P38 mitogen-activated protein kinase promotes dedifferentiation of primary articular chondrocytes in monolayer culture.

Articular cartilage is an avascular tissue with poor regenerative capacity following injury, a contributing factor to joint degenerative disease. Cell-based therapies for cartilage tissue regeneration have rapidly advanced; however, expansion of autologous chondrocytes in vitro using standard methods causes 'dedifferentiation' into fibroblastic cells. Mitogen-activated protein kinase (MAPK) signalling is crucial for chondrocyte metabolism and matrix production, and changes in MAPK signals can affect the phenotype of cultured cells. We investigated the effects of inhibition of MAPK signalling on chondrocyte dedifferentiation during monolayer culture. Blockade of extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK) signalling caused a significant increase in cartilage gene expression, however, also caused up-regulation of fibrotic gene expression. Inhibition of p38 MAPK (p38) caused a significant up-regulation of collagen type II while suppressing collagen type I expression. P38 inhibition also resulted in consistently more organized secretion of collagen type II protein deposits on cell culture surfaces. Follow-on pellet culture of treated cells revealed that MAPK inhibition reduced cell migration from the pellet. ERK and JNK inhibition caused more collagen type I accumulation in pellets versus controls while p38 inhibition strongly promoted collagen type II accumulation with no effect on collagen type I. Blockade of all three MAPKs caused increased GAG content in pellets. These results indicate a role for MAPK signalling in chondrocyte phenotype loss during monolayer culture, with a strong contribution from p38 signalling. Thus, blockade of p38 enhances chondrocyte phenotype in monolayer culture and may promote more efficient cartilage tissue regeneration for cell-based therapies.

MicroRNA93 regulates proliferation and differentiation of normal and malignant breast stem cells.

MicroRNAs (miRNAs) play important roles in normal cellular differentiation and oncogenesis. microRNA93 (mir-93), a member of the mir106b-25 cluster, located in intron 13 of the MCM7 gene, although frequently overexpressed in human malignancies may also function as a tumor suppressor gene. Using a series of breast cancer cell lines representing different stages of differentiation and mouse xenograft models, we demonstrate that mir-93 modulates the fate of breast cancer stem cells (BCSCs) by regulating their proliferation and differentiation states. In "claudin(low)" SUM159 cells, expression of mir-93 induces Mesenchymal-Epithelial Transition (MET) associated with downregulation of TGFß signaling and downregulates multiple stem cell regulatory genes, including JAK1, STAT3, AKT3, SOX4, EZH1, and HMGA2, resulting in cancer stem cell (CSC) depletion. Enforced expression of mir-93 completely blocks tumor development in mammary fat pads and development of metastases following intracardiac injection in mouse xenografts. The effect of mir-93 on the CSC population is dependent on the cellular differentiation state, with mir-93 expression increasing the CSC population in MCF7 cells that display a more differentiated "luminal" phenotype. mir-93 also regulates the proliferation and differentiation of normal breast stem cells isolated from reduction mammoplasties. These studies demonstrate that miRNAs can regulate the states and fates of normal and malignant mammary stem cells, findings which have important biological and clinical implications.

Breast cancer stem cells are regulated by mesenchymal stem cells through cytokine networks.

We have used in vitro and mouse xenograft models to examine the interaction between breast cancer stem cells (CSC) and bone marrow-derived mesenchymal stem cells (MSC). We show that both of these cell populations are organized in a cellular hierarchy in which primitive aldehyde dehydrogenase expressing mesenchymal cells regulate breast CSCs through cytokine loops involving IL6 and CXCL7. In NOD/SCID mice, labeled MSCs introduced into the tibia traffic to sites of growing breast tumor xenografts where they accelerated tumor growth by increasing the breast CSC population. With immunochemistry, we identified MSC-CSC niches in these tumor xenografts as well as in frozen sections from primary human breast cancers. Bone marrow-derived MSCs may accelerate human breast tumor growth by generating cytokine networks that regulate the CSC population.

[Validation of a sampling method for the channels of a flexible endoscope which is experimentally contaminated]. / Validation d'une méthode de prélèvement des canaux d'un endoscope souple contaminé expérimentalement.

The increase of endoscopic actions and infectious risks explains of interest in endoscopic material maintenance processing. This purpose study was to set to the point and validate a microbiological flexibles endoscopes channels (suction, biopsy, air/water) sample method. An inoculum made of faeces contents material and microorganisms suspension (Pseudomonas aeruginosa or Candida albicans), at low or high concentration, was injected into digestive endoscope's channels. For each microorganism, five trials were carried out at both of the two concentrations. After incubation, a global channels sample was performed with a recovering solution, then a counting was made. A variance analysis (ANOVA) concerning restated measures was performed for the whole results. The mean difference between the number of microorganisms injected and those recovered's is 0.114 log (IC95% [+0.088; +0.140]). It does not vary significantly according to the micro-organisms (p = 0.69) and the inoculum (p = 0.70). The coefficient of variation which is 0.525 (min: 0.010; max: 0.260) shows this method can be reproduced. This technique are allowed to be recommended to evaluate and to validate manuals and automatics endoscopes maintenance processing.

[Automatization of colorimetric serum zinc determination using the Bayer RA 1000 autoanalyzer]. / Automatisation de la détermination colorimétrique du zinc sérique sur Bayer RA 1000.

A colorimetric method for the determination of zinc in serum was adapted for use with the Bayer RA 1000 autoanalyzer. Zinc is dissociated from proteins by guanidinium hydrochloride and complexed with 2-(5-bromo-2-pyridylazo)-5-(N-propyl-N- sulfopropylamino-)phenol (5-Br-PAPS). Precision (within-run CVs from 1.4 to 4.2%; between-run CVs from 3.5 to 8.1%), accuracy (93 to 103%) and detection limit (1.15 mumol/l) were sufficient for routine determinations. Moreover, results obtained with the proposed Bayer RA 1000 method were in good agreement with those obtained by flame atomic absorption spectrometry (FAAS) which could be considered the reference method. For 119 sera, the correlation coefficient r was found to be 0.958 and the regression line was as follows: serum zinc (RA 1000) = 0.96 serum zinc (FAAS) + 0.8 mumol/l. Moreover, 98% of the differences between colorimetry and FAAS were within the range Md +/- 1.96 SDd, where Md is the mean of the differences and SDd is the standard deviation of the differences.

Replacement of ureteric segments by intubated neo-ureterotomies (modified Davis technique) using autologous bladder and omentum in dogs.

In 14 dogs, using a modified Davis technique, excised segments of ureters were replaced by free, partial-thickness bladder grafts vascularised by omentum. Eight of the 14 dogs had satisfactory radiological appearances of the operated upper tracts at 12 months, and 9 of the 14 were unobstructed urodynamically at this time. Graft segments did not appear to contract. Urothelial and smooth muscle regeneration was observed.

Inferior vena caval tamponade following partial hepatectomy for liver injury.

A case of acute obstruction of the inferior vena cava after partial hepatectomy for liver injury is described. The underlying haemodynamics and prevention of this complication are discussed.

Pimozide attenuates lever pressing for water reinforcement in rats.

Rats were trained to lever-press for water on a schedule of continuous reinforcement, then tested every fourth session on five occasions either under conditions of non-reinforcement or following injections of the dopamine receptor blocker pimozide (0.5 or 1.0 mg/kg) or the injection vehicle. The low dose of pimozide did not significantly attenuate responding until the fifth session. The high dose attenuated responding on all occasions, with residual responding decreasing progressively across repeated drug sessions. Responding in the pimozide conditions was never less than that of the non-reinforced control group. Responding in each condition was strongest in the early minutes of a session. After five sessions, rats were switched from the pimozide condition to the non-reinforced condition (or vice-versa) for one additional test day. Decreased responding continued for rats transferred from non-reinforcement to pimozide though not for rats transferred from pimozide to non-reinforcement. These data suggest a role for brain dopamine in behavior; they reflect the same patterns as have been seen with food reinforcement and with several centrally-acting reinforcers.