Comparison of human isogeneic Wharton's jelly MSCs and iPSC-derived MSCs reveals differentiation-dependent metabolic responses to IFNG stimulation.

Variability among donors, non-standardized methods for isolation, and characterization contribute to mesenchymal stem/stromal cell (MSC) heterogeneity. Induced pluripotent stem cell (iPSCs)-derived MSCs would circumvent many of current issues and enable large-scale production of standardized cellular therapy. To explore differences between native MSCs (nMSCs) and iPSC-derived MSCs (iMSCs), we developed isogeneic lines from Wharton's jelly (WJ) from the umbilical cords of two donors (#12 and #13) under xeno-free conditions. Next, we reprogrammed them into iPSCs (iPSC12 and iPSC13) and subsequently differentiated them back into iMSCs (iMSC12 and iMSC13) using two different protocols, which we named ARG and TEX. We assessed their differentiation capability, transcriptome, immunomodulatory potential, and interferon-γ (IFNG)-induced changes in metabolome. Our data demonstrated that although both differentiation protocols yield iMSCs similar to their parental nMSCs, there are substantial differences. The ARG protocol resulted in iMSCs with a strong immunomodulatory potential and lower plasticity and proliferation rate, whereas the TEX protocol raised iMSCs with a higher proliferation rate, better differentiation potential, though weak immunomodulatory response. Our data suggest that, following a careful selection and screening of donors, nMSCs from umbilical's cord WJ can be easily reprogrammed into iPSCs, providing an unlimited source of material for differentiation into iMSCs. However, the differentiation protocol should be chosen depending on their clinical use.

Effects of maternal obesity on Wharton's Jelly mesenchymal stromal cells.

We investigated whether maternal metabolic environment affects mesenchymal stromal/stem cells (MSCs) from umbilical cord's Wharton's Jelly (WJ) on a molecular level, and potentially render them unsuitable for clinical use in multiple recipients. In this pilot study on umbilical cords post partum from healthy non-obese (BMI = 19-25; n = 7) and obese (BMI ≥ 30; n = 7) donors undergoing elective Cesarean section, we found that WJ MSC from obese donors showed slower population doubling and a stronger immunosuppressive activity. Genome-wide DNA methylation of triple positive (CD73+CD90+CD105+) WJ MSCs found 67 genes with at least one CpG site where the methylation difference was ≥0.2 in four or more obese donors. Only one gene, PNPLA7, demonstrated significant difference on methylome, transcriptome and protein level. Although the number of analysed donors is limited, our data suggest that the altered metabolic environment related to excessive body weight might bear consequences on the WJ MSCs.

Genome-wide effects of MELK-inhibitor in triple-negative breast cancer cells indicate context-dependent response with p53 as a key determinant.

Triple-negative breast cancer (TNBC) is an aggressive, highly recurrent breast cancer subtype, affecting approximately one-fifth of all breast cancer patients. Subpopulations of treatment-resistant cancer stem cells within the tumors are considered to contribute to disease recurrence. A potential druggable target for such cells is the maternal embryonic leucine-zipper kinase (MELK). MELK expression is upregulated in mammary stem cells and in undifferentiated cancers, where it correlates with poor prognosis and potentially mediates treatment resistance. Several MELK inhibitors have been developed, of which one, OTSSP167, is currently in clinical trials. In order to better understand how MELK and its inhibition influence TNBC, we verified its anti-proliferative and apoptotic effects in claudin-low TNBC cell lines MDA-MB-231 and SUM-159 using MTS assays and/or trypan blue viability assays together with analysis of PARP cleavage. Then, using microarrays, we explored which genes were affected by OTSSP167. We demonstrate that different sets of genes are regulated in MDA-MB-231 and SUM-159, but in both cell lines genes involved in cell cycle, mitosis and protein metabolism and folding were regulated. We identified p53 (TP53) as a potential upstream regulator of the regulated genes. Using western blot we found that OTSSP167 downregulates mutant p53 in all tested TNBC cell lines (MDA-MB-231, SUM-159, and BT-549), but upregulates wild-type p53 in the luminal A subtype MCF-7 cell line. We propose that OTSSP167 might have context-dependent or off-target effects, but that one consistent mechanism of action could involve the destabilization of mutant p53.

Ligand Independent and Subtype-Selective Actions of Thyroid Hormone Receptors in Human Adipose Derived Stem Cells.

Thyroid hormone (TH) receptors (TRs α and ß) are homologous ligand-dependent transcription factors (TFs). While the TRs display distinct actions in development, metabolic regulation and other processes, comparisons of TRα and TRß dependent gene regulation mostly reveal similar mechanisms of action and few TR subtype specific genes. Here, we show that TRα predominates in multipotent human adipose derived stem cells (hADSC) whereas TRß is expressed at lower levels and is upregulated during hADSC differentiation. The TRs display several unusual properties in parental hADSC. First, TRs display predominantly cytoplasmic intracellular distribution and major TRα variants TRα1 and TRα2 colocalize with mitochondria. Second, knockdown experiments reveal that endogenous TRs influence hADSC cell morphology and expression of hundreds of genes in the absence of hormone, but do not respond to exogenous TH. Third, TRα and TRß affect hADSC in completely distinct ways; TRα regulates cell cycle associated processes while TRß may repress aspects of differentiation. TRα splice variant specific knockdown reveals that TRα1 and TRα2 both contribute to TRα-dependent gene expression in a gene specific manner. We propose that TRs work in a non-canonical and hormone independent manner in hADSC and that prominent subtype-specific activities emerge in the context of these unusual actions.

Hemolytic anemia in patients receiving daily dapsone for the treatment of leprosy.

INTRODUCTION: Multidrug therapy for leprosy is currently done with dapsone, clofazimine and rifampicin. Dapsone is known to cause hemolytic anemia (HA) and this adverse event during MDT seems to be more frequent than reported. The aim of this report is to discuss and grade HA due to dapsone during MDT treatment for leprosy. METHODS: This is a retrospective study of 194 leprosy patients from a Leprosy Control Programme Unit in Vit6ria-ES, Brazil. RESULTS: HA was observed in 48 (24.7%) patients and occurred within the first 3 months in 51% of these. Mean hematocrit levels fell from 38.5 to 31.5 and hemoglobin from 12.8 to 10.3. CONCLUSION: Dapsone used in the MDT regime for leprosy decreases the hematocrit and hemoglobin levels due to a low grade hemolysis, which can result in significant anemia.

Adverse effects from multi-drug therapy in leprosy: a Brazilian study.

INTRODUCTION: The WHO MDT for leprosy treatment was officially introduced in Brazil in 1991 and comprises three drugs: dapsone, rifampicin and clofazimine. There are few good studies on the frequency of side-effects attributable to MDT in Brazil. METHODS: A retrospective and descriptive study carried out in a LCP in Vitória, State of Espirito Santo, Brazil. A specific and detailed protocol about side-effects was prepared and filled in from the patient records. RESULTS: One hundred ninety four patients' records were analysed looking for side-effects attributable to MDT. Side-effects were attributed to at least one MDT component in 88 (45%) patients and 85 had side-effects due to dapsone, 24 due to rifampicin and 18 due to clofazimine. 185 episodes were identified. The suspected drug was stopped in 47 out of 88 episodes (24% patients); 46 had dapsone stopped, 5 had rifampicin stopped and no-one had clofazimine stopped. CONCLUSION: Side-effects attributed to MDT is more frequent than previously described, resulting in interruption of treatment in many patients.