Development and Validation of a New LC-MS/MS Bioanalytical Method for the Simultaneous Determination of Levodopa, Levodopa Methyl Ester, and Carbidopa in Human Plasma Samples.

Levodopa (L-DOPA) treatment, combined with the administration of dopa-decarboxylase inhibitors (DDCIs), is still the most effective symptomatic treatment of Parkinson's disease (PD). Although its efficacy in the early stage of the disease has been confirmed, its complex pharmacokinetics (PK) increases the variability of the intra-individual motor response, thus amplifying the risk of motor/non-motor fluctuations and dyskinesia. Moreover, it has been demonstrated that L-DOPA PK is strongly influenced by several clinical, therapeutic, and lifestyle variables (e.g., dietary proteins). L-DOPA therapeutic monitoring is therefore crucial to provide personalized therapy, hence improving drug efficacy and safety. To this aim, we have developed and validated an ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method to quantify L-DOPA, levodopa methyl ester (LDME), and the DDCI carbidopa in human plasma. The compounds were extracted by protein precipitation and samples were analyzed with a triple quadrupole mass spectrometer. The method showed good selectivity and specificity for all compounds. No carryover was observed, and dilution integrity was demonstrated. No matrix effect could be retrieved; intra-day and inter-day precision and accuracy values met the acceptance criteria. Reinjection reproducibility was assessed. The described method was successfully applied to a 45-year-old male patient to compare the pharmacokinetic behavior of an L-DOPA-based medical treatment involving commercially available Mucuna pruriens extracts and an LDME/carbidopa (100/25 mg) formulation.

A comprehensive report of long-term stability data for a range ATMPs: A need to develop guidelines for safe and harmonized stability studies.

BACKGROUND AIMS: Advanced therapy medicinal products (ATMPs) are novel drugs based on genes, cells or tissues developed to treat many different diseases. Stability studies of each new ATMP need to be performed to define its shelf life and guarantee efficacy and safety upon infusion, and these are presently based on guidelines originally drafted for standard pharmaceutical drugs, which have properties and are stored in conditions quite different from cell products. The aim of this report is to provide evidence-based information for stability studies on ATMPs that will facilitate the interlaboratory harmonization of practices in this area. METHODS: We have collected and analyzed the results of stability studies on 19 different cell-based experimental ATMPs, produced by five authorized cell factories forming the Lombardy "Plagencell network" for use in 36 approved phase I/II clinical trials; most were cryopreserved and stored in liquid nitrogen vapors for 1 to 13 years. RESULTS: The cell attributes collected in stability studies included cell viability, immunophenotype and potency assays, in particular immunosuppression, cytotoxicity, cytokine release and proliferation/differentiation capacity. Microbiological attributes including sterility, endotoxin levels and mycoplasma contamination were also analyzed. All drug products (DPs), cryopreserved in various excipients containing 10% DMSO and in different primary containers, were very stable long term at <-150°C and did not show any tendency for diminished viability or efficacy for up to 13.5 years. CONCLUSIONS: Our data indicate that new guidelines for stability studies, specific for ATMPs and based on risk analyses, should be drafted to harmonize practices, significantly reduce the costs of stability studies without diminishing safety. Some specific suggestions are presented in the discussion.

Third-party bone marrow-derived mesenchymal stromal cell infusion before liver transplantation: A randomized controlled trial.

Mesenchymal stromal cells (MSC) have emerged as a promising therapy to minimize the immunosuppressive regimen or induce tolerance in solid organ transplantation. In this randomized open-label phase Ib/IIa clinical trial, 20 liver transplant patients were randomly allocated (1:1) to receive a single pretransplant intravenous infusion of third-party bone marrow-derived MSC or standard of care alone. The primary endpoint was the safety profile of MSC administration during the 1-year follow-up. In all, 19 patients completed the study, and none of those who received MSC experienced infusion-related complications. The incidence of serious and non-serious adverse events was similar in the two groups. Circulating Treg/memory Treg and tolerant NK subset of CD56bright NK cells increased slightly over baseline, albeit not to a statistically significant extent, in MSC-treated patients but not in the control group. Graft function and survival, as well as histologic parameters and intragraft expression of tolerance-associated transcripts in 1-year protocol biopsies were similar in the two groups. In conclusion, pretransplant MSC infusion in liver transplant recipients was safe and induced mild positive changes in immunoregulatory T and NK cells in the peripheral blood. This study opens the way for a trial on possible tolerogenic efficacy of MSC in liver transplantation. ClinicalTrials.gov identifier: NCT02260375.

Utility of routine evaluation of sterility of cellular therapy products with or without extensive manipulation: Best practices and clinical significance.

BACKGROUND: We analyzed the results of routine sterility testing performed in our center over the last 10 years, in the context both hematopoietic stem cell transplantation (HSCT) and Advanced Therapeutic Medicinal Products (ATMPs). METHODS: For sterility tests 14-day cultures were performed in culture media detecting aerobic and anaerobic microorganisms. RESULTS: In this study, 22/1643 (1.3%) of apheretic products for autologous or allogeneic HSCT were contaminated, whereas 14/73 bone marrow (BM) harvests (17.8%) were positive. In 22 cases, the contaminated HSCs were infused to patients, but there was no evidence of any adverse impact of contamination on the hematologic engraftment or on infections. Indeed none of the five positive hemocultures detected in patients following infusion could be linked to the contaminated stem cell product. Our Cell Factory also generated 286 ATMPs in good manufacturing practice (GMP) conditions since 2007 and all final products were sterile. In three cases of mesenchymal stromal cell expansions, the starting BM harvests were contaminated, but the cell products at the end of expansion were sterile, presumably thanks to the presence of an antibiotic in the culture medium. DISCUSSION: The decreased rate of contamination of cell harvests observed with time suggests that routine sterility testing and communication of the results to the collecting centers may improve clinical practices. Furthermore, we recommend the use of antibiotics in the medium for ATMP expansion, to decrease the likelihood of expanding microorganisms within clean rooms. Finally we discuss the costs of sterility testing of ATMPs by GMP-approved external laboratories.

Molecular events underlying interleukin-6 independence in a subclone of the CMA-03 multiple myeloma cell line.

We explored the molecular mechanisms involved in the establishement of CMA-03/06, an IL-6-independent variant of the multiple myeloma cell line CMA-03 previously generated in our Institution. CMA-03/06 cells grow in the absence of IL-6 with a doubling time comparable with that of CMA-03 cells; neither the addition of IL6 (IL-6) to the culture medium nor co-culture with multipotent mesenchymal stromal cells increases the proliferation rate, although they maintain the responsiveness to IL-6 stimulation as demonstrated by STAT1, STAT3, and STAT5 induction. IL-6 independence of CMA-03/06 cells is not apparently due to the development of an autocrine IL-6 loop, nor to the observed moderate constitutive activation of STAT5 and STAT3, since STAT3 silencing does not affect cell viability or proliferation. When compared to the parental cell line, CMA-03/06 cells showed an activated pattern of the NF-κB pathway. This finding is supported by gene expression profiling (GEP) analysis identifying an appreciable fraction of modulated genes (28/308) in the CMA-03/06 subclone reported to be involved in this pathway. Furthermore, although more resistant to apoptotic stimuli compared to the parental cell line, CMA-03/06 cells display a higher sensibility to NF-κB inhibition induced by bortezomib. Finally, GEP analysis suggests an involvement of a number of cytokines, which might contribute to IL-6 independence of CMA-03/06 by stimulating growth and antiapoptotic processes. In conclusion, the parental cell-line CMA-03 and its variant CMA-03/06 represent a suitable model to further investigate molecular mechanisms involved in the IL-6-independent growth of myeloma cells.

Treatment of graft versus host disease with mesenchymal stromal cells: a phase I study on 40 adult and pediatric patients.

This phase I multicenter study was aimed at assessing the feasibility and safety of intravenous administration of third party bone marrow-derived mesenchymal stromal cells (MSC) expanded in platelet lysate in 40 patients (15 children and 25 adults), experiencing steroid-resistant grade II to IV graft-versus-host disease (GVHD). Patients received a median of 3 MSC infusions after having failed conventional immunosuppressive therapy. A median cell dose of 1.5 × 10(6)/kg per infusion was administered. No acute toxicity was reported. Overall, 86 adverse events and serious adverse events were reported in the study, most of which (72.1%) were of infectious nature. Overall response rate, measured at 28 days after the last MSC injection, was 67.5%, with 27.5% complete response. The latter was significantly more frequent in patients exhibiting grade II GVHD as compared with higher grades (61.5% versus 11.1%, P = .002) and was borderline significant in children as compared with adults (46.7 versus 16.0%, P = .065). Overall survival at 1 and 2 years from the first MSC administration was 50.0% and 38.6%, with a median survival time of 1.1 years. In conclusion, MSC can be safely administered on top of conventional immunosuppression for steroid resistant GVHD treatment. Eudract Number 2008-007869-23, NCT01764100.

Mesenchymal stromal cells and kidney transplantation: pretransplant infusion protects from graft dysfunction while fostering immunoregulation.

Bone marrow-derived mesenchymal stromal cells (MSC) have emerged as useful cell population for immunomodulation therapy in transplantation. Moving this concept towards clinical application, however, should be critically assessed by a tailor-made step-wise approach. Here, we report results of the second step of the multistep MSC-based clinical protocol in kidney transplantation. We examined in two living-related kidney transplant recipients whether: (i) pre-transplant (DAY-1) infusion of autologous MSC protected from the development of acute graft dysfunction previously reported in patients given MSC post-transplant, (ii) avoiding basiliximab in the induction regimen improved the MSC-induced Treg expansion previously reported with therapy including this anti-CD25-antibody. In patient 3, MSC treatment was uneventful and graft function remained normal during 1 year follow-up. In patient 4, acute cellular rejection occurred 2 weeks post-transplant. Both patients had excellent graft function at the last observation. Circulating memory CD8(+) T cells and donor-specific CD8(+) T-cell cytolytic response were reduced in MSC-treated patients, not in transplant controls not given MSC. CD4(+) FoxP3(+) Treg expansion was comparable in MSC-treated patients with or without basiliximab induction. Thus, pre-transplant MSC no longer negatively affect kidney graft at least to the point of impairing graft function, and maintained MSC-immunomodulatory properties. Induction therapy without basiliximab does not offer any advantage on CD4(+) FoxP3(+) Treg expansion (ClinicalTrials.gov number: NCT 00752479).

Potency assays and biomarkers for cell-based advanced therapy medicinal products.

Advanced Therapy Medicinal Products (ATMPs) based on somatic cells expanded in vitro, with or without genetic modification, is a rapidly growing area of drug development, even more so following the marketing approval of several such products. ATMPs are produced according to Good Manufacturing Practice (GMP) in authorized laboratories. Potency assays are a fundamental aspect of the quality control of the end cell products and ideally could become useful biomarkers of efficacy in vivo. Here we summarize the state of the art with regard to potency assays used for the assessment of the quality of the major ATMPs used clinic settings. We also review the data available on biomarkers that may substitute more complex functional potency tests and predict the efficacy in vivo of these cell-based drugs.

A phase I study of autologous mesenchymal stromal cells for severe steroid-dependent nephrotic syndrome.

BACKGROUNDSevere forms of idiopathic nephrotic syndrome (INS) require prolonged immunosuppressive therapies and repeated courses of high-dose glucocorticoids. Mesenchymal stromal cells (MSCs) have promising immunomodulatory properties that may be employed therapeutically to reduce patient exposure to medications and their side effects.METHODSWe performed a phase I open-label trial assessing safety and feasibility of autologous bone marrow-derived MSCs (BM-MSCs) in children and young adults with severe forms of steroid-dependent nephrotic syndrome. Following autologous BM-MSC preparation and infusion, oral immunosuppression was tapered. Safety, efficacy, and immunomodulatory effects in vivo were monitored for 12 months.RESULTSSixteen patients (10 children, 6 adults) were treated. Adverse events were limited and not related to BM-MSC infusions. All patients relapsed during follow-up, but in the 10 treated children, time to first relapse was delayed (P = 0.02) and number of relapses was reduced (P = 0.002) after BM-MSC infusion, compared with the previous 12 months. Cumulative prednisone dose was also reduced at 12 months compared with baseline (P < 0.05). No treatment benefit was observed in adults.In children, despite tapering of immunosuppression, clinical benefit was mirrored by a significant reduction in total CD19+, mature, and memory B cells and an increase in regulatory T cells in vivo up to 3-6 months following BM-MSC infusionCONCLUSIONTreatment with autologous BM-MSCs is feasible and safely reduces relapses and immunosuppression at 12 months in children with severe steroid-dependent INS. Immunomodulatory studies suggest that repeating MSC infusions at 3-6 months may sustain benefit.TRIAL REGISTRATIONEudraCT 2016-004804-77.FUNDINGAIFA Ricerca Indipendente 2016-02364623.

Life-sparing effect of human cord blood-mesenchymal stem cells in experimental acute kidney injury.

In search for new sources of mesenchymal stem cells (MSCs) for renal repair in acute kidney injury (AKI), we investigated the potential of human cord blood (CB)-MSCs to cure mice with AKI. Infusion of CB-MSCs in immunodeficient mice with cisplatin-induced AKI ameliorated both renal function and tubular cell injury, and prolonged survival. Transplanted CB-MSCs localized in peritubular areas, limited capillary alterations and neutrophil infiltration. Apoptosis reduced and tubular cell proliferation increased by virtue of stem cell capacity to produce growth factors. The reno-protective effect of CB-MSCs was further confirmed by their ability to inhibit oxidative damage and to induce the prosurvival factor Akt in tubular cells. The evidence that CB-MSCs in vitro increased the production of growth factors and inhibited IL-1 beta and TNFalpha synthesis when cocultured with damaged proximal tubular cells indicates a regenerative and anti-inflammatory action of stem cell treatment. Altogether these results highlight the potential of human CB-MSCs as future cell therapy for testing in human AKI.

Minimally manipulated whole human umbilical cord is a rich source of clinical-grade human mesenchymal stromal cells expanded in human platelet lysate.

BACKGROUND AIMS: Mesenchymal stromal cells (MSC) have recently been identified as a therapeutic option in several clinical conditions. Whereas bone marrow (BM) is considered the main source of MSC (BM-MSC), the invasive technique required for collection and the decline in allogeneic donations call for alternative sources. Human umbilical cord (UC) represents an easily available source of MSC (UC-MSC). METHODS: Sections of full-term UC were transferred to cell culture flasks and cultured in 5% human platelet lysate (PL)-enriched medium. Neither enzymatic digestion nor blood vessel removal was performed. After 2 weeks, the adherent cells were harvested (P1), replated at low density and expanded for two consecutive rounds (P2 and P3). RESULTS: We isolated and expanded MSC from 9/9 UC. UC-MSC expanded with a mean fold increase (FI) of 42 735 ± 16 195 from P1 to P3 in a mean of 29 ± 2 days. By processing the entire cord unit, we theoretically could have reached a median of 9.5 × 10(10) cells (ranging from 1.0 × 10(10) to 29.0 × 10(10)). UC-MSC expressed standard surface markers; they contained more colony-forming unit (CFU)-fibroblast (F) and seemed less committed towards osteogenic, chondrogenic and adipogenic lineages than BM-MSC. They showed immunosuppressive properties both in vitro and in an in vivo chronic Graft versus Host disease (cGvHD) mouse model. Both array-Comparative Genomic Hybridization (CGH) analysis and karyotyping revealed no chromosome alterations at the end of the expansion. Animal studies revealed no tumorigenicity in vivo. CONCLUSIONS: UC constitute a convenient and very rich source of MSC for the production of third-party 'clinical doses' of cells under good manufacturing practice (GMP) conditions.

Feasibility and safety of adoptive immunotherapy with CIK cells after cord blood transplantation.

Five patients with aggressive acute leukemias who had relapsed after cord blood transplantation were treated with cord blood derived cytokine-induced killer (CIK) cells. These were obtained by ex vivo expansion, using as starting material the washouts of the cord blood units, left over at the end of the transplant. We did not observe any acute or delayed adverse event, and observed 1 partial response in 1 patient concomitantly with the development of acute grade III graft-versus-host disease (GVHD). These observations show the relatively low toxicity of cord blood-derived CIK cells and, more importantly, the feasibility of this immunotherapy program for patients who could not otherwise benefit from donor lymphocyte infusions.

Platelet-lysate-expanded mesenchymal stromal cells as a salvage therapy for severe resistant graft-versus-host disease in a pediatric population.

Despite advances in graft-versus-host-disease (GVHD) treatment, it is estimated that overall survival (OS) at 2 years for hematopoietic cell transplantation (HCT) recipients who experience steroid-resistant GVHD is 10%. Among recent therapeutic approaches for GVHD treatment, mesenchymal stromal cells (MSCs) hold a key position. We describe a multicenter experience of 11 pediatric patients diagnosed with acute or chronic GVHD (aGVHD, cGVHD) treated for compassionate use with GMP-grade unrelated HLA-disparate donors' bone marrow-derived MSCs, expanded in platelet-lysate (PL)-containing medium. Eleven patients (aged 4-15 years) received intravenous (i.v.) MSCs for aGVHD or cGVHD, which was resistant to multiple lines of immunosuppression. The median dose was 1.2 x 10(6)/kg (range: 0.7-3.7 x 10(6)/kg). No acute side effects were observed, and no late side effects were reported at a median follow-up of 8 months (range: 4-18 months). Overall response was obtained in 71.4% of patients, with complete response in 23.8% of cases. None of our patients presented GVHD progression upon MSC administration, but 4 patients presented GVHD recurrence 2 to 5 months after infusion. Two patients developed chronic limited GVHD. This study underlines the safety of PL-expanded MSC use in children. MSC efficacy seems to be greater in aGVHD than in cGVHD, even after failure of multiple lines of immunosuppression.

Protective Effects of Human Nonrenal and Renal Stromal Cells and Their Conditioned Media in a Rat Model of Chronic Kidney Disease.

Mesenchymal stromal cells (MSCs) are emerging as a novel therapeutic option for limiting chronic kidney disease progression. Conditioned medium (CM) containing bioactive compounds could convey similar benefits, avoiding the potential risks of cell therapy. This study compared the efficacy of nonrenal and renal cell-based therapy with the corresponding CM in rats with renal mass reduction (RMR). Infusions of human kidney stromal cells (kPSCs) and CM-kPSCs, but not umbilical cord (uc) MSCs or CM-ucMSCs, reduced proteinuria and preserved podocyte number and nephrin expression in RMR rats. Glomerular fibrosis, microvascular rarefaction, and apoptosis were reduced by all treatments, while the peritubular microvascular loss was reduced by kPSCs and CM-kPSCs treatment only. Importantly, kPSCs and CM-kPSCs reduced NG2-positive pericytes, and all therapies reduced α-smooth muscle actin expression, indicating reduced myofibroblast expansion. Treatment with kPSCs also significantly inhibited the accumulation of ED1-positive macrophages in the renal interstitium of RMR rats. These findings demonstrate that the CM of ucMSCs and kPSCs confers similar renoprotection as the cells. kPSCs and CM-kPSCs may be superior in attenuating chronic renal injury as a cell source.

The washouts of discarded bone marrow collection bags and filters are a very abundant source of hMSCs.

BACKGROUND AIMS: Human multipotent mesenchymal stromal cells (hMSC) are considered good candidates for a growing spectrum of cell therapies. We have validated a protocol that makes use of the washouts of discarded collection sets, left over at the end of the filtration of bone marrow (BM) explants performed for hematopoietic stem cell (HSC) transplantation. METHODS: The method consists of direct plating of cells without density-gradient isolation followed by two detachment steps and expansion in 5% human platelet lysate (hPL). RESULTS: In a median of 26 days, 14 bags for adult patients and nine bags for pediatric patients for a standard dose of 1x10(6) hMSC/kg body weight could be prepared from the expansion of a fraction of the cells recovered from seven independent washouts. Moreover, 151 vials could be frozen from the remaining cells. The theoretical full expansion of all the frozen vials (validated by the expansion of two independent vials) could have allowed the production of 173 bags for adults and 348 bags for pediatric patients. CONCLUSIONS: The washouts of discarded bags and filters left over at the end of routine BM explants filtration are a very abundant source of hMSC precursors that can be easily utilized for clinical applications.

Induction of Mouse Lung Injury by Endotracheal Injection of Bleomycin.

Pulmonary fibrosis is a hallmark of several human lung diseases with a different etiology. Since current therapies are rather limited, mouse models continue to be an essential tool for developing new antifibrotic strategies. Here we provide an effective method to investigate in vivo antifibrotic activity of human mesenchymal stromal cells obtained from whole umbilical cord (hUC-MSC) in attenuating bleomycin-induced lung injury. C57BL/6 mice receive a single endotracheal injection of bleomycin (1.5 U/kg body weight) followed by a double infusion of hUC-MSC (2.5 x 105) into the tail vein, 24 h and 7 days after the bleomycin administration. Upon sacrifice at days 8, 14, or 21, inflammatory and fibrotic changes, collagen content, and hUC-MSC presence in explanted lung tissue are analyzed. The injection of bleomycin into the mouse trachea allows the direct targeting of the lungs, leading to extensive pulmonary inflammation and fibrosis. The systemic administration of a double dose of hUC-MSC results in the early blunting of the bleomycin-induced lung injury. Intravenously infused hUC-MSC are transiently engrafted into the mouse lungs, where they exert their anti-inflammatory and antifibrotic activity. In conclusion, this protocol has been successfully applied for the preclinical testing of hUC-MSC in an experimental mouse model of human pulmonary fibrosis. However, this technique can be easily extended both to study the effect of different endotracheally administered substances on the pathophysiology of the lungs and to validate new anti-inflammatory and antifibrotic systemic therapies.

A dual-phase xenon TPC for scintillation and ionisation yield measurements in liquid xenon.

A small-scale, two-phase (liquid/gas) xenon time projection chamber (Xurich II) was designed, constructed and is under operation at the University of Zürich. Its main purpose is to investigate the microphysics of particle interactions in liquid xenon at energies below 50 keV, which are relevant for rare event searches using xenon as target material. Here we describe in detail the detector, its associated infrastructure, and the signal identification algorithm developed for processing and analysing the data. We present the first characterisation of the new instrument with calibration data from an internal 83 m Kr source. The zero-field light yield is 15.0 and 14.0 photoelectrons/keV at 9.4 and 32.1 keV, respectively, and the corresponding values at an electron drift field of 1 kV/cm are 10.8 and 7.9 photoelectrons/keV. The charge yields at these energies are 28 and 31 electrons/keV, with the proportional scintillation yield of 24 photoelectrons per one electron extracted into the gas phase, and an electron lifetime of 200  µ s. The relative energy resolution, σ / E , is 11.9 and 5.8% at 9.4 and 32.1 keV, respectively using a linear combination of the scintillation and ionisation signals. We conclude with measurements of the electron drift velocity at various electric fields, and compare these to literature values.

Therapeutic potential of stromal cells of non-renal or renal origin in experimental chronic kidney disease.

BACKGROUND: Mesenchymal stromal cell (MSC)-based therapy is a promising strategy for preventing the progression of chronic kidney disease (CKD), with the potential to induce tissue regeneration. In search of the best cellular source we compared, in the rat model of adriamycin (ADR) nephropathy, the regenerative potential of human stromal cells of non-renal origin, such as bone marrow (bm) MSCs and umbilical cord (uc) MSCs, with that of newly discovered stromal cells of renal origin, the kidney perivascular cells (kPSCs) known to exhibit tissue-specific properties. METHODS: The therapeutic effect of repeated infusions of human bmMSCs, ucMSCs, kPSCs (1.5 × 106 cells/rats) or conditioned medium from ucMSCs was studied in athymic rats with ADR-induced nephropathy (7.9 mg/kg). The ability of the three stromal cell populations to engraft the damaged kidney was evaluated by detecting the presence of human nuclear antigenpos cells. Glomerular podocyte loss and endothelial damage, sclerotic lesions and inflammation were assessed at 14 and 28 days. In-vitro experiments with a transwell system were performed to investigate the effects of different stromal cell populations on parietal epithelial cells (PECs) activated or not with albumin or angiotensin II for 24 h. RESULTS: Infusions of non-renal and renal stromal cells resulted in a comparable engraftment into the kidney, in the peritubular areas and around the glomerular structures. All three cell populations limited podocyte loss and glomerular endothelial cell injury, and attenuated the formation of podocyte and PEC bridges. This translated into a reduction of glomerulosclerosis and fibrosis. Human ucMSCs had an anti-inflammatory effect superior to that of the other stromal cells, reducing macrophage infiltration and inducing polarisation towards the M2 macrophage phenotype. Conditioned medium from ucMSCs shared the same renoprotective effects of the cells. Consistent with in-vivo data, bmMSCs and kPSCs, but even more so ucMSCs, limited proliferation, migratory potential and extracellular matrix production of activated PECs, when cultured in a transwell system. CONCLUSIONS: Our data indicate that either non-renal or renal stromal cells induce renal tissue repair, highlighting ucMSCs and their conditioned medium as the most reliable clinical therapeutic tool for CKD patients.