Nathalie Saulnier

Objectives: The aim of this case series was to collect preliminary data on safety and efficacy of treating cats suffering from refractory feline chronic gingivostomatitis with a single intravenous therapy of cryopreserved placenta‐derived mesenchymal stromal cells.Materials and Methods: We planned the prospective inclusion of cats suffering from refractory chronic gingivostomatitis in three veterinary clinics. All cats received a single infusion of 10×10⁶ cryopreserved cells. Follow‐up evaluations were done at day 15 and at 2‐, 3‐ and 6‐months following infusion. Clinical disease severity was evaluated by dental specialists using a published stomatitis disease activity index scoring system coupled with an owners' assessment questionnaire.Results: All eight cats attended all follow up visits. Cryopreserved ready‐to‐use placenta‐derived cells administered systemically were safe and resulted in notable clinical improvement in all cats as reported by stomatitis disease activity index scoring and owner's survey.Clinical Significance: Infusion of cryopreserved freshly thawed placenta‐derived mesenchymal stromal cells appears to promote clinical and consequently behavioural benefits in cats with refractory chronic gingivostomatitis after having undergone full‐mouth or premolar–molar tooth extraction. Future randomised studies are required to confirm safety and efficacy of this treatment.

As in humans, osteoarthritis (OA) causes considerable economic loss to the equine industry. New hopes for cartilage repair have emerged with the matrix-associated autologous chondrocyte implantation (MACI). Nevertheless, its limitation is due to the dedifferentiation occurring during the chondrocyte amplification phase, leading to the loss of its capacity to produce a hyaline extracellular matrix (ECM). To enhance the MACI therapy efficiency, we have developed a strategy for chondrocyte redifferentiation, and demonstrated its feasibility in the equine model. Thus, to mimic the cartilage microenvironment, the equine dedifferentiated chondrocytes were cultured in type I/III collagen sponges for 7 days under hypoxia in the presence of BMP-2. In addition, chondrocytes were transfected by siRNA targeting Col1a1 and Htra1 mRNAs, which are overexpressed during dedifferentiation and OA. To investigate the quality of the neo-synthesized ECM, specific and atypical cartilage markers were evaluated by RT-qPCR and Western blot. Our results show that the combination of 3D hypoxia cell culture, BMP-2 (Bone morphogenetic protein-2), and RNA interference, increases the chondrocytes functional indexes (Col2a1/Col1a1, Acan/Col1a1), leading to an effective chondrocyte redifferentiation. These data represent a proof of concept for this process of application, in vitro, in the equine model, and will lead to the improvement of the MACI efficiency for cartilage tissue engineering therapy in preclinical/clinical trials, both in equine and human medicine.

The aim of the present study was to investigate whether microarray gene expression analysis can be used to predict lymph node status in gastric cancer. Twenty-nine patients undergoing gastrectomy for cancer were enrolled and subdivided according to the pathologic nodal involvement of their disease (N+ vs N0). Molecular profiling was performed by cDNA microarray on tumor tissue and healthy mucosa. Data were processed to identify differently expressed genes. Selected genes were categorized with gene ontology. Compared to healthy gastric mucosa, 52 genes were differently expressed in N+ patients, and 50 genes in N0 patients. Forty-five genes were similarly regulated in N+ and N0 patients, whereas 12 genes were differently expressed between N+ and N0 patients. Seven genes were exclusively expressed in N+ patients: Egr-1 was upregulated; Claudin-18, AKR1C2, Cathepsin E, CA II, TFF 1, and progastricsin were downregulated. Five genes were exclusively expressed in N0 patients: Complement C5 receptor 1, PLA2/VII, and MMP- 9 were upregulated; MAO-A and ID-4 were downregulated. Microarray analysis could be a valuable tool to identify genes associated with lymph node metastasis in gastric cancer. This technique could improve the selection of patients with locally advanced disease who are candidates for extended lymph node dissection, multimodal treatment options, or alternative therapeutic strategies.

The gene expression pattern of glioblastoma (GBM) is well documented but the expression profile of brain adjacent to tumor is not yet analysed. This may help to understand the oncogenic pathway of GBM development. We have established the genome-wide expression profiles of samples isolated from GBM tumor mass, white matter adjacent to tumor (apparently free of tumor cells), and white matter controls by using the Affymetrix HG-U133 arrays. Array-CGH (aCGH) was also performed to detect genomic alterations. Among genes dysregulated in peritumoral white matter, 15 were over-expressed, while 42 were down-regulated when compared to white matter controls. A similar expression profile was detected in GBM cells. Growth, proliferation and cell motility/adhesion-associated genes were up-regulated while genes involved in neurogenesis were down-regulated. Furthermore, several tumor suppressor genes along with the KLRC1 (a member of natural killer receptor) were also down-regulated in the peritumoral brain tissue. Several mosaic genomic lesions were detected by aCGH, mostly in tumor samples and several GBM-associated mosaic genomic lesions were also present in the peritumoral brain tissue, with a similar mosaicism pattern. Our data could be explained by a dilution of genes expressed from tumor cells infiltrating the peritumour tissue. Alternatively, these findings could be substained by a relevant amount of “apparently normal” cells presenting a gene profile compatible with a precancerous state or even “quiescent” cancer cells. Otherwise, the recurrent tumor may arise from both infiltrating tumor cells and from an interaction and recruitment of apparently normal cells in the peritumor tissue by infiltrating tumor cells.

Multipotent mesenchymal stem cells with extensive self-renewal properties can be easily isolated and rapidly expanded in culture from small volumes of amniotic fluid. These cells, namely, amniotic fluid-stromal cells (AFSCs), can be regarded as an attractive source for tissue engineering purposes, being phenotypically and genetically stable, plus overcoming all the safety and ethical issues related to the use of embryonic/fetal cells. LMP3 is a novel osteoinductive molecule acting upstream to the main osteogenic pathways. This study is aimed at delineating the basic molecular events underlying LMP3-induced osteogenesis, using AFSCs as a cellular model to focus on the molecular features underlying the multipotency/differentiation switch. For this purpose, AFSCs were isolated and characterizedin vitroand transfected with a defective adenoviral vector expressing the human LMP3. LMP3 induced the successful osteogenic differentiation of AFSC by inducing the expression of osteogenic markers and osteospecific transcription factors. Moreover, LMP3 induced an early repression of the kruppel-like factor-4, implicated in MSC stemness maintenance. KLF4 repression was released upon LMP3 silencing, indicating that this event could be reasonably considered among the basic molecular events that govern the proliferation/differentiation switch during LMP3-induced osteogenic differentiation of AFSC.

The prevalence of individuals allergic to latex, exhibiting cross-hypersensitivity with plant-derived food has been frequently reported as the so-called latex-fruit syndrome. Nonetheless, molecular mechanisms underlying allergy to latex and/or fruit are poorly understood. The aims of this study were to identify candidate genes that may be associated with the pathogenesis of allergy to latex and/or vegetable food, and to assess if similar molecular pathways are involved in both types of hypersensitivity. DNA microarray analysis was performed to screen the molecular profiles of peripheral blood mononuclear cells isolated from patients with allergy to latex, to fruit, or with latex-fruit syndrome, and from control healthy subjects. Molecular profiling identified an overlapping dataset of genes commonly regulated in all the atopic patients enrolled in this study, suggesting that similar molecular mechanisms are involved in the pathogenesis of allergy to the fruit and/or latex. Several regulators of the innate and acquired immunity reported to polarize the immunological response towards a Th2-mediated immune response were overexpressed in the patients. Evidences suggested that the expression of T-regulatory cells might be defective in allergic patients, as a consequence of a dysregulation of some inflammatory cytokines. Finally, several transcription factors that may be responsible for the Th1/Th2 imbalance were modulated in allergic patients. This study identified relevant genes that may help to elucidate the molecular mechanisms underlying allergic disease. Knowledges of critical targets, along with transcription factors regulating gene activity may facilitate the development of new therapeutic options.

The mitogen-activated protein kinases (MAPK) ERK1 and ERK2 are among the major signal transduction molecules but little is known about their specific functions in vivo. ERK activity is provided by two isoforms, ERK1 and ERK2, which are ubiquitously expressed and share activators and substrates. However, there are not in vivo studies which have reported a role for ERK1 or ERK2 in HSCs and the bone marrow microenvironment. The present study shows that the ERK1-deficient mice present a mild osteopetrosis phenotype. The lodging and the homing abilities of the ERK1^−/− HSC are impaired, suggesting that the ERK1^−/−-defective environment may affect the engrafment of HSCs. Serial transplantations demonstrate that ERK1 is involved in the maintenance of an appropriate medullar microenvironment, but that the intrinsic properties of HSCs are not altered by the ERK1^−/− defective microenvironment. Deletion of ERK1 impaired in vitro and in vivo osteoclastogenesis while osteoblasts were unaffected. As osteoclasts derive from precursors of the monocyte/macrophage lineage, investigation of the monocytic compartment was performed. In vivo analysis of the myeloid lineage progenitors revealed that the frequency of CMPs increased by approximately 1.3-fold, while the frequency of GMPs significantly decreased by almost 2-fold, compared with the respective WT compartments. The overall mononuclear-phagocyte lineage development was compromised in these mice due to a reduced expression of the M-CSF receptor on myeloid progenitors. These results show that the cellular targets of ERK1 are M-CSFR-responsive cells, upstream to osteoclasts. While ERK1 is well known to be activated by M-CSF, the present results are the first to point out an ERK1-dependent M-CSFR regulation on hematopoietic progenitors. This study reinforces the hypothesis of an active cross-talk between HSCs, their progeny and bone cells in the maintenance of the homeostasis of these compartments.

Several in vitro assays have been proposed to identify cancer stem cells (CSCs), including immunophenotyping, sphere assay and side population (SP) assay. CD133 antigen has been proposed as a CSC marker in colon cancer (CC). However, no functional data are available to date and conflicting results have been reported regarding its role as true CSC marker. Here we set out to identify a molecular signature associated with potential CSC. CD133⁺ cells isolated from the CaCo‐2 CC cell line were analysed by microarray molecular profiling compared to CD133⁻ counterparts. Various differentially expressed genes were identified and the most relevant transcripts found to be over‐expressed in CD133⁺ cells were evaluated by quantitative RT‐PCR in the CD133⁺ fractions isolated from several CC cell lines. In the attempt to find a correlation between putative CSCs, isolated by means of CD133 immunophenotyping and the SP approach, we demonstrated a significant enrichment of CD133⁺ cells within the SP fraction of CC cells, and comparison of the gene expression profiles revealed that Endothelin‐1 (END‐1) and nuclear receptor subfamily 4, group A, member 2 (NR4A2) transcripts are highly expressed in both CD133⁺ and SP fractions of CC cells. Moreover, depletion of CD133 by siRNA induced a significant attenuation of END‐1 and NR4A2 expression levels in CaCo‐2 cells, while expression of all three molecules decreased during sodium butyrate‐induced differentiation. In conclusion, we have identified a molecular signature associated with potential CSCs and showed for the first time the existence of a functional relationship between CD133, END‐1 and NR4A2 expression in colon cancer cells. Copyright © 2011 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.