Mesenchymal stem/stromal cells (MSCs) are increasingly explored for the treatment of degenerative and inflammatory diseases in human and veterinary medicine. One of the key characteristics of MSCs is that they modulate inflammation mainly through the secretion of soluble mediators. However, despite widespread clinical use, knowledge regarding the effector mechanisms of equine MSCs, and consequently their effectiveness in the treatment of diseases, is still unknown. The objectives of this study were to determine the mechanisms underlying inhibition of lymphocyte proliferation by equine bone marrow-derived MSCs, and to evaluate the effect of pre-conditioning of equine MSCs with different pro-inflammatory cytokines on inhibition of lymphocyte proliferation. We determined that inhibition of lymphocyte proliferation by equine MSCs depends on activity of prostaglandin-endoperoxide synthase 2 and indoleamine 2,3-dioxygenase. Additionally, pre-conditioning of MSCs with TNF-α, IFN-γ or their combination significantly increased the expression of prostaglandin-endoperoxide synthase 2, indoleamine 2,3-dioxygenase, iNOS and IL-6. This upregulation correlated with an increased inhibitory effect of MSCs on lymphocyte proliferation. In conclusion, pre-conditioning of bone marrow-derived MSC increases their inhibitory effect on lymphocyte proliferation in horses.

The absence of optimal treatments for Diabetic Nephropathy (DN) highlights the importance of the search for novel therapeutic targets. The vascular endothelial growth factor receptor 2 (VEGFR2) pathway is activated in experimental and human DN, but the effects of its blockade in experimental models of DN is still controversial. Here, we test the effects of a therapeutic anti-VEGFR2 treatment, using a VEGFR2 kinase inhibitor, on the progression of renal damage in the BTBR ob/ob (leptin deficiency mutation) mice. This experimental diabetic model develops histological characteristics mimicking the key features of advanced human DN. A VEGFR2 pathway-activation blockade using the VEGFR2 kinase inhibitor SU5416, starting after kidney disease development, improves renal function, glomerular damage (mesangial matrix expansion and basement membrane thickening), tubulointerstitial inflammation and tubular atrophy, compared to untreated diabetic mice. The downstream mechanisms involved in these beneficial effects of VEGFR2 blockade include gene expression restoration of podocyte markers and downregulation of renal injury biomarkers and pro-inflammatory mediators. Several ligands can activate VEGFR2, including the canonical ligands VEGFs and GREMLIN. Activation of a GREMLIN/VEGFR2 pathway, but not other ligands, is correlated with renal damage progression in BTBR ob/ob diabetic mice. RNA sequencing analysis of GREMLIN-regulated genes confirm the modulation of proinflammatory genes and related-molecular pathways. Overall, these data show that a GREMLIN/VEGFR2 pathway activation is involved in diabetic kidney disease and could potentially be a novel therapeutic target in this clinical condition.

Multipotent stromal cells (MSCs) are envisioned as a powerful therapeutic tool. As they home into tumors, secrete trophic and vasculogenic factors, and suppress immune response their role in carcinogenesis is a matter of controversy. Worldwide oral squamous cell carcinoma (OSCC) is the fifth most common epithelial cancer. Our aim was to determine whether MSC administration at precancerous stage modifies the natural progression of OSCC. OSCC was induced in Syrian hamsters by topical application of DMBA in the buccal pouch. At papilloma stage, the vehicle or 3×10 allogenic bone marrow-derived MSCs were locally administered. Four weeks later, the lesions were studied according to: volume, stratification (histology), proliferation (Ki-67), apoptosis (Caspase 3 cleaved), vasculature (ASMA), inflammation (Leukocyte infiltrate), differentiation (CK1 and CK4) and gene expression profile (mRNA). Tumors found in individuals that received MSCs were smaller than those presented in the vehicle group (87±80 versus 54±62mm, p<0.05). The rate of proliferation was two times lower and the apoptosis was 2.5 times higher in lesions treated with MSCs than in untreated ones. While the laters presented dedifferentiated cells, the former maintained differentiated cells (cytokeratin and gene expression profile similar to normal tissue). Thus, MSC administration at papilloma stage precludes tumor growth and epithelial dedifferentiation of OSCC.

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BackgroundMesenchymal stem cells (MSCs) are a heterogeneous subset of stromal cells currently tested for multiple therapeutic purposes. Their potential to home into tumors, to secrete trophic/vasculogenic factors, and to suppress immune response raises questions regarding their biosafety. Our aim was to evaluate whether systemically administered allogeneic MSCs modify the natural progression of precancerous lesions and whether their putative effect depends on cancer stage and/or cell dose.MethodsOral squamous cell carcinoma (OSCC) was induced in Syrian golden hamsters by topical application of 7,12-dimethylbenz[a]anthracene in one buccal pouch. At hyperplasia, dysplasia, or papilloma stage, animals received intracardially the vehicle or 0.7 × 106, 7 × 106, or 21 × 106 allogeneic bone marrow-derived MSCs/kg. OSCC progression was assessed according to the presence of erythroplakia and leukoplakia, extent of inflammation and vascularization, and appearance, volume, and staging of tumors. Also, the homing of donor cells was studied.ResultsPrecancerous lesions progressed from hyperplasia to dysplasia in 2 weeks, from dysplasia to papilloma in 3 weeks, and from papilloma to carcinoma in 4 weeks. This time course was unmodified by the systemic administration of MSCs at hyperplasia or dysplasia stages. When MSCs were administered at papilloma stage, lesions did not progress to carcinoma stage. Tumors developed in hamsters receiving 0.7 × 106 or 7 × 106 MSCs/kg at hyperplasia stage were significantly smaller than those found in control animals (25 ± 4 or 23 ± 4 mm3 versus 72 ± 19 mm3, p < 0.05). Similar results were obtained when 0.7 × 106, 7 × 106, or 21 × 106 MSCs/kg were administered at papilloma stage (44 ± 15, 28 ± 7, or 28 ± 5 mm3 versus 104 ± 26 mm3, p < 0.05). For dysplasia stage, only the lower concentration of MSCs reached statistical significance (21 ± 9 mm3 versus 94 ± 39 mm3, p < 0.05). Animals receiving 21 × 106 MSCs/kg at hyperplasia stage developed tumors larger than those found in animals that received the vehicle (147 ± 47 mm3 versus 72 ± 19 mm3, p < 0.05). Donor cells were rarely found in precancerous lesions.ConclusionsSystemically administered allogeneic MSCs do not aggravate the progression of precancerous lesions. Moreover, they preclude cancer progression and tumor growth.Electronic supplementary materialThe online version of this article (10.1186/s13287-018-0878-1) contains supplementary material, which is available to authorized users.

Diabetic nephropathy (DN) is a multifactorial disease characterized by hyperglycemia and close interaction of hemodynamic, metabolic and inflammatory factors. Nuclear factor-κB (NF-κB) is a principal matchmaker linking hyperglycemia and inflammation. The present work investigates the cell-permeable peptide containing the inhibitor of kappa B kinase γ (IKKγ)/NF-κB essential modulator (NEMO)-binding domain (NBD) as therapeutic option to modulate inflammation in a preclinical model of type 2 diabetes (T2D) with DN. Black and tan, brachyuric obese/obese mice were randomized into 4 interventions groups: Active NBD peptide (10 and 6 µg/g body weight); Inactive mutant peptide (10 µg/g); and vehicle control. In vivo/ex vivo fluorescence imaging revealed efficient delivery of NBD peptide, systemic biodistribution and selective renal metabolization. In vivo administration of active NBD peptide improved albuminuria (>40% reduction on average) and kidney damage, decreased podocyte loss and basement membrane thickness, and modulated the expression of proinflammatory and oxidative stress markers. In vitro, NBD blocked IKK-mediated NF-κB induction and target gene expression in mesangial cells exposed to diabetic-like milieu. These results constitute the first nephroprotective effect of NBD peptide in a T2D mouse model that recapitulates the kidney lesions observed in DN patients. Targeting IKK-dependent NF-κB activation could be a therapeutic strategy to combat kidney inflammation in DN.

Cyclosporine (CsA) and tacrolimus (TAC) are immunosuppressant drugs characterized by a narrow therapeutic range and high pharmacokinetic variability. The effect of polymorphisms in genes related to the metabolism and transport of these drugs, namely CYP3A4, CYP3A5, MDR1 and POR genes, has been evaluated in diverse populations. However, the impact of these polymorphisms on drug disposition is not well established in Latin American populations. Using TaqMan® probes, we determined the allelic frequency of seven variants in CYP3A4, CYP3A5, MDR1 and POR in 139 Chilean renal transplant recipients, of which 89 were treated with CsA and 50 with TAC. We tested associations between variants and trough and/or 2-hour concentrations, normalized by dose (C0/D and C2/D) at specific time points post-transplant. We found that CYP3A5*3/*3 carriers required lower doses of TAC. In TAC treated patients, most CYP3A5*3/*3 carriers presented higher C0/D and a high proportion of patients with C0 levels outside the therapeutic range relative to other genotypes. These results reinforce the value of considering CYP3A5 genotypes alongside therapeutic drug monitoring for TAC treated Chilean kidney recipients.

Mesenchymal stem/stromal cells (MSCs) therapy has been a cornerstone of regenerative medicine in humans and animals since their identification in 1968. MSCs can interact and modulate the activity of practically all cellular components of the immune response, either through cell-cell contact or paracrine secretion of soluble mediators, which makes them an attractive alternative to conventional therapies for the treatment of chronic inflammatory and immune-mediated diseases. Many of the mechanisms described as necessary for MSCs to modulate the immune/inflammatory response appear to be dependent on the animal species and source. Although there is evidence demonstrating an in vitro immunomodulatory effect of MSCs, there are disparate results between the beneficial effect of MSCs in preclinical models and their actual use in clinical diseases. This discordance might be due to cells' limited survival or impaired function in the inflammatory environment after transplantation. This limited efficacy may be due to several factors, including the small amount of MSCs inoculated, MSC administration late in the course of the disease, low MSC survival rates in vivo, cryopreservation and thawing effects, and impaired MSC potency/biological activity. Multiple physical and chemical pre-conditioning strategies can enhance the survival rate and potency of MSCs; this paper focuses on hypoxic conditions, with inflammatory cytokines, or with different pattern recognition receptor ligands. These different pre-conditioning strategies can modify MSCs metabolism, gene expression, proliferation, and survivability after transplantation.