Impact of chronic exposure to bevacizumab on EpCAM-based detection of circulating tumor cells

BACKGROUND: Circulating tumor cells (CTCs) are often undetected through the immunomagnetic epithelial cell adhesion molecule (EpCAM)-based CellSearch(®) System in breast and colorectal cancer (CRC) patients treated with bevacizumab (BEV), where low CTC numbers have been reported even in patients with evidence of progression of disease. To date, the reasons for this discrepancy have not been clarified. This study was carried out to investigate the molecular and phenotypic changes in CRC cells after chronic exposure to BEV in vitro. METHODS: The human CRC cell line WiDr was exposed to a clinically relevant dose of BEV for 3 months in vitro. The expression of epithelial and mesenchymal markers and EpCAM isoforms was determined by western blotting and immunofluorescence. To evaluate the impact of EpCAM variant isoforms expression on CTC enumeration by CellSearch(®), untreated and treated colon cancer cells were spiked into 7.5 mL of blood from a healthy donor and enumerated by CellSearch(®). RESULTS: Chronic exposure of CRC cell line to BEV induced decreased expression of EpCAM 40 kDa isoform and increased expression EpCAM 42 kDa isoform, together with a decreased expression of cytokeratins (CK), while no evidence of epithelial to mesenchymal transition (EMT) in treated cells was observed. The recovery rate of cells through CellSearch(®) was gradually reduced in course of treatment with BEV, being 84%, 70% and 40% at 1, 2 and 3 months, respectively. CONCLUSIONS: We hypothesize that BEV may prevent CellSearch(®) from capturing CTCs through altering EpCAM isoforms

Spin–orbit coupling controlling the superconducting dome of artificial superlattices of quantum wells

While it is known that a resonant amplification of Tc in two-gap superconductors can be driven by using the Fano-Feshbach resonance tuning the chemical potential near a Lifshitz transition, little is known on tuning the Tc resonance by cooperative interplay of the Rashba spin-orbit coupling (RSOC) joint with phonon mediated (e-ph) pairing at selected k-space spots. Here we present first-principles quantum calculation of superconductivity in an artificial heterostructure of metallic quantum wells with 3 nm period where quantum size effects give two-gap superconductivity with RSOC controlled by the internal electric field at the interface between the nanoscale metallic layers intercalated by insulating spacer layers. The key results of this work show that fundamental quantum mechanics effects including RSCO at the nanoscale (Mazziotti et al Phys. Rev. B, 103, 024523, 2021) provide key tools in applied physics for quantitative material design of unconventional high temperature superconductors at ambient pressure. We discuss the superconducting domes where Tc is a function of either the Lifshitz parameter (?) measuring the distance from the topological Lifshitz transition for the appearing of a new small Fermi surface due to quantum size effects with finite spin-orbit coupling and the variable e-ph coupling g in the appearing second Fermi surface linked with the softening of the phonon energy cut off.Comment: 13 pages, 8 figure

Born to sing! Song development in a singing primate

In animal vocal communication, the development of adult-like vocalization is fundamental to interact appropriately with conspecifics. However, the factors that guide ontogenetic changes in the acoustic features remain poorly understood. In contrast with a historical view of nonhuman primate vocal production as substantially innate, recent research suggests that inheritance and physiological modification can only explain some of the developmental changes in call structure during growth. A particular case of acoustic communication is the indris’ singing behavior, a peculiar case among Strepsirrhine primates. Thanks to a decade of intense data collection, this work provides the first long-term quantitative analysis on song development in a singing primate. To understand the ontogeny of such a complex vocal output, we investigated juvenile and sub-adult indris’ vocal behavior, and we found that young individuals started participating in the chorus years earlier than previously reported. Our results indicated that spectro-temporal song parameters underwent essential changes during growth. In particular, the age and sex of the emitter influenced the indris’ vocal activity. We found that frequency parameters showed consistent changes across the sexes, but the temporal features showed different developmental trajectories for males and females. Given the low level of morphological sexual dimorphism and the marked differences in vocal behavior, we hypothesize that factors like social influences and auditory feedback may affect songs’ features, resulting in high vocal flexibility in juvenile indris. This trait may be pivotal in a species that engages in choruses with rapid vocal turn-taking

Expression Of Mir-34a In T-cells Infected By Human T-lymphotropic Virus 1

Human T-lymphotropic virus 1 (HTLV-1) immortalizes T-cells and is the causative agent of adult T-cell leukemia/lymphoma (ATLL). HTLV-1 replication and transformation are governed by multiple interactions between viral regulatory proteins and host cell factors that remain to be fully elucidated. The present study investigated the impact of HTLV-1 infection on the expression of miR-34a, a microRNA whose expression is downregulated in many types of cancer. Results of RT-PCR assays showed that five out of six HTLV-1-positive cell lines expressed higher levels of miR-34a compared to normal PBMC or purified CD4+ T-cells. ATLL cell line ED, which did not express miR-34a, showed methylation of the miR-34a promoter. Newly infected PBMC and samples from 10 ATLL patients also showed a prominent increase in miR-34a expression compared to PBMC controls. The primary miR-34a transcript expressed in infected cell line C91PL contained binding motifs for NF-kappa B and p53. Pharmacological inhibition of NF-kappa B with Bay 11-7082 indicated that this pathway contributes to sustain miR-34a levels in infected cells. Treatment of infected cell lines with the p53 activator nutlin-3a resulted in a further increase in miR-34a levels, thus confirming it as a transcriptional target of p53. Nutlin-3a-treated cells showed downregulation of known miR-34a targets including the deacetylase SIRT1, which was accompanied by increased acetylation of p53, a substrate of SIRT1. Transfection of C91PL cells with a miR-34a mimic also led to downregulation of mRNA targets including SIRT1 as well as the pro-apoptotic factor BAX. Unlike nutlin-3a, the miR-34a mimic did not cause cell cycle arrest or reduce cell viability. On the other hand, sequestration of miR-34a with a sponge construct resulted in an increase in death of C91PL cells. These findings provide evidence for a functional role for miR-34a in fine-tuning the expression of target genes that influence the turnover of HTLV-1-infected cells

Neonatal isolated coronary artery dilatation: to treat (and how) or not to treat, that’s the question!

We describe the clinical presentation of a newborn, who received diagnosis of diffuse right coronary artery dilatation and a high origin of the same vessel, in the absence of intrauterine hypoxia or other identifiable causes of coronary artery ectasia. The relevance of the case is in the complex aetiology of congenital heart malformation. In addition, we also highlight the difficult therapeutic management of these patients, in lack of guidelines for newborn population

Conceptual Design of a Soft X‐ray SASE‐FEL Source

FELs based on SASE are believed to be powerful tools to explore the frontiers of basic sciences, from physics to chemistry to biology. Intense R&D programs have started in the USA and Europe in order to understand the SASE physics and to prove the feasibility of these sources. The allocation of considerable resources in the Italian National Research Plan (PNR) brought about the formation of a CNR‐ENEA‐INFN‐University of Roma "Tor Vergata" study group. A conceptual design study has been developed and possible schemes for linac sources have been investigated, bringing to the SPARX proposal. We report in this paper the results of a preliminary start to end simulation concerning one option we are considering based on an S‐band normal conducting linac with high brightness photoinjector integrated in a RF compressor

Role of miR-34a in HTLV-1-infected T-cells

Human T-cell leukaemia virus type 1 (HTLV-1) is the etiological agent of an aggressive neoplasm of CD4+ T-cells termed adult T-cell leukaemia/lymphoma (ATLL) and a neurodegenerative disease termed tropical spastic paraparesis/HTLV-associated myelopathy (TSP/HAM). To better understand the pathways controlling HTLV-1 infection, persistence and transformation, our laboratory is investigating the interplay between the virus and the cellular microRNA (miRNA) network. This study was focused on miR-34a, which is known as a tumor suppressor in other contexts, but is highly expressed in HTLV-1 infected cell lines, newly infected PBMCs and ATLL samples. Further studies of HTLV-1-infected cell lines C91PL and MT-2 treated with the NF-κB inhibitor Bay 11-7082 indicated that the NF-κB pathway contributes to sustain miR-34a expression in this cellular context. Identification of the primary miR-34a precursor (pri-miRNA) produced in C91PL cells through the 5’RACE technique revealed that the main pri-miR-34a species present in this cell line corresponded to a two-exon transcript previously identified in cells of different lineage. The presence of an NF-κB binding site near the transcription start site of the primiR-34a provided further evidence that miR-34a expression is driven by this pathway in HTLV-1-infected cells. Treatment of C91PL and MT-2 cells with the MDM2 inhibitor Nutlin-3a resulted in stabilization of p53, a further increase in the levels of miR-34a and downregulation of several of its targets, including SIRT1, a deacetylase whose substrates include p53, the inhibitor of apoptosis protein (IAP) BIRC5, which codes for Survivin, and the transcription factor E2F3, which is involved in controlling cell cycle. Interestingly, although Nutlin-3a induced G1 arrest in both cell lines, MT-2 cells entered late apoptosis, while C91PL cells showed signs of senescence. The last part of this project was aimed at investigating the effects of Nutlin-3a on viral gene expression and directly assessing the impact of miR-34a in C91PL cells by electroporating a miR-34a mimic. We observed that Nultin-3a treatment of C91PL cells increased levels of the majority of the alternatively spliced HTLV-1 transcripts. Although the introduction of the miR-34a mimic into C91PL cells resulted in a reduction in the expression of key targets of the miRNA, it did not induce cell cycle arrest, death, senescence, or alterations in viral gene expression, suggesting that other factors come into play in the response to Nutlin-3a treatment.Il virus T-linfotropico umano di tipo 1 (HTLV-1, Human T-cell leukaemia virus type 1) è l’agente eziologico della leucemia/linfoma a cellule T dell’adulto (ATLL, adult T-cell leukaemia/lymphoma) e della paraparesi spastica tropicale/mielopatia associata ad HTLV (TSP/HAM, Tropical spastic paraparesis/HTLV-associated myelopathy), una patologia degenerativa del sistema nervoso centrale. Al fine di comprendere i meccanismi volti a regolare l’infezione da HTLV-1, la sua persistenza e la capacità di indurre trasformazione cellulare, il nostro laboratorio ha investigato le relazioni tra virus e microRNA (miRNA). In particolar modo, il presente lavoro si è incentrato sull’analisi dell’espressione di miR-34a, noto oncosoppressore in altri contesti tumorali, il quale risulta espresso ad alti livelli in campioni derivanti da pazienti ATLL, linee cellulari stabilmente infettate da HTLV-1 e PBMC infettati de novo. Uno studio approfondito effettuato su linee cellulari infettate da HTLV-1, C91PL e MT-2, trattate con un inibitore di NF-κB, Bay 11-7082, ha rivelato come il pathway di NF-κB sia coinvolto nell’aumento di espressione di miR-34a in questo contesto cellulare. Inoltre, la determinazione del precursore di miR-34a (pri-miRNA) effettuata in cellule C91PL mediante la tecnica denominata 5’RACE, ha permesso di identificare un trascritto costituito da due esoni, precedentemente descritto in una diversa linea. Inoltre, trattamenti delle linee cellulari C91PL e MT-2 con un inibitore di MDM2, Nutlin-3a, hanno portato alla stabilizzazione di p53, ad un incremento dei livelli di miR-34a e alla riduzione dei suoi mRNA bersaglio, tra cui SIRT1 (una deacetilasi tra i cui substrati è annoverato anche p53), BIRC5 (codificante Survivin, una proteina inibitrice dell’apoptosi) ed E2F3 (un fattore di trascrizione implicato nel controllo del ciclo cellulare). È interessante notare come Nutlin-3a, nonostante causi l’arresto del ciclo cellulare in fase G1 in entrambe le linee, induca un’apoptosi tardiva in cellule MT-2, e porti invece le cellule C91PL ad uno stato di senescenza. Infine, il progetto si è rivolto a investigare gli effetti di Nutlin-3a sull’espressione genica virale, nonchè l’impatto diretto di miR-34a su cellule C91PL mediante elettroporazione con un “miR-34a mimic”. I nostri esperimenti ci hanno permesso di osservare come cellule C91PL trattate con Nutlin-3a presentino livelli aumentati dei trascritti di HTLV-1. Tuttavia, l’introduzione di miR-34a-mimic in cellule C91PL, pur confermando una riduzione nei livelli di espressione dei bersagli di miR-34a, non ha mostrato induzione di arresto del ciclo cellulare, morte o senescenza, così come non ha alterato l’espressione dei geni virali, suggerendo che anche altri fattori possano agire in risposta ai trattamenti con Nutlin-3a

Role of miR-34a in HTLV-1-infected T-cells

Human T-cell leukaemia virus type 1 (HTLV-1) is the etiological agent of an aggressive neoplasm of CD4+ T-cells termed adult T-cell leukaemia/lymphoma (ATLL) and a neurodegenerative disease termed tropical spastic paraparesis/HTLV-associated myelopathy (TSP/HAM). To better understand the pathways controlling HTLV-1 infection, persistence and transformation, our laboratory is investigating the interplay between the virus and the cellular microRNA (miRNA) network. This study was focused on miR-34a, which is known as a tumor suppressor in other contexts, but is highly expressed in HTLV-1 infected cell lines, newly infected PBMCs and ATLL samples. Further studies of HTLV-1-infected cell lines C91PL and MT-2 treated with the NF-κB inhibitor Bay 11-7082 indicated that the NF-κB pathway contributes to sustain miR-34a expression in this cellular context. Identification of the primary miR-34a precursor (pri-miRNA) produced in C91PL cells through the 5’RACE technique revealed that the main pri-miR-34a species present in this cell line corresponded to a two-exon transcript previously identified in cells of different lineage. The presence of an NF-κB binding site near the transcription start site of the primiR-34a provided further evidence that miR-34a expression is driven by this pathway in HTLV-1-infected cells. Treatment of C91PL and MT-2 cells with the MDM2 inhibitor Nutlin-3a resulted in stabilization of p53, a further increase in the levels of miR-34a and downregulation of several of its targets, including SIRT1, a deacetylase whose substrates include p53, the inhibitor of apoptosis protein (IAP) BIRC5, which codes for Survivin, and the transcription factor E2F3, which is involved in controlling cell cycle. Interestingly, although Nutlin-3a induced G1 arrest in both cell lines, MT-2 cells entered late apoptosis, while C91PL cells showed signs of senescence. The last part of this project was aimed at investigating the effects of Nutlin-3a on viral gene expression and directly assessing the impact of miR-34a in C91PL cells by electroporating a miR-34a mimic. We observed that Nultin-3a treatment of C91PL cells increased levels of the majority of the alternatively spliced HTLV-1 transcripts. Although the introduction of the miR-34a mimic into C91PL cells resulted in a reduction in the expression of key targets of the miRNA, it did not induce cell cycle arrest, death, senescence, or alterations in viral gene expression, suggesting that other factors come into play in the response to Nutlin-3a treatment

Antenne a schiera planari a bassa osservabilità radar

La tesi illustra il progetto di antenne(singole e ad array)a patch e a slot realizzate su superfici selettive in frequenza (FSS) che consentono di ridurre la Radar Cross Section rispetto al caso in cui si utilizzi un piano di massa perfettamente conduttore. Nella prima parte si analizzano le prestazioni di una singola antenna a patch e a slot sia su piano di massa che su FSS; successivamente vengono descritte le proprietà delle FSS in termini di campo riflesso e trasmesso e infine si analizza il comportamento di un array di slot e di patch su superficie selettiva e su piano di massa perfettamente conduttore. L'ultima parte della tesi mostra il confronto tra i risultati ottenuti per le diverse antenne progettate