1,721,023 research outputs found
Heparanase e rene. Regolazione e ruolo dell'heparanase a livello tubulare e nella nefropatia diabetica.
Full text linkHeparanase (HPSE) is an interesting player in the arena of proteinuric disease and several investigators proposed that this enzyme is responsible for heparan sulphate (HS) reduction in glomerular basement membrane in diabetic nephropathy (DN). However, few data are available about HPSE expression, regulation and role in proximal tubular epithelial cells (PTEC), particularly in diabetes.
In this study we demonstrate that this molecule is a key mediator in homeostasis of PTEC in diabetic milieu possibly playing a role in the DN pathogenesis.
We showed that HPSE is constitutively expressed in PTEC suggesting that it has a physiological role. HPSE is up-regulated by mediators involved in DN such as albumin and glycated albumin (AGE), but not by high-glucose. Albumin and AGE-induced HPSE up-regulation is PI3K/AKT dependent.
Albumin and especially AGE cause a significant decrease in HS staining. In order to investigate the effects derived from HPSE up-regulation and distinguish from those caused directly by albumin and AGE we generated a HK2 PTEC line stably silenced for HPSE. In HPSE silenced cells, HS was significantly increased respect to HK2 wt cells and treatment with albumin or AGE had no effect suggesting that in tubular cells, HS levels depend on HPSE regulation. We also discovered that HS levels aren't regulated only by the cleaving activity of the endoglycosidase HPSE but also from the regulation of HSPG core proteins. In fact, Syndecan-1 core protein and mRNA expression were down-regulated by albumin and AGE in w.t. tubular cells while in HPSE silenced cells they failed to induce any Syndecan-1 modulation. Moreover in untreated HPSE silenced cells, Syndecan-1 is up-regulated at both gene transcription and protein expression in comparison with w.t. cells. Preliminary data also indicate that HPSE is involved in the regulation of FGF-2 signaling.
We has also verified that exist a correlation between heparanase and metalloproteinases (MMP), infact in HPSE-silenced cells these extracellular matrix degrading enzymes results down-regulated both at gene transcription and activity levels in comparison with wt cells. MMPs are also up-regulated by albumin and AGE stimulation in a heparanase-independet way.
In conclusion, we showed that HPSE is implicated in different mechanisms possibly involved in the tubulointerstitial damage of the DN.
We have alco confirmed that Sulodexide, a drug commonly used in DN treatment which is effective in reducing proteinuria, is a potent heparanase inhibitor with a IC50 of 50mg/ml.
Finally, we observed that heparanase plasma activity is up-regulated in nephropatic patient, expecially in transplant rejection, respect to healthy subjects. These data must be related to other parameter of kidney functionality to recognize heparanase activity as a useful marker of nephropathy.L'heparanase (HPSE) sta emergendo come un importante fattore nel complesso sistema delle nefropatie proteinuriche e percio diversi autori hanno suggerito che questo enzima possa essere responsabile della riduzione dell'heparan solfato (HS) nella membrana basale glomerulare nel corso della nefropatia diabetica (ND). Attualmente sono disponibili poche informazioni riguardo l'espressione, la regolazione e il ruolo dell'heparanase nelle cellule epiteliali del tubulo prossimale (PTEC) in particolare nel diabete.
In questo studio è stato dimostrato che questo enzima è un mediatore chiave nell'omeostasi delle PTEC nell'ambiente diabetico e può svolgere un ruolo nella patogenesi della ND. È stato evidenziato che l'heparanase è espressa costitutivamente nelle PTEC suggerendo che essa svolga un ruolo fisiologico in queste cellule. La sovra-espressione dell'heparanase è mediata da alcuni fattori tipici della ND, quali l'albumina e i prodotti di avanzata glicazione (AGE), ma non dall'alto glucosio. L'albumina e gli AGE portatano alla sovra-espressione dell'heparanase attraverso la via PI3K/AKT.
L'albumina e particolarmente gli AGE causano una significativa riduzione dell'HS. Al fine di discriminare gli effetti dovuti alla sovra-espressione dell'heparanase da quelli causati direttamente dall'albumina e dagli AGE abbiamo generato una linea cellulare (HK2) stabilmente silenziata per l'heparanase. Nelle cellule HPSE-silenziate l'HS è significativamente aumentato rispetto alle cellule HK2 wt e il trattamento con albumina e AGE non ha prodotto effetti suggerendo che nelle PTEC i livelli di HS dipendono dalla regolazione dell'heparanase.
È inoltre emerso che i livelli di HS non sono regolati solamente dall'attività endoglicosidasica dell'heparanase ma anche dalla regolazione del core proteico dei proteoglicani dell'HS. Infatti, il core proteico del sindecano-1 e la sua espressione genica sono ridotte nelle cellule tubulari wt trattate con albumina ed AGE mentre nelle cellule HPSE silenziate questi mediatori non riescono ad indurre alcuna modulazione del sindecano-1. Nelle cellule HPSE-silenziate, inoltre, il sindecano-1 risulta significativamente incrementato sia a livello di trascrizione genica che della relativa espressione proteica rispetto alle cellule wt. Ulteriori dati confermano che l'heparanase è coinvolta nelle vie di trasduzione del segnale dell'FGF-2.
È stato poi verificato che esiste una correlazione fra heparanase e metalloproteasi (MMP), infatti nelle cellule HPSE silenziate questi enzimi di degradazione della matrice extracellulare risultano ridotti sia a livello di espressione genica che di attività . Anche le MMP risultano incrementate dalla stimolazione con albumina e AGE ma in maniera indipendente dall'heparanase.
Inoltre il Sulodexide, un farmaco comunemente usato nel trattamento della ND e che è risultato efficace nel ridurre la proteinuria, è effettivamente un buon inibitore dell'heparanase con un IC50 di 50mg/ml.
Infine è stato osservato che l'attività plasmatica dell'heparanase risulta aumentata in pazienti con nefropatie, speciamemte in pazienti con rigetto di trapianto, rispetto a soggetti sani. Al fine di evidenziare l'utilita di questi valori come marcatori di qualche condizione nefropatica dovranno essere correlati ad altri parametri di funzionalià renale.
In conclusione, si ritiene di avere dimostrato un ruolo attivo dell'l'heparanase nei meccanismi di danno e patogenesi della ND
LGI1 Affects survival of neuroblastoma cells by inhibiting signalling through phosphoinositide 3-Kinase
No full textOverexpression of the leucine-rich, glioma-inactivated 1 (LGI1) gene in neuroblastoma cells inhibited proliferation and efficiently induced apoptosis. Cell clones stably transfected with LGI1 cDNA showed greater mortality during a period of serum starvation in comparison with control cells stably transfected with empty vector. This observation suggested hindrance of the PI3K/Akt pathway, a central transducer of survival stimuli elicited by serum growth factors. Treatment with inhibitors of PI3K significantly increased the death of control cells but substantially failed to influence LGI1 cell death, which was greatest independently of the presence of inhibitors. Blockage of the PI3K/Akt pathway in LGI1 cells was confirmed by the lack of serum-induced Akt phosphorylation, in contrast with the strong response of control cells. Instead, serum-induced phosphorylation of ERK1/2 was not impaired by the expression of LGI1. This study showed that overexpression of LGI1 caused neuroblastoma cell death by blocking activation of the PI3K/Akt pathway. Thus, the possibility of upregulating LGI1 expression may be a novel strategy in suppressing oncogenesis and metastasis sustained by excessive activation of the PI3K/Akt pathway
The Story of SPATA2 (Spermatogenesis-Associated Protein 2): From Sertoli Cells to Pancreatic Beta-Cells
No full text
The role of SPATA2 in TNF signaling, cancer, and spermatogenesis
Full text linkThe activation of TNF receptors can lead to cell death with a mechanism of cell necrosis regulated genetically and distinct from apoptosis which is defined as necroptosis. Necroptosis has been one of the most studied emerging cell death/signaling pathways in recent years, especially in light of the role of this process in human disease. However, not all regulatory components of TNF signaling have been identified in relation to both physiological and pathological conditions. In 2008, Spata2 (Spermatogenesis-associated protein 2) was identified as one of the seven fundamental genes for the cellular signaling network that regulates necroptosis and apoptosis. This gene had been cloned by our group and named Spata2 as its expression was found to be elevated in the testis compared to other tissues, localized at the Sertoli cell level and FSH-dependent. More recently, it has been demonstrated that deletion of Spata2 gene causes increased inhibin α expression and attenuated fertility in male mice. However, more importantly, five recently published reports have highlighted that SPATA2 is crucial for recruiting CYLD to the TNFR1 signaling complex thus promoting its activation leading to TNF-induced cell death. Loss of SPATA2 increases transcriptional activation of NF-kB and limits TNF-induced necroptosis. Here we will discuss these important findings regarding SPATA2 and, in particular, focus attention on the evidence that suggests a role for this protein in the TNF signaling pathway
The story of SPATA2 (Spermatogenesis-associated Protein 2): from Sertoli cells to pancreatic beta-cells.
Full text linkIn an attempt to isolate new spermatogenesis-associated genes, pd1 was initially identified and cloned as a novel human cDNA sequence from testis cDNA library. The novel gene was submitted to GenBank under accession n degrees U28164 in 1996. PD1 expression was demonstrated at the Sertoli cell level with a production which appeared to be under the influence of neighbouring spermatogenic cells. The rat orthologue of human pd1 was further cloned and, according to the Gene Nomenclature Committee, was renamed spata2 (spermatogenesis-associated protein 2) gene on the basis of its FSH-dependent up-regulation and developmental expression. The analysis of the human and rat cDNA sequences disclosed an open reading frame for a protein of 520 and 511 amino acids respectively, with an overall identity of 85%. Subsequently, a zebrafish orthologue of the human spata2 gene was identified. The consensus open reading frame (1650 bp) encodes a polypeptide of 550 amino acids, which shares 37% identity with the human spata2. By means of whole-mount in situ hybridisation it has been shown that spata2 transcripts are maternally derived and become strongly localised in the central nervous system at early developmental stages. At the same time, RT-PCR analysis demonstrated that several adult zebrafish tissues expressed high level of spata2 mRNA providing evidence that this gene may have a broader function than previously described. More recently, novel findings have highlighted a potential role of spata2 during pancreatic development and beta-cell proliferation. In this review we will discuss spata2 gene expression and regulation as well as focus on novel evidence, which suggests a role for this protein in pancreatic beta-cell function
Glycosaminoglycans, proteoglycans and sulodexide and the endothelium: biological roles and pharmacological effects.
No full textExpression of LGI1 Impairs Proliferation and Survival of HeLa Cells
Full text linkThe LGI1 gene was suggested to function as tumor suppressor for its ability to reduce malignant features of glioblastoma cells. In support to this proposal were the findings that overexpression of LGI1 in neuroblastoma cells inhibited proliferation and induced apoptosis. In this study we performed stable LGI1 expression in HeLa cells to examine whether the noxious effect of LGI1 might be extended to cancer cells of diverse origin. HeLa cell clones stably expressing LGI1 exhibited a significant impairment of proliferation and a consistent increase of cell death when compared with control cells lacking expression of LGI1. Expression of LGI1 increased the activity of apoptosis effectors caspase-3/7; furthermore it downregulated the antiapoptotic BCL2 gene and upregulated the proapoptotic BAX gene expression, suggesting that the cause of HeLa cells death might be an increased susceptibility to apoptosis induced by LGI1. The results suggested that LGI1 is capable to restrain growth and survival of adenocarcinoma cells such as HeLa
Elevated serum xylitol levels and cardiovascular risk: an active component or an innocent bystander?
Full text linkHeparanase: A Multitasking Protein Involved in Extracellular Matrix (ECM) Remodeling and Intracellular Events
Full text linkHeparanase (HPSE) has been defined as a multitasking protein that exhibits a peculiar enzymatic activity towards HS chains but which simultaneously performs other non-enzymatic functions. Through its enzymatic activity, HPSE catalyzes the cutting of the side chains of heparan sulfate (HS) proteoglycans, thus contributing to the remodeling of the extracellular matrix and of the basal membranes. Furthermore, thanks to this activity, HPSE also promotes the release and diffusion of various HS-linked molecules like growth factors, cytokines and enzymes. In addition to being an enzyme, HPSE has been shown to possess the ability to trigger different signaling pathways by interacting with transmembrane proteins. In normal tissue and in physiological conditions, HPSE exhibits only low levels of expression restricted only to keratinocytes, trophoblast, platelets and mast cells and leukocytes. On the contrary, in pathological conditions, such as in tumor progression and metastasis, inflammation and fibrosis, it is overexpressed. With this brief review, we intend to provide an update on the current knowledge about the different role of HPSE protein exerted by its enzymatic and non-enzymatic activity
- …
