Long-term effects of different stem cells in genetic models of Cystic Kidney Disease

Cystic kidney diseases are a global public problem, as the population of patients is increasing at a rate of approximately 7% per year. Unfortunately, up to now there is a lack of efficient therapies that can prevent the progressive loss of renal function. Drug treatment is of limited. The only therapeutic alternatives are dialysis or kidney transplantation. Nevertheless, the human and economic impact of the diseases to affected individuals and to medical community and society alike is enormous. Therefore new therapies are urgently needed. Stem cell application represents a promising therapeutic approach. MSC therapies have been used extensively, in the recent years, as a possible therapy for other kidney diseases with the aim of slowing down the course of the disease. However, many aspects remain unclear or are under debate like the finding of an appropriate source of MSCs, the understanding of their modes of action and, not least, the dose and timing of the administration. Aim of this study was to evaluate the potential therapeutic effects of two different types of stem cells, and the derived conditioned media, in two animal models resembling human cystic kidney diseases. In order to achieve this goal, firstly we characterized, on a long-term basis, two different genetic animal models: the PKD/Mhm (Cy/+) and the PCK rats. Afterwards, we performed a 6 months trial, to test the long-term effects of human ASC and human ABCB5+ cells and ASC derived CM and ABCB5+ derived CoCM+. Animals were classified in four different groups, depending on the treatment received: (i) group that did not received treatment, (ii) ASC derived CM or ABCB5+ derive CoCM+ group, (iii) i.p. ASC or ABCB5+ group and (iv) i.v. ASC or ABCB5+ group. The progression of the disease and the effect of the treatments were determined on the basis of plasma and urine biochemistry, transcutaneous measurement of renal function, histology evaluation and gene expression profiling. Our results found a different disease severity in the two models. PCK rats presented, since the beginning of the project, worse clinical manifestations compared to PKD/Mhm (Cy/+) probably due to the different genetic background. ABCB5+ and ASC treatments led to an improvement in renal function reflected by GFR, plasma levels of creatinine, albuminuria and proteinuria, in PKD/Mhm (Cy/+) model. Histological evaluation of the kidney revealed a reduction of apoptosis and cell proliferation. Moreover, comparable genetic changes were reported after these treatments. Also the ABCB5+ derived CoCM+ treatment proved to ameliorate the biochemical parameters, while ASC derived CM treatment had a lesser pronounced outcome. Both after ABCB5+ derived CoCM+ or ASC derived CM administration the histological changes in apoptosis and proliferation reduction was observed. Ultimately, these animals undergo to genetic changes similar to the one observed after the cells treatment. Concerning the PCK model. ABCB5+ and ABCB5+ derived CoCM+ treatments slightly ameliorated kidney function, plasma and urine parameter and likewise the levels of apoptotic and proliferative positive markers. Administration of ASC or ASC derived CM improved the renal function and decreased the apoptotic and proliferative positive cells but had only a mild effect on other parameter involved in the kidney function. Additionally, the gene expression profile did not highlight significant genetic changes after those treatments, while ABCB5+ and ABCB5+ derived CoCM+ administrations induced beneficial genetic changes. In conclusion, our results demonstrate that ABCB5+ or ASC cells administration, either i.v. or i.p., and ABCB5+ derived CoCM+ might be a valid alternative therapy for cystic kidney diseases. However, before a possible application in the clinical field further studies need to be carried out in order to better define the mode of action and the side effects of these therapies

DNAM-1 chimeric receptor-engineered NK cells: a new frontier for CAR-NK cell-based immunotherapy

DNAM-1 is a major NK cell activating receptor and, together with NKG2D and NCRs, by binding specific ligands, strongly contributes to mediating the killing of tumor or virus-infected cells. DNAM-1 specifically recognizes PVR and Nectin-2 ligands that are expressed on some virus-infected cells and on a broad spectrum of tumor cells of both hematological and solid malignancies. So far, while NK cells engineered for different antigen chimeric receptors (CARs) or chimeric NKG2D receptor have been extensively tested in preclinical and clinical studies, the use of DNAM-1 chimeric receptor-engineered NK cells has been proposed only in our recent proof-of-concept study and deserves further development. The aim of this perspective study is to describe the rationale for using this novel tool as a new anti-cancer immunotherapy

Combined treatment with inhibitors of ErbB Receptors and Hh signaling pathways is more effective than single treatment in reducing the growth of malignant mesothelioma both in vitro and in vivo

Malignant mesothelioma (MM) is a rare orphan aggressive neoplasia with low survival rates. Among the other signaling pathways, ErbB receptors and Hh signaling are deregulated in MM. Thus, molecules involved in these signaling pathways could be used for targeted therapy approaches. The aim of this study was to evaluate the effects of inhibitors of Hh- (GANT-61) and ErbB receptors (Afatinib)-mediated signaling pathways, when used alone or in combination, on growth, cell cycle, cell death and autophagy, modulation of molecules involved in transduction pathways, in three human MM cell lines of different histotypes. The efficacy of the combined treatment was also evaluated in a murine epithelioid MM cell line both in vitro and in vivo. This study demonstrated that combined treatment with two inhibitors counteracting the activation of two different signaling pathways involved in neoplastic transformation and progression, such as those activated by ErbB and Hh signaling, is more effective than the single treatments in reducing MM growth in vitro and in vivo. This study may have clinical implications for the development of targeted therapy approaches for MM

The proper interplay between the expression of Spo11 splice isoforms and the structure of the pseudoautosomal region promotes XY chromosomes recombination

XY chromosome missegregation is relatively common in humans and can lead to sterility or the generation of aneuploid spermatozoa. a leading cause of XY missegregation in mammals is the lack of formation of double-strand breaks (DSBs) in the pseudoautosomal region (PAR), a defect that may occur in mice due to faulty expression of Spo11 splice isoforms. using a knock-in (ki) mouse that expresses only the single Spo11 ss splice isoform, here we demonstrate that by varying the genetic background of mice, the length of chromatin loops extending from the PAR axis and the XY recombination proficiency varies.in spermatocytes of C57(Spo11 ss ki/-) mice, in which loops are relatively short, recombination/synapsis between XY is fairly normal. In contrast, in cells of C57/129(Spo11 ss ki/-) males where PAR loops are relatively long, formation of DSBs in the PAR (more frequently the Y-PAR) and XY synapsis fails at a high rate, and mice produce sperm with sex-chromosomal aneuploidy. However, if the entire set of Spo11 splicing isoforms is expressed by a wild type allele in the C57/129 background, XY recombination and synapsis is recovered. By generating a Spo11aki mouse model, we prove that concomitant expression of SPO11 ss and SPO11a isoforms, boosts DSB formation in the PAR. Based on these findings, we propose that SPO11 splice isoforms cooperate functionally in promoting recombination in the PAR, constraining XY asynapsis defects that may arise due to differences in the conformation of the PAR between mouse strains

Phylogenetic and Phylodynamic Analyses of HCV Strains Circulating among Patients Using Injectable Drugs in Central Italy

Approximately 71 million people worldwide are infected with the hepatitis C virus (HCV). Injectable drug use represents the most common route of transmission in Europe and other developed countries. We studied the molecular characteristics of the HCV infection among mono-infected people who used drugs (PWUD) in Italy. Among 208 PWUD with anti-HCV antibodies, 101 (48.6%) were HCV RNA-positive, the majority (47%) were infected with the HCV genotype (Gt)1a, followed by Gt3a (34.9%), Gt4 (9.1%), Gt1b (4.5%), and Gt2 (4.5%). Bayesian phylogenetic analyses of clustered HCV NS5B sequences from 66 HCV-positive PWUDs with available plasma samples indicated age and neighborhood proximity as the most common characteristics between closely related HCV strains. Population dynamics, as measured by a coalescent Bayesian skyline analysis, revealed an increase in HCV Gt1a infections from the mid-1980s to mid-1990s. While HCV Gt3a infections were first detected in the 1980s, patient numbers with this genotype subtype remained relatively constant. For both Gt1a and Gt3a, Birth–Death Bayesian Skyline analyses produced higher reproduction numbers post 2014. For earlier time intervals, slow growths were observed for both Gt1a and Gt3a with reproduction numbers (Re) of approximately 1. The evolutionary rates for Gt1a and Gt3a were estimated as 2.23 × 10−4 and 3.85 × 10−4, respectively

Potential Therapeutic Effects of Long-Term Stem Cell Administration: Impact on the Gene Profile and Kidney Function of PKD/Mhm (Cy/+) Rats

Cystic kidney disease (CKD) is a heterogeneous group of genetic disorders and one of the most common causes of end-stage renal disease. Here, we investigate the potential effects of long-term human stem cell treatment on kidney function and the gene expression profile of PKD/Mhm (Cy/+) rats. Human adipose-derived stromal cells (ASC) and human skin-derived ABCB5+ stromal cells (2 × 106) were infused intravenously or intraperitoneally monthly, over 6 months. Additionally, ASC and ABCB5+-derived conditioned media were administrated intraperitoneally. The gene expression profile results showed a significant reprogramming of metabolism-related pathways along with downregulation of the cAMP, NF-kB and apoptosis pathways. During the experimental period, we measured the principal renal parameters as well as renal function using an innovative non-invasive transcutaneous device. All together, these analyses show a moderate amelioration of renal function in the ABCB5+ and ASC-treated groups. Additionally, ABCB5+ and ASC-derived conditioned media treatments lead to milder but still promising improvements. Even though further analyses have to be performed, the preliminary results obtained in this study can lay the foundations for a novel therapeutic approach with the application of cell-based therapy in CKD

Stem/Stromal Cells for Treatment of Kidney Injuries With Focus on Preclinical Models

Within the last years, the use of stem cells (embryonic, induced pluripotent stem cells, or hematopoietic stem cells), Progenitor cells (e.g., endothelial progenitor cells), and most intensely mesenchymal stromal cells (MSC) has emerged as a promising cell-based therapy for several diseases including nephropathy. For patients with end-stage renal disease (ESRD), dialysis or finally organ transplantation are the only therapeutic modalities available. Since ESRD is associated with a high healthcare expenditure, MSC therapy represents an innovative approach. In a variety of preclinical and clinical studies, MSC have shown to exert renoprotective properties, mediated mainly by paracrine effects, immunomodulation, regulation of inflammation, secretion of several trophic factors, and possibly differentiation to renal precursors. However, studies are highly diverse; thus, knowledge is still limited regarding the exact mode of action, source of MSC in comparison to other stem cell types, administration route and dose, tracking of cells and documentation of therapeutic efficacy by new imaging techniques and tissue visualization. The aim of this review is to provide a summary of published studies of stem cell therapy in acute and chronic kidney injury, diabetic nephropathy, polycystic kidney disease, and kidney transplantation. Preclinical studies with allogeneic or xenogeneic cell therapy were first addressed, followed by a summary of clinical trials carried out with autologous or allogeneic hMSC. Studies were analyzed with respect to source of cell type, mechanism of action etc

H2AFX and MDC1 promote maintenance of genomic integrity in male germ cells

In somatic cells, H2afx and Mdc1 are close functional partners in DNA repair and damage response. However, it is not known whether they are also involved in the maintenance of genome integrity in meiosis. By analyzing chromosome dynamics in H2afx spermatocytes, we found that the synapsis of autosomes and X-Y chromosomes was impaired in a fraction of cells. Such defects correlated with an abnormal recombination profile. Conversely, Mdc1 was dispensable for the synapsis of the autosomes and played only a minor role in X-Y synapsis, compared with the action of H2afx. This suggested that those genes have non-overlapping functions in chromosome synapsis. However, we observed that both genes play a similar role in the assembly of MLH3 onto chromosomes, a key step in crossover formation. Moreover, we show that H2afx and Mdc1 cooperate in promoting the activation of the recombination-dependent checkpoint, a mechanism that restrains the differentiation of cells with unrepaired DSBs. This occurs by a mechanism that involves P53. Overall, our data show that, in male germ cells, H2afx and Mdc1 promote the maintenance of genome integrity