RESUMEN
Due to its dual role as reporter and therapy gene, the sodium iodide symporter (NIS) allows noninvasive imaging of functional NIS expression by (123)I-scintigraphy or (124)I-PET imaging before the application of a therapeutic dose of (131)I. NIS expression provides a novel mechanism for the evaluation of mesenchymal stem cells (MSCs) as gene delivery vehicles for tumor therapy. In the current study, we stably transfected bone marrow-derived CD34(-) MSCs with NIS cDNA (NIS-MSC), which revealed high levels of functional NIS protein expression. In mixed populations of NIS-MSCs and hepatocellular cancer (HCC) cells, clonogenic assays showed a 55% reduction of HCC cell survival after (131)I application. We then investigated body distribution of NIS-MSCs by (123)I-scintigraphy and (124)I-PET imaging following intravenous (i.v.) injection of NIS-MSCs in a HCC xenograft mouse model demonstrating active MSC recruitment into the tumor stroma which was confirmed by immunohistochemistry and ex vivo γ-counter analysis. Three cycles of systemic MSC-mediated NIS gene delivery followed by (131)I application resulted in a significant delay in tumor growth. Our results demonstrate tumor-specific accumulation and therapeutic efficacy of radioiodine after MSC-mediated NIS gene delivery in HCC tumors, opening the prospect of NIS-mediated radionuclide therapy of metastatic cancer using MSCs as gene delivery vehicles.
Asunto(s)
Carcinoma Hepatocelular/radioterapia , Técnicas de Transferencia de Gen , Neoplasias Hepáticas/radioterapia , Células Madre Mesenquimatosas/patología , Simportadores/genética , Animales , Antígenos Transformadores de Poliomavirus/genética , Antígenos Transformadores de Poliomavirus/metabolismo , Western Blotting , Carcinoma Hepatocelular/genética , Supervivencia Celular , Femenino , Técnica del Anticuerpo Fluorescente , Terapia Genética/métodos , Células Hep G2 , Humanos , Radioisótopos de Yodo/farmacocinética , Neoplasias Hepáticas/genética , Células Madre Mesenquimatosas/metabolismo , Ratones , Ratones Desnudos , ARN Mensajero/genética , ARN Mensajero/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa , Simportadores/metabolismo , Transfección/métodosRESUMEN
BACKGROUND: Nodular inflammatory cell infiltrates with defined microarchitecture, i.e. tertiary lymphoid organs, develop in the tubulointerstitium during chronic renal inflammation. CCR6 and the corresponding ligand CCL20 are involved in the formation of gut-associated lymphatic tissue. We hypothesized that CCR6 might be involved in the formation of nodular infiltrates in the kidney. METHODS: CCR6- and CD20-positive B cells were localized in renal biopsies with IgA nephropathy (n = 13), membranous nephropathy (n = 12), crescentic glomerulonephritis (cGN, n = 11) and chronic interstitial nephritis (n = 13), and in pre-implantation biopsies as controls (n = 8). The mRNA expression of CCR6 and the ligand CCL20 was quantified by real-time RT-PCR in 51 renal biopsies of the same disease entities. RESULTS: In the pre-transplant biopsies, CCR6 was expressed by endothelial cells of peritubular and glomerular capillaries. In patients with glomerulonephritis, infiltrating cells were positive particularly in areas of nodular inflammatory cell accumulations. A major part of the CCR6-positive cells were CD20-positive B cells, but a part of the CD3-positive T cells were also found to be positive. The constitutive expression of CCR6 on the endothelium of glomerular capillaries was lost in biopsies with progressive injury. Tubular epithelial cells expressed CCR6 in inflamed kidneys, most commonly on the basolateral side. CONCLUSIONS: CCR6 and the corresponding ligand CCL20 might therefore be involved in the recruitment of T and B cells to organized nodular infiltrates in chronic renal inflammation. The functional role of endothelial CCR6 needs to be evaluated in further studies.
Asunto(s)
Quimiocina CCL20/metabolismo , Glomerulonefritis por IGA/metabolismo , Glomerulonefritis/metabolismo , Inflamación/metabolismo , Fallo Renal Crónico/metabolismo , Nefritis Intersticial/metabolismo , Receptores CCR6/metabolismo , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Estudios de Casos y Controles , Quimiocina CCL20/genética , Niño , Preescolar , Femenino , Técnica del Anticuerpo Fluorescente , Humanos , Técnicas para Inmunoenzimas , Lactante , Recién Nacido , Masculino , Persona de Mediana Edad , ARN Mensajero/genética , Receptores CCR6/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Adulto JovenRESUMEN
The Alport syndrome is a hereditary glomerular disease linked to structural abnormalities of collagen IV. In a mouse model of Alport syndrome, the interstitial lymphocyte influx was important for disease progression. CXCR3 is a chemokine receptor involved in lymphocyte recruitment to the kidney. We hypothesized that CXCR3-positive T cells might be involved in human Alport syndrome. Immunohistochemistry was performed on formalin-fixed, paraffin-embedded biopsies from 17 patients with Alport syndrome, 10 with immunoglobulin A (IgA) nephropathy, and 11 healthy donor kidneys. We investigated the expression of the alpha5 chain of collagen IV to confirm the morphologic diagnosis, the chemokine receptor CXCR3 and CD3-positive T cells. Alport syndrome biopsies demonstrated a complete loss of the alpha5 chain of collagen IV from the glomerular basement membrane and the morphologic features consistent with Alport syndrome on electron microscopy. A prominent number of CXCR3-positive cells were found in the tubulointerstitium. Most of the CXCR3-positive cells were CD3-positive T cells, demonstrated by double-labeling in selected biopsies. The number of CXCR3-positive cells in kidneys with Alport syndrome correlated with serum creatinine (P < .05) and with morphologic features of a progressive disease (eg, interstitial fibrosis, glomerulosclerosis, and tubular atrophy). The severity of interstitial CXCR3-positive cell influx was similar in Alport syndrome as compared to immunoglobulin A nephropathy. The noninflammatory glomerular lesion of Alport syndrome is associated with prominent interstitial accumulation of CD3- and CXCR3-positive lymphocytes. The degree of infiltration correlated with renal function. We speculate that targeting T lymphocytes, for example, by CXCR3 blocking agents, might be a novel approach to inhibit disease progression in patients with Alport syndrome.
Asunto(s)
Nefritis Hereditaria/inmunología , Nefritis Intersticial/inmunología , Adolescente , Adulto , Animales , Complejo CD3/metabolismo , Niño , Preescolar , Femenino , Humanos , Masculino , Nefritis Hereditaria/patología , Nefritis Intersticial/patología , Receptores CXCR3/metabolismo , Linfocitos T/inmunología , Linfocitos T/metabolismo , Adulto JovenRESUMEN
The Duffy antigen/receptor for chemokines (DARC) is a chemokine-binding protein that is expressed on erythrocytes and renal endothelial cells. DARC-mediated endothelial transcytosis of chemokines may facilitate the renal recruitment of macrophages and T cells, as has been suggested for neutrophils. We studied the role of Darc in two mouse models of prolonged renal inflammation, one that primarily involves the tubulointerstitium (unilateral ureteral obstruction), and one that requires an adaptive immune response that leads to glomerulonephritis (accelerated nephrotoxic nephritis). Renal expression of Darc and its ligands was increased in both models. Leukocytes effectively infiltrated obstructed kidneys in Darc-deficient mice with pronounced T-cell infiltration at early time points. Development of interstitial fibrosis was comparable in both genotypes. Nephrotoxic nephritis was inducible in Darc-deficient mice, with both an increased humoral immune response and functional impairment during the early phase of disease. Leukocytes efficiently infiltrated kidneys of Darc-deficient mice, with increased cell numbers at early but not late time points. Taken together, renal inflammation developed more rapidly in DARC-deficient mice, without affecting the extent of renal injury at later time points. Thus, genetic elimination of Darc in mice does not prevent the development of renal infiltrates and may even enhance such development during the early phases of interstitial and glomerular diseases in mouse models of prolonged renal inflammation.
Asunto(s)
Quimiotaxis de Leucocito , Sistema del Grupo Sanguíneo Duffy , Glomerulonefritis , Macrófagos , Receptores de Superficie Celular , Linfocitos T , Animales , Ratones , Quimiocinas/inmunología , Quimiotaxis de Leucocito/inmunología , Sistema del Grupo Sanguíneo Duffy/genética , Sistema del Grupo Sanguíneo Duffy/inmunología , Ensayo de Inmunoadsorción Enzimática , Fibrosis , Citometría de Flujo , Glomerulonefritis/inmunología , Glomerulonefritis/patología , Procesamiento de Imagen Asistido por Computador , Inmunoglobulina G/sangre , Inmunohistoquímica , Macrófagos/inmunología , Ratones Endogámicos C57BL , Ratones Noqueados , Receptores de Superficie Celular/deficiencia , Receptores de Superficie Celular/genética , Receptores de Superficie Celular/inmunología , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Linfocitos T/inmunologíaRESUMEN
Local B-cell infiltrates play a role in tissue fibrosis, neolymphangiogenesis, and renal allograft survival. We sought to characterize the B-cell infiltrates, factors involved in B-cell recruitment, and lymphangiogenesis in renal interstitial injury (ie, acute and chronic interstitial nephritis and chronic IgA nephropathy). CD20-positive B cells formed a prominent part of the interstitial infiltrating cells. Together with CD3-positive T cells, the CD20-positive B cells formed larger nodular structures. CD10-positive pre-B cells were rare, and the majority were mature CD27-positive B cells. Proliferating B cells were detected within nodular infiltrates. The level of mRNA expression of the chemokine CXCL13 was increased and correlated with CD20 mRNA in the tubulointerstitial space. CXCL13 protein was predominantly found at sites of nodular infiltrates, in association with CXCR5-positive B cells. Furthermore, sites of chronic interstitial inflammation were associated with a high number of lymphatic vessels. B-cell infiltrates form a prominent part of the interstitial infiltrates both in primary interstitial lesions and in IgA nephropathy. CXCR5-positive B cells might be recruited via the chemokine CXCL13 and seem to contribute to the formation of intrarenal lymphoid follicle-like structures. These might represent an intrarenal immune system.