduction phenomenon). In this work, through personal results and published works, the authors considers all the effects of mechanical stresses and the possible hypothesis.

[Introduction to vascular engineering]. / Introduction à l'ingénierie vasculaire.

Vascular engineering developed through advancing knowledge of the physiological and pathological processes operating in cells and tissues subjected to environmental mechanical stress. We review briefly the more relevant cellular and general mechanism characteristic of vascular tissue examining the performances obtained with prosthetic materials (allografts, synthetic grafts) and current research and development of new vessel substitutes (biohybrids or biovessels).

[Treatment of mycosis fungoides by PUVAtherapy. Report on 18 cases (author's transl)]. / Traitement du mycosis fongoïde par la puvathérapie. a propos de 18 cas.

A group of 18 patients with mycosis fungoides (M.F.) was treated by PUVAtherapy. According to VanScott classification, they have been set in:--5 parapsoriasis in large plaques = 5 stages I;--6 stages II;--3 stages III;--2 stages IV;--1Sézary syndrom.--1 erythrodermia. There was complete clearing of 12 patients (66 p. 100); 2 patients (11 p. 100) improved cutaneous lesions without a complete clearing, and there was no response to treatment for 3 patients (22 p. 100) (1 with erythrodermia, 1 with Sézary syndrom and 1 stage IV, and 1 forsaking). The method applied here was different in several points: orally administered methoxalen were given according to the body area (mg/m2) and some of the patients had twice a day puvatherapy. Puvatherapy seems to be the least dangerous and most effective treatment for the patients in the early stages of mycosis fongoides (I, II) and nitrogen mustard, electron therapy, corticoids and even mono- or polychimiotherapy could be associated to puvatherapy, if necessary, for the stages III and IV.