Littérature scientifique sur le sujet « Staminali renali »

Lo sviluppo di sempre nuove conoscenze sulle cellule staminali e il contemporaneo progresso delle Techniche basate sull'ingegneria tissutale permettono, ad oggi, di ampliare gli orizzonti della medicina rigenerativa e le sue applicazioni in ambito terapeutico. In questo articolo sono riportate le Techniche e le possibilità di base inerenti la creazione de novo di tessuti ed organi in vitro, evidenziandone sia i benefici, sia le problematiche di maggior rilievo che rendono tali approcci ancora poco applicabili in terapia, come la questione correlata al rigetto di natura immunitaria. Unendo la tematica del rigetto autoimmune e dell'ingegneria tissutale sarà quindi introdotta una possibile alternativa per la creazione di strutture renali totalmente create in vitro, ma anche totalmente funzionali.

Nella malattia renale cronica, il rischio di fallimento d’organo dipende dal grado di severità della proteinuria, la quale è determinata dal numero di podociti persi e dalla conseguente fibrosi glomerulare. Esistono, tuttavia, numerose evidenze cliniche e sperimentali che suggeriscono la possibilità di remissione della malattia renale e, in alcuni casi, persino di regressione del danno, quando ancora l’istologia dell’organo non risulti totalmente compromessa. Tali risultati sono ottenuti in particolare mediante l’impiego di terapie con effetti anti-proteinurici. Nuove evidenze sperimentali suggeriscono perché il blocco della perdita di proteine urinarie permetta la remissione della malattia renale cronica. In un recente articolo di Peired et al., è, per la prima volta, dimostrato come l’albuminuria blocchi il processo rigenerativo a causa del sequestro attuato dall’albumina ai danni della vitamina A, noto agente differenziativo per popolazioni di progenitori staminali presenti in vari organi. La conseguente perdita della vitamina A, complessata all’albumina, con le urine, impedisce, quindi, l’attivazione dei progenitori renali residenti nella capsula di Bowman, bloccando sul nascere la risposta rigenerativa e permettendo la progressione della malattia renale cronica.

Abstract Background: MODY 5 is an infrequently reported form of monogenic diabetes attributed to deletions of chromosome 17q12 with impaired expression of HNF1β (Hepatocyte Nuclear Factor 1 Beta). We report a patient who presented with hyperglycemia and hypomagnesemia with an eventual diagnosis of MODY5 that responded to treatment with the SGLT2 inhibitor (SGLT2i), canagliflozin. Clinical case: A 64-year-old male diagnosed with diabetes at age 60 and treated with glimepiride and metformin (HbA1c 6.1–6.2% without hypoglycemia) presented to establish care. He had a prior history of hypermagnesuric hypomagnesemia (serum Magnesium (Mg) 1.4 ± 0.3 mg/dl with Fractional Excretion of Mg (FEMg) 32 ± 3% [values >3–4% in the setting of normal renal function and hypomagnesemia indicates renal Mg loss]) that developed while taking Triamterene- HCTZ for hypertension, prompting discontinuation. Treatment with oral and intravenous (IV) Magnesium along with Amiloride failed to normalize serum Mg levels. Genetic evaluation revealed 17q12 deletion consistent with a diagnosis of MODY 5. Referral to nephrology resulted in discontinuation of glimepiride and addition of canagliflozin 100 mg titrated to 300 mg daily. Mg levels normalized (serum Mg level 1.9 ± 0.1 mg/dL) within 8 weeks of canagliflozin therapy, allowing discontinuation of IV Mg and patient reported improvement in physical stamina and quality of life. At his 1 year follow up visit, his serum Mg remains stable at 1.8mg/dl with a FEMg of 22 ± 2.5%. His current therapy includes Metformin 2 gm, Canagliflozin 300 mg, Amiloride 10 mg tid, and oral Mg. Discussion: Prior to introduction of the SGLT2i, MODY 5 patients required oral and IV Mg repletion in combination with amiloride to achieve near normal Mg levels. Several clinical trials with SGLT2i demonstrated dose and agent dependent improvement in Mg levels in patients with type 2 diabetes. Proposed explanations include reductions in eGFR, changes in intraluminal electrical potential, and activation of renin angiotensin aldosterone system. Mg replacement results not only in symptomatic improvement, but has also been demonstrated to reduce risk of stroke and all-cause mortality. In summary, SGLT2 inhibitors in patients with MODY 5 can be effective in restoring normal Mg levels, improving quality of life, and reducing all cause mortality.

8002 Background: High-risk (HR) cytogenetic is associated with poor outcome in transplant eligible (TE) newly diagnosed myeloma multiple myeloma (NDMM). The triplet combination carfilzomib lenalidomide and dexamethasone (KRD) plus transplantation demonstrated high efficacy with favorable safety profile in TE-NDMM patients (FORTE). The addition of daratumumab (Dara) to frontline therapy also improved response rate and progression free-survival in TE-NDMM patients (CASSIOPEIA, GRIFFIN). Double transplant also improved outcome of HR TE NDMM patients (EMN02, STAMINA). The phase 2 trial 2018-04 from the Intergroupe Francophone du Myelome (IFM) is evaluating an intensive strategy with Dara-KRD induction and consolidation plus double transplant in HR TE NDMM (NCT03606577). Methods: HR MM was defined by the presence of del17p, t(4;14) and/or t(14;16). Stategy includes Dara-KRD induction (6 cycles), autologous stem cell transplantation (ASCT), Dara-KRD consolidation (4 cycles), second ASCT, Dara-lenalidomide maintenance. The primary endpoint was the feasibility of this intensive strategy. Here, we report efficacy and safety analysis of Dara-KRD induction. Results: Fifty patients with previously untreated NDMM were included from july 2019 to march 2021 in 11 IFM centers Median age was 57 (range 38 -65). ISS stage 3 was present in 12 (24%) patients. Based on inclusion criteria, all patients had HR cytogenetic, including 17p deletion (n = 20, 40%), t(4;14) (n = 26, 52%) or t(14;16) (n = 10,20%). Forty-six patients completed Dara-KRD induction. Two patients discontinued treatment due to severe adverse event (COVID-19 infection, n = 1 ; drug-induced hepatitis, n = 1) and 2 patients discontinued treatment due to disease progression. Grade 3-4 treatment related adverse event ( > 5% of patients) were neutropenia (38%), anemia (14%), thrombocytopenia (8%), infection (6%), renal insufficiency (6%) and deep-vein thrombosis (6%). Two patients (6%) experienced stem-cell collection failure. Overall response rate was 96%, including 92 % > very good partial response. Among 37 (/46) evaluable patients post induction, Minimal Residual Disease negativity rate (NGS, 10-5) was 62%. Conclusions: Dara-KRD as induction prior ASCT is safe and allows deep responses in TE NDMM patients with high-risk cytogenetic profile. IFM 2018-04 study is ongoing and longer follow-up is needed to evaluate safety and efficacy of the overall strategy with Dara-KRD induction and consolidation plus double transplant in this subset of HR patients. Clinical trial information: NCT03606577.

8002 Background: High-risk (HR) cytogenetic is associated with poor outcome in transplant eligible (TE) newly diagnosed myeloma multiple myeloma (NDMM). The triplet combination carfilzomib lenalidomide and dexamethasone (KRD) plus transplantation demonstrated high efficacy with favorable safety profile in TE-NDMM patients (FORTE). The addition of daratumumab (Dara) to frontline therapy also improved response rate and progression free-survival in TE-NDMM patients (CASSIOPEIA, GRIFFIN). Double transplant also improved outcome of HR TE NDMM patients (EMN02, STAMINA). The phase 2 trial 2018-04 from the Intergroupe Francophone du Myelome (IFM) is evaluating an intensive strategy with Dara-KRD induction and consolidation plus double transplant in HR TE NDMM (NCT03606577). Methods: HR MM was defined by the presence of del17p, t(4;14) and/or t(14;16). Stategy includes Dara-KRD induction (6 cycles), autologous stem cell transplantation (ASCT), Dara-KRD consolidation (4 cycles), second ASCT, Dara-lenalidomide maintenance. The primary endpoint was the feasibility of this intensive strategy. Here, we report efficacy and safety analysis of Dara-KRD induction. Results: Fifty patients with previously untreated NDMM were included from july 2019 to march 2021 in 11 IFM centers Median age was 57 (range 38 -65). ISS stage 3 was present in 12 (24%) patients. Based on inclusion criteria, all patients had HR cytogenetic, including 17p deletion (n = 20, 40%), t(4;14) (n = 26, 52%) or t(14;16) (n = 10,20%). Forty-six patients completed Dara-KRD induction. Two patients discontinued treatment due to severe adverse event (COVID-19 infection, n = 1 ; drug-induced hepatitis, n = 1) and 2 patients discontinued treatment due to disease progression. Grade 3-4 treatment related adverse event ( > 5% of patients) were neutropenia (38%), anemia (14%), thrombocytopenia (8%), infection (6%), renal insufficiency (6%) and deep-vein thrombosis (6%). Two patients (6%) experienced stem-cell collection failure. Overall response rate was 96%, including 92 % > very good partial response. Among 37 (/46) evaluable patients post induction, Minimal Residual Disease negativity rate (NGS, 10-5) was 62%. Conclusions: Dara-KRD as induction prior ASCT is safe and allows deep responses in TE NDMM patients with high-risk cytogenetic profile. IFM 2018-04 study is ongoing and longer follow-up is needed to evaluate safety and efficacy of the overall strategy with Dara-KRD induction and consolidation plus double transplant in this subset of HR patients. Clinical trial information: NCT03606577.

La fragilità è una sindrome geriatrica definita da un progressivo declino età-correlato di diverse facoltà fisiologiche, che si traduce in una ridotta funzionalità d’organo e in un’aumentata vulnerabilità in condizioni di stress. Fried e coll. propongono una definizione operativa delle fragilità. La letteratura riporta un’alta prevalenza di fragilità in individui affetti da insufficienza renale cronica. Tra questi soggetti, il rischio di sviluppare fragilità è aumentato di due/tre volte rispetto ai soggetti sani. Ad oggi, la definizione e la valutazione della fragilità in questi pazienti è ancora controversa. Tuttavia quello che si sa è che l’invecchiamento è un processo strettamente correlato alla ridotta capacità delle cellule staminali di auto-rinnovarsi e differenziarsi. Quest’alterazione delle funzioni delle cellule staminali può svolgere un ruolo chiave nella fisiopatologia delle malattie associate all’invecchiamento, inclusa la disfunzione renale. Il nostro gruppo ha identificato e isolato le cellule staminali renali adulte a partire da colture clonali di nefrosfere umane. All’interno di queste sono presenti cellule a diversa differenziazione e maturazione, tra queste anche le staminali identificate come PKHhigh/CD133+/CD24-, multipotenti e in grado di ripopolare scaffold renali decellularizzati. Date queste premesse l'obiettivo del progetto è valutare gli effetti di condizioni biologiche legate alla fragilità sul comportamento delle cellule staminali renali adulte umane e capire se tali condizioni sono in grado di esaurire il pool di cellule staminali e alterarne la funzione. In primo luogo abbiamo arruolato soggetti fragili, pre-fragili e non-fragili e giovani sani come controllo e raccolto i rispettivi plasmi e cellule mononucleate dal sangue periferico (PBMC). Sia nelle PBMC che nelle cellule staminali/progenitrici ematopoietiche circolanti abbiamo valutato il danno al DNA osservando una percentuale di cellule positive al danno statisticamente più alta nei pazienti fragili rispetto agli altri gruppi. Per valutare il reale effetto delle condizioni biologiche legate alla fragilità sulle proprietà delle RSC, le colture di NS, ottenute da nefrectomie, sono state trattate con il plasma dei soggetti arruolati. Abbiamo valutato dapprima le capacità di autorinnovamento delle cellule trattate e osserviamo una significativa diminuzione dell'efficienza di formazione della sfera, indice di autorinnovamento, nei soggetti fragili rispetto ai non fragili e ai giovani. Successivamente, abbiamo valutato il danno al DNA, i ROS intracellulari, la proliferazione e la vitalità nelle cellule staminali/progenitrici renali ottenute dopo la dissociazione delle NS. Non sono state evidenziate differenze nella vitalità e nella proliferazione cellulare tra i gruppi, mentre il danno al DNA e i ROS intracellulari sono aumentati nelle cellule delle NS trattate con plasma di anziani fragili rispetto a quelle trattate con gli altri plasmi. Ciò potrebbe indicare che la diminuzione della capacità di autorinnovamento nelle cellule trattate con il plasma di pazienti fragili e un aumento del danno al DNA e dei ROS intracellulari non sono correlati con la morte o la proliferazione cellulare, ma con un'elevata presenza di mediatori infiammatori e ROS nel plasma dei pazienti fragili. Per confermare questi dati abbiamo analizzato lo stress ossidativo e il profilo di 40 citochine infiammatorie nel plasma dei soggetti arruolati. Si ha un aumento dello stress ossidativo nel plasma dei soggetti fragili rispetto agli altri gruppi, così come un’aumentata presenza o l’esclusività di alcune citochine infiammatorie. Questi dati preliminari suggeriscono che esiste una combinazione di stress ossidativo e citochine pro-infiammatorie nel plasma di pazienti fragili che contribuiscono ad aumentare il danno al DNA e i ROS intracellulari alterando conseguentemente le caratteristiche di staminalità delle cellule delle NS.
Frailty is a geriatric syndrome that can be defined as an age-related progressive impairment of multiple physiological systems, resulting in a significantly reduced capacity to compensate for external stressors. Fried and colleagues proposed a phenotype characterization of frailty through five physical criteria, so this can be possible only after the onset of clinical manifestations without the possibility of a precocious diagnosis. Several studies report a high prevalence of frailty in both old and young individuals with kidney dysfunction, and this further increases with advancing age and progressive decline of renal function. Elderly individuals with chronic kidney disease (CKD) are two to three times more likely to be frail than those with normal renal function. However, the relationship between CKD and frailty is still unclear. The aging process can have adverse effects on stem cells; their self-renewal ability declines and their differentiation potential into the various cell types is altered. Aging-induced exhaustion and deterioration of stem cell pool and functions may play a key role in the pathophysiology of aging-associated diseases, including kidney dysfunction. Our group isolated a pure population of multipotent renal stem-like cells by a functional approach, taking advantage from the ability of renal stem cells (RSC) to grow as nephrospheres (NS). Investigating the expression of renal progenitor markers described in literature, our group identified in NS a homogeneous PKHhigh/CD133+/CD24- cell population displaying in vitro stem-cell properties, able to repopulate human decellularized renal scaffold and exhibiting multipotency. In this scenario, we tested whether in the organism of elderly and frail people there are biological conditions able to alter RSC behavior, justifying the high prevalence of chronic kidney dysfunction in the frail status and its severity. First, we recruited frail, pre-frail and non-frail subjects, and young subjects as controls and we obtained whole blood that was separated into plasma and PBMC. We studied DNA damage in both PBMC and circulating hematopoietic progenitor/stem cells (cHPSC) and we observed a statistically higher percentage of cells positive for DNA damage in frail patients compared to all the other groups. To assess the real effect of biological conditions related to frailty on adult RSC properties, NS cultures, obtained from nephrectomies, were treated with 10% plasma of enrolled frail and non-frail subjects and healthy young. We first evaluated the self-renewal abilities of treated cells and we observe a significant decrease in sphere forming efficiency, indication of self-renewal, in frail subjects compared to both non-frail and young people. Subsequently, we evaluated DNA damage, intracellular ROS, proliferation and viability in renal stem/progenitor cells obtained after NS dissociation after plasma treatment. We find no differences in viability and proliferation between groups, while DNA damage and intracellular ROS increased in NS cells treated with plasma of frail seniors compared to those treated with the other plasmas. This might indicate that the decrease of self-renewal ability in cell treated with plasma of frail patients and an increase of DNA damage and intracellular ROS are not correlated to cell death or proliferation, but with a high presence of inflammatory mediators and ROS in the plasma of frail patients. To confirm these data we analyzed the oxidative stress and the profile of 40 inflammatory cytokines on plasma of enrolled subjects. We observed an increase in oxidative stress and osome inflammatory cytokines in frail plasma compared to other plasmas. These preliminary data suggested that there is a combination of oxidative stress and pro-inflammatory cytokines in plasma of frail patients that contribute to increase DNA damage and intracellular ROS and consequently alter stem characteristics of NS cells.

In recent years, numerous cancers have been described as having a "cancer stem cell" (CSC) population also known as "cancer initiating cells". CSCs refer to a subset of tumour cells that has the ability to self-renew and generate the diverse cells that comprise the tumor. Their name derives from their "stem-like" properties and ability to continually sustain tumorigenesis. CSCs have the same properties that define a normal tissue adult stem cell, even if they are aberrant: self-renewal and differentiation. Renal cell carcinoma (RCC) accounts for about 3% of adult cancers and is among the 10 most common malignancies in Europe. RCC has several morphological subtypes and clear cell RCC accounts for ~80% of cases. RCC has a late diagnosis and is therapy resistant. In renal pathologies there are situation in which the presence and function of adult renal stem cells may have clinical relevance. In the last years the existence of different sources of renal stem cells has been proposed even if the phenotype of a resident stem cell of the kidney has not been exhaustively described. Even in RCC the definition of a kidney cancer stem cell may have a role for better understanding renal cell carcinoma biology. A number of approaches based on the exploitation of renal stem cell markers have allowed the prospective isolation of human renal stem cells, even if the relative promiscuity of these markers limits their usefulness when highly purified stem cells are needed. So we decided to use a functional approach for the isolation of normal and cancer stem cells of the kidney by culturing the cells in suspension, non-adherent conditions, at low density with specific growth factors. In these conditions only a little percentage of cells survives growing as spherical aggregates in suspension. We called them "nephrospheres". Using fluorescent lipophilic dyes PKH, we demonstrated the clonal origin of the spheres and the presence of a heterogeneous population inside the spheres. In fact the dye dilutes in active replicating cells while is retained in quiescent cells; we can observe in normal and cancer nephrospheres some most fluorescent cells, the quiescent stem cells, and some less fluorescent or not fluorescent cells, that are the active replicating progenitors. We performed a characterization of the nephrospheres by immunofluorescence after cytospin or FACS evaluating the expression of some epithelial and stem cell markers. By Real Time PCR we found that some genes related with stemness or involved in the maintenance of pluripotency are overexpressed in normal and cancer nephrospheres if compared with the corresponding differentiated primary cell cultures. We then evaluated the differentiative abilities of the cells derived from normal nephrospheres by culturing the cells in specific media and semisolid substrates; the cells are able to differentiate into epithelial and neuronal-like phenotype and to form tubular/glomerular-like tridimensional structures. We the isolated the stem cell population form normal nephrospheres on the basis of the PKH fluorescence. We identified 3 PKH populations: PKHhigh population, with a high level of fluorescence, PKHlow population, with a low level of fluorescence, and PKHneg population, negative for PKH. The populations were separated with cell sorting and cultivated to form spheres. Only PKHhigh cells were able to generate new spheres, demostrating that the PKHhigh cells represent the stem cell population inside the nephrospheres. Normal and cancer PKHhigh cells will be deeply characterized in the future.

While amphetamine and especially methamphetamine (speed) is being misused by all social classes in order to increase stamina, intellectual expansion, endurance, and euphoria, the drug 3,4-methylenedioxy-N-methylamphetamine (MDMA) (ecstasy) is preferentially abused by the younger generation for the feeling of empathy, the touching within, and enhancement of the senses. Acute intoxication differs in regard to their effects on the person. The predominant sympathetic overstimulation after methamphetamine results in cardiovascular and CNS hyperactivity accompanied by agitation and seizures, while tachycardia is a prodrome of fibrillation. The excess hypertonia often leads into myocardial infarction and may even induce cerebral haemorrhage. MDMA intoxication often seen in the emergency department is predominantly characterized by hyperthermia, the most important condition to treat, followed by rhabdomyolysis and acute renal failure. Since there is no specific antidote available, in both cases therapy consists of treatment until the acute effects are gone.