Phenotypical and functional characteristics of mesenchymal stem cells from bone marrow: comparison of culture using different media supplemented with human platelet lysate or fetal bovine serum.

INTRODUCTION: Mesenchymal stem cells (MSCs) are multipotent cells able to differentiate into several mesenchymal lineages, classically derived from bone marrow (BM) but potentially from umbilical cord blood (UCB). Although they are becoming a good tool for regenerative medicine, they usually need to be expanded in fetal bovine serum (FBS)-supplemented media. Human platelet lysate (HPL) has recently been proposed as substitute for safety reasons, but it is not yet clear how this supplement influences the properties of expanded MSCs. METHODS: In the present study, we compared the effect of various media combining autologous HPL with or without FBS on phenotypic, proliferative and functional (differentiation, cytokine secretion profile) characteristics of human BM-derived MSCs. RESULTS: Despite less expression of adipogenic and osteogenic markers, MSCs cultured in HPL-supplemented media fully differentiated along osteoblastic, adipogenic, chondrogenic and vascular smooth muscle lineages. The analyses of particular specific proteins expressed during osteogenic differentiation (calcium-sensing receptor (CaSR) and parathormone receptor (PTHR)) showed their decrease at D0 before any induction for MSC cultured with HPL mostly at high percentage (10%HPL). The cytokine dosage showed a clear increase of proliferation capacity and interleukin (IL)-6 and IL-8 secretion. CONCLUSIONS: This study shows that MSCs can be expanded in media supplemented with HPL that can totally replace FBS. HPL-supplemented media not only preserves their phenotype as well as their differentiation capacity, but also shortens culture time by increasing their growth rate.

Immunophenotyping of hematopoietic progenitor cells: Comparison between cord blood and adult mobilized blood grafts.

AIM: To study the immunophenotype of hematopoietic progenitor cells from cord blood (CB) grafts (n = 39) in comparison with adult apheresis grafts (AG, n = 229) and pre-apheresis peripheral blood (PAPB) samples (n = 908) using flow cytometry analysis. METHODS: First, we performed a qualitative analysis of CD34+ cell sub-populations in both CB and PAPB grafts using the standardized ISHAGE protocol and a wide panel of 20 monoclonal antibodies. Next, we studied some parameters, such as the age of mothers and the weight of newborns, which can influence the quality and the quantity of CD34+ cells from CB. RESULTS: We found that the percentage of apoptotic cells was high in CB in comparison to PAPB (PAPB: 4.6% ± 2.6% vs CB: 53.4% ± 5.2%, P < 0.001). In CB, the weight of newborn and the age of the mother have the influence on CD34+ cells. The follow-up of Ag CD133 in the ISHAGE double platform protocol in association with CD45, CD34 and the 7'AAD shows an equal rate between the two cell populations CD133+CD45+CD34+ high and CD34+CD45+ high with a higher percentage. So, is the inclusion of Ac CD133 necessary in the present panel included in the ISHAGE method? Last part, we showed a significant presence of interferon γ in CB in comparison to PAPB, the annexin showing the high number of apoptotic cells in CB. CONCLUSION: This study demonstrates that many different obstetric factors must be taken into account when processing and cryo-banking umbilical CB units for transplantation.

Stromal cell-derived factor 1 polymorphism in patients infected with HIV and implications for AIDS progression in Tunisia.

BACKGROUND: An interesting finding in the epidemiology of human immunodeficiency virus (HIV) infection is that certain mutations in genes coding for chemokines, and their receptors and ligands, may confer resistance or susceptibility to HIV-1 infection and acquired immunodeficiency syndrome (AIDS) progression. The mutation most frequently studied is stromal cell-derived factor (SDF)1-3'A, a single nucleotide polymorphism in the 3' untranslated region at the 801 position of the SDF1 gene, which seems to be associated with susceptibility or resistance to diseases, including AIDS. We examined the frequency of the above polymorphisms in the Tunisian population, and evaluated their contribution to a protective genetic background against HIV infection and progression. METHODS AND MATERIALS: One hundred forty blood samples from HIV-infected patients from the Cellular Immunology Research Laboratory at the National Blood Transfusion Center were compared with those of 164 random blood donors from the same center. Genotyping was initially performed by polymerase chain reaction (PCR) analysis. SDF1 PCR product genomic regions were further subjected to restriction fragment length polymorphism analysis for genotype determination. Screening for the SDF1 polymorphism in the HIV-infected population yielded 56 heterozygous (40%), 52 mutation homozygous (37.1%), and 32 wild-type homozygous (22.8%) subjects. In contrast, in our healthy population, we found 70/164 heterozygous (42.6%), nine mutation homozygous (5.4%), and 85 wild-type homozygous (51.8%) subjects. The allele frequencies in the HIV-infected and healthy populations were f(SD1 3'A) = 57.1%, f(SDF1) = 42.8%, f(SDF1 3'A) = 26.8%, and f(SDF1) = 73.1%, respectively. The allelic and genotypic frequencies of the SDF1 3'A in our population show significantly higher distribution profiles compared with those observed in other Caucasian, European, and African American populations. Our results were examined by χ(2) test and appear to confirm an association between polymorphism and AIDS progression. A higher odds ratio (>1) was found for the SDF1-3'A allele than for the wild-type allele (<1). CONCLUSION: This result seems to confirm that the SDF1-3'A allele is associated with acceleration and progression from HIV infection to AIDS in the Tunisian population.

[Effect of triple antiretroviral therapy on Tunisian AIDS profile: study of 139 cases]. / Impact de la trithérapie anti-retrovirale sur le profil evolutif du sideen Tunisien: etude de 139 cas.

We report a retrospective study to estimate highly active antiretroviral therapy (HAART) effect in 139 HIV infected patients. Four criteria are studied: prevalence of opportunistic infections, CD4 cell count evolution, viral load progression and mortality. Gastrointestinal side effects are the most common clinical adverse reaction (61.1 percent), and hematological side effects are the most common biological adverse reaction (61.2 percent). During the 22.8 months (3 months to 6 years) follow-up average period, CD4 cell counts remained above 500 per cubic millimeter in only 25.8 percent of cases, while 63.5 percent of patients had a viral load below 400 copies per milliliter. During the study on patients receiving HAART, opportunistic infections appeared in 17.3 percent of cases (24 cases) and mortality in 6.4 percent of cases.

[Prevalence of anti-GAD autoantibodies in Tunisian children with type 1 diabetes]. / Prévalence des auto anticorps anti-GAD chez des enfants Tunisiens diabétiques de type 1.

The prevalence of GAD antibodies and its correlation with some autoimmune markers: ICA (islet cells antibodies), antinuclear antibodies, thyroid antithymune and antimicrosomes antibodies, were studied in 84 Tunisian type 1 diabetic children. The prevalence of GAD antibodies was 51.2% and decreased as a function of increasing duration of the disease. Their frequency was 84.6% in children with newly diagnosed diabetes (within 6 months of diagnosis) and only 29.41% in those with a longer duration of the diabetes (more than 5 years). ICA were present less frequently (21.4% of the children). 10.7% of the studied samples were positive with the two antibodies (GAD ab ICA), and 40% were positive only with GAD antibodies. We conclude that the GAD antibodies seems to be more associated to the development of type 1 diabetes than ICA. They are detected more frequently in patients with long standing disease, that's make their determination very interesting as diagnostic and predictive marker.