Mario Serio

Isolation and Characterization of Multipotent Progenitor Cells from the Bowman’s Capsule of Adult Human Kidneys

Regenerative medicine represents a critical clinical goal for patients with ESRD, but the identification of renal adult multipotent progenitor cells has remained elusive. It is demonstrated that in human adult kidneys, a subset of parietal epithelial cells (PEC) in the Bowmans capsule exhibit coexpression of the stem cell markers CD24 and CD133 and of the stem cell-specific transcription factors Oct-4 and BmI-1, in the absence of lineage-specific markers. This CD24+CD133+ PEC population, which could be purified from cultured capsulated glomeruli, revealed self-renewal potential and a high cloning efficiency. Under appropriate culture conditions, individual clones of CD24+CD133+ PEC could be induced to generate mature, functional, tubular cells with phenotypic features of proximal and/or distal tubules, osteogenic cells, adipocytes, and cells that exhibited phenotypic and functional features of neuronal cells. The injection of CD24+CD133+ PEC but not of CD24-CD133- renal cells into SCID mice that had acute renal failure resulted in the regeneration of tubular structures of different portions of the nephron. More important, treatment of acute renal failure with CD24+CD133+ PEC significantly ameliorated the morphologic and functional kidney damage. This study demonstrates the existence and provides the characterization of a population of resident multipotent progenitor cells in adult human glomeruli, potentially opening new avenues for the development of regenerative medicine in patients who have renal diseases.

An Alternatively Spliced Variant of CXCR3 Mediates the Inhibition of Endothelial Cell Growth Induced by IP-10, Mig, and I-TAC, and Acts as Functional Receptor for Platelet Factor 4

The chemokines CXCL9/Mig, CXCL10/IP-10, and CXCL11/I-TAC regulate lymphocyte chemotaxis, mediate vascular pericyte proliferation, and act as angiostatic agents, thus inhibiting tumor growth. These multiple activities are apparently mediated by a unique G protein–coupled receptor, termed CXCR3. The chemokine CXCL4/PF4 shares several activities with CXCL9, CXCL10, and CXCL11, including a powerful angiostatic effect, but its specific receptor is still unknown. Here, we describe a distinct, previously unrecognized receptor named CXCR3-B, derived from an alternative splicing of the CXCR3 gene that mediates the angiostatic activity of CXCR3 ligands and also acts as functional receptor for CXCL4. Human microvascular endothelial cell line-1 (HMEC-1), transfected with either the known CXCR3 (renamed CXCR3-A) or CXCR3-B, bound CXCL9, CXCL10, and CXCL11, whereas CXCL4 showed high affinity only for CXCR3-B. Overexpression of CXCR3-A induced an increase of survival, whereas overexpression of CXCR3-B dramatically reduced DNA synthesis and up-regulated apoptotic HMEC-1 death through activation of distinct signal transduction pathways. Remarkably, primary cultures of human microvascular endothelial cells, whose growth is inhibited by CXCL9, CXCL10, CXCL11, and CXCL4, expressed CXCR3-B, but not CXCR3-A. Finally, monoclonal antibodies raised to selectively recognize CXCR3-B reacted with endothelial cells from neoplastic tissues, providing evidence that CXCR3-B is also expressed in vivo and may account for the angiostatic effects of CXC chemokines.

Regeneration of Glomerular Podocytes by Human Renal Progenitors

Depletion of podocytes, common to glomerular diseases in general, plays a role in the pathogenesis of glomerulosclerosis. Whether podocyte injury in adulthood can be repaired has not been established. Here, we demonstrate that in the adult human kidney, CD133+CD24+ cells consist of a hierarchical population of progenitors that are arranged in a precise sequence within Bowmans capsule and exhibit heterogeneous potential for differentiation and regeneration. Cells localized to the urinary pole that expressed CD133 and CD24, but not podocyte markers (CD133+CD24+PDX- cells), could regenerate both tubular cells and podocytes. In contrast, cells localized between the urinary pole and vascular pole that expressed both progenitor and podocytes markers (CD133+CD24+PDX+) could regenerate only podocytes. Finally, cells localized to the vascular pole did not exhibit progenitor markers, but displayed phenotypic features of differentiated podocytes (CD133-CD24-PDX+ cells). Injection of CD133+CD24+PDX- cells, but not CD133+CD24+PDX+ or CD133-CD24- cells, into mice with adriamycin-induced nephropathy reduced proteinuria and improved chronic glomerular damage, suggesting that CD133+CD24+PDX- cells could potentially treat glomerular disorders characterized by podocyte injury, proteinuria, and progressive glomerulosclerosis.

Cell cycle–dependent expression of CXC chemokine receptor 3 by endothelial cells mediates angiostatic activity

Endothelial cell receptors for the angiostatic chemokines IFN-gamma-inducible protein of 10 kDa (IP-10) and monokine induced by IFN-gamma (Mig) have not yet been identified, and the mechanisms responsible for the effects of these chemokines on angiogenesis are still unclear. IP-10 and Mig share a common functional receptor on activated T lymphocytes, named CXC chemokine receptor 3 (CXCR3). Using in situ hybridization and immunohistochemistry, we show that CXCR3 is expressed by a small percentage of microvascular endothelial cells in several human normal and pathological tissues. Primary cultures of human microvascular endothelial cells (HMVECs) likewise express CXCR3, although this expression is limited to the S/G2-M phase of their cell cycle. Both IP-10 and Mig, as well as the IFN-gamma-inducible T-cell alpha chemoattractant (I-TAC), which all share high-affinity binding for CXCR3, block HMVEC proliferation in vitro, an effect that can be inhibited by an anti-CXCR3 antibody. These data provide definitive evidence of CXCR3 expression by HMVEC and open new avenues for therapeutic interventions in all conditions in which an angiostatic effect may be beneficial.

Essential but differential role for CXCR4 and CXCR7 in the therapeutic homingof human renal progenitor cells

Recently, we have identified a population of renal progenitor cells in human kidneys showing regenerative potential for injured renal tissue of SCID mice. We demonstrate here that among all known chemokine receptors, human renal progenitor cells exhibit high expression of both stromal-derived factor-1 (SDF-1) receptors, CXCR4 and CXCR7. In SCID mice with acute renal failure (ARF), SDF-1 was strongly up-regulated in resident cells surrounding necrotic areas. In the same mice, intravenously injected renal stem/progenitor cells engrafted into injured renal tissue decreased the severity of ARF and prevented renal fibrosis. These beneficial effects were abolished by blocking either CXCR4 or CXCR7, which dramatically reduced the number of engrafting renal progenitor cells. However, although SDF-1–induced migration of renal progenitor cells was only abolished by an anti-CXCR4 antibody, transendothelial migration required the activity of both CXCR4 and CXCR7, with CXCR7 being essential for renal progenitor cell adhesion to endothelial cells. Moreover, CXCR7 but not CXCR4 was responsible for the SDF-1–induced renal progenitor cell survival. Collectively, these findings suggest that CXCR4 and CXCR7 play an essential, but differential, role in the therapeutic homing of human renal progenitor cells in ARF, with important implications for the development of stem cell–based therapies.

Regenerative Potential of Embryonic Renal Multipotent Progenitors in Acute Renal Failure

Bone marrow-and adult kidney-derived stem/progenitor cells hold promise in the development of therapies for renal failure. Here is reported the identification and characterization of renal multipotent progenitors in human embryonic kidneys that share CD24 and CD133 surface expression with adult renal progenitors and have the capacity for self-renewal and multilineage differentiation. It was found that these CD24+CD133+ cells constitute the early primordial nephron but progressively disappear during nephron development until they become selectively localized to the urinary pole of Bowmans capsule. When isolated and injected into SCID mice with acute renal failure from glycerol-induced rhabdomyolysis, these cells regenerated different portions of the nephron, reduced tissue necrosis and fibrosis, and significantly improved renal function. No tumorigenic potential was observed. It is concluded that CD24+CD133+ cells represent a subset of multipotent embryonic progenitors that persist in human kidneys from early stages of nephrogenesis. The ability of these cells to repair renal damage, together with their apparent lack of tumorigenicity, suggests their potential in the treatment of renal failure.

Characterization of human adult stem-cell populations isolated from visceral and subcutaneous adipose tissue

Adipose tissue is a dynamic endocrine organ with a central role in metabolism regulation. Functional differences in adipose tissue seem associated with the regional distribution of fat depots, in particular in subcutaneous and visceral omental pads. Here, we report for the first time the isolation of human adipose‐derived adult stem cells from visceral omental and subcutaneous fat (V‐ASCs and S‐ASCs, respectively) from the same subject. Immunophenotyping shows that plastic culturing selects homogeneous cell populations of V‐ASCs and S‐ASCs from the corresponding stromal vascular fractions (SVFs), sharing typical markers of mesenchymal stem cells. Electron microscopy and electrophysiological and real‐time RT‐PCR analyses confirm the mesenchymal stem nature of both V‐ASCs and S‐ASCs, while no significant differences in a limited pattern of cytokine/chemokine expression can be detected. Similar to S‐ASCs, V‐ASCs can differentiate in vitro toward adipogenic, osteogenic, chondrogenic, muscular, and neuronal lineages, as demonstrated by histochemical, immunofluorescence, real‐time RT‐PCR, and electrophysiological analyses, suggesting the multipotency of such adult stem cells. Our data demonstrate that both visceral and subcutaneous adipose tissues are a source of pluripotent stem cells with multigermline potential. However, the visceral rather than the subcutaneous ASC could represent a more appropriate in vitro cell model for investigating the molecular mechanisms implicated in the pathophysiology of metabolic disorders such as obesity.—Baglioni, S., Francalanci, M., Squecco, R., Lombardi, A., Cantini, G., Angeli, R., Gelmini, S., Guasti, D., Benvenuti, S., Annunziato, F., Bani, D., Liotta, F., Francini, F., Perigli, G., Serio, M., Luconi, M. Characterization of human adult stem‐cell populations isolated from visceral and subcutaneous adipose tissue. FASEB J. 23, 3494–3505 (2009). www.fasebj.org

Pleiotropic effects of thiazolidinediones: Taking a look beyond antidiabetic activity

Thiazolidinediones (TZD) [Troglitazone (TRO), Pioglitazone (PGZ), Rosiglitazone, (RGZ)] are a novel class of antidiabetic drugs for patients with Type-2 diabetes mellitus (T2DM) able to decrease blood glucose, working through a reduction of insulin resistance. The family of TZD exerts its effect specifically bound to peroxisome proliferator- activated receptor ψ (PPARψ). This is a member of the nuclear hormone receptor superfamily of ligand-dependent transcription factors, together with PPARa and dgB. Although PPARψ is essentially expressed in adipose tissue, it has also been found in endothelial cells, macrophages, vascular smooth muscle cells, glomerular mesangial cells, hepatic stellate cells and in several cancer cell lines. In these cells, the PPARψ activation by TZD determines modulatory effects on growth factor release, production of cytokine, cell proliferation and migration, extracellular matrix remodeling and control on cell cycle progression and differentiation. In addition, TZD have been shown to have a potent antioxidant effect. This review, taking a quick look beyond the antidiabetic activity of PPARψ, shows the dramatic ranging of medical implications that the use of TZD could have modulating the PPARψ activity in several diseases with a strong social impact, such as insulin resistance syndrome, chronic inflammation, atherosclerosis and cancer.

Androgen Receptor Expression in Prostate Carcinoma Cells Suppresses α6β4 Integrin-Mediated Invasive Phenotype1

Prostate cancer cells may lose androgen-sensitivity after androgen ablation therapy, becoming highly invasive and metastatic. The biological mechanisms responsible for higher tumurogenicity of androgen-independent prostate carcinomas are not entirely known. We demonstrate that androgen receptor regulation of adhesion and invasion of prostate cancer cells through modulation of α6β4 integrin expression may be one of the molecular mechanisms responsible of this phenomenon. We found that protein and gene expressions of α6 and β4 subunits were strongly reduced in the androgen-sensitive cell line LNCaP respect to the androgen-independent PC3 and that transfection of PC3 cells with a full-length androgen receptor expression vector resulted in a decreased expression of α6β4 integrin, reduced adhesion on laminin, and suppressed Matrigel invasion. Growth in soft agar was also suppressed in androgen receptor-positive PC3 clones. Treatment of androgen receptor positive clones with the synthetic androgen R1881 further...

PLASMA ANDROGENS IN WOMEN WITH HYPERPROLACTINAEMIC AMENORRHOES

Cortisol, androstenedione, testosterone, dehydroepiandrosterone sulphate (DHAS) and free dehydroepiandrosterone (DHA) were measured in plasma of ten women affected by amenorrhoea with hyperprolactinaemia and eleven women affected by secondary hypothalamic amenorrhoea; twelve normal women at the second day of the menstrual cycle were used as controls. All subjects were hospitalized and 17‐ketosteroids, 170H‐corticosteroids and total dehydroepiandrosterone were also measured in urine.