Accuracy of five electronic foramen locators with different operating systems: na ex vivo study

Objective: The aim of this study was to evaluate, ex vivo, the precision of five electronic root canal length measurement devices (ERCLMDs) with different operating systems: the Root ZX, Mini Apex Locator, Propex II, iPex, and RomiApex A-15, and the possible influence of the positioning of the instrument tips short of the apical foramen. Material and Methods: Forty-two mandibular bicuspids had their real canal lengths (RL) previously determined. Electronic measurements were performed 1.0 mm short of the apical foramen (-1.0), followed by measurements at the apical foramen (0.0). The data resulting from the comparison of the ERCLMD measurements and the RL were evaluated by the Wilcoxon and Friedman tests at a significance level of 5%. Results: Considering the measurements performed at 0.0 and -1.0, the precision rates for the ERCLMDs were: 73.5% and 47.1% (Root ZX), 73.5% and 55.9% (Mini Apex Locator), 67.6% and 41.1% (Propex II), 61.7% and 44.1% (iPex), and 79.4% and 44.1% (RomiApex A-15), respectively, considering ±0.5 mm of tolerance. Regarding the mean discrepancies, no differences were observed at 0.0; however, in the measurements at -1.0, the iPex, a multi-frequency ERCLMD, had significantly more discrepant readings short of the apical foramen than the other devices, except for the Propex II, which had intermediate results. When the ERCLMDs measurements at -1.0 were compared with those at 0.0, the Propex II, iPex and RomiApex A-15 presented significantly higher discrepancies in their readings. Conclusions: Under the conditions of the present study, all the ERCLMDs provided acceptable measurements at the 0.0 position. However, at the -1.0 position, the ERCLMDs had a lower precision, with statistically significant differences for the Propex II, iPex, and RomiApex A-15.

Recondicionamento pulmonar ex vivo: uma nova era para o transplante pulmonar

O transplante pulmonar consolidou-se como a melhor opção terapêutica para diversas pneumopatias terminais. O baixo número de doadores viáveis ainda persiste como uma grande limitação ao aumento do número de transplantes de pulmão, causando alta mortalidade na lista de espera. Diferentemente do transplante de outros órgãos sólidos, a maior limitação do transplante pulmonar não é o número absoluto de doadores e sim a viabilidade desses órgãos, que é reduzida devido às agressões ao pulmão ocasionadas pela morte encefálica e aos cuidados na UTI. Diversas são as propostas para o aumento do número de doadores: intensificação das campanhas de doação, o uso de doadores com coração parado, transplante pulmonar lobar intervivos e maior flexibilidade dos critérios para aceitação de doadores de pulmão. Todavia, a proposta que atrai a atenção de diversos grupos de transplante pulmonar é a perfusão pulmonar ex vivo, principalmente pela perspectiva de recuperação de pulmões inicialmente descartados. Esse sistema consiste na reperfusão e ventilação do bloco pulmonar isolado em um circuito de circulação extracorpórea modificado. Devido aos bons resultados apresentados e à perspectiva de aumento no número de órgãos aptos a transplante, diversos grupos têm estudado a técnica. Pesquisadores na Suécia, Canadá, Áustria, Inglaterra, Espanha e Brasil já possuem experiência sólida com o método e introduziram algumas variações. O objetivo deste artigo foi revisar o desenvolvimento, o estado da arte e as perspectivas futuras do modelo ex vivo de perfusão e recondicionamento pulmonar.

Isotiocianati come potenziali farmaci antileucemici: identificazione in vitro ed ex vivo del profilo molecolare e cellulare

Il presente studio ha come obbiettivo lo sviluppo di composti di origine naturale come potenziali farmaci antitumorali, attraverso la definizione dei loro specifici target cellulari e molecolari su diversi modelli cellulari ad alta predittività. Gli isotiocianati, contenuti nei vegetali appartenenti alla famiglia delle Crucifereae, sono dotati di una comprovata capacità di inibire la formazione di tumori in modelli animali preventivamente trattati con cancerogeni. Questa attività è riconducibile principalmente alla modulazione degli enzimi coinvolti nell’attivazione/detossificazione di xenobiotici e ad effetti citostatici e citossici, osservati su numerose linee cellulari. Un isotiocianato particolarmente promettente è il sulforafane (SFN). La ricerca condotta durante il periodo di dottorato si è, quindi, focalizzata sull’isotiocianato SFN e in particolare sulla sua capacità di modulare specifici eventi cellulari e molecolari coinvolti nel processo di leucemogenesi. Inizialmente è stato indagato il potenziale citostatico e citotossico del SFN su una linea cellulare T linfoblastoide (cellule Jurkat), con particolare attenzione agli effetti sulla proliferazione cellulare, all’induzione di apoptosi/necrosi e all’analisi di alcuni dei meccanismi molecolari coinvolti negli effetti citostatici e citotossici dell’isotiocianato ( livelli proteici di p53, bax e bcl-2). Successivamente, poiché requisiti fondamentali di un antitumorale sono selettività d’azione e scarsa tossicità, è stato indagato il potenziale citostatico e citotossico dell’isotiocianato SFN sulla controparte non trasformata delle cellule leucemiche T linfoblastoidi, analizzando gli stessi eventi studiati su cellule tumorali e alcuni dei meccanismi molecolari coinvolti (livelli proteici di ciclina D2, ciclina D3, chinasi ciclina dipendente (CDK) 4 e CDK6 ). Il SFN si è dimostrato in grado di indurre apoptosi sulle cellule Jurkat e di inibirne la proliferazione, mediante un blocco in fase G2/M del ciclo cellulare e un incremento dei livelli di p53 e bax. Il SFN è in grado di indurre effetti citostatici e citotossici anche su linfociti T non trasformati. Tuttavia, le dosi necessarie per esibire tali effetti sono ben più elevate di quelle attive su cellule leucemiche. Una tappa importante nello sviluppo di un farmaco antitumorale è, la definizione, dove possibile, dei suoi effetti in un modello ex vivo, altamente predittivo di quella che sarà la risposta farmacologica in vivo. Sono stati quindi valutati gli effetti del SFN su colture primarie di blasti provenienti da pazienti affetti da diversi tipi di leucemia , sia mieloide che linfoblastica. Il SFN non sembra possedere alcuna attività su campioni da pazienti affetti da LLC, mentre un importante attività proapoptotica si registra nei campioni da pazienti affetti da LMA, dove l’effetto del SFN è sorprendentemente marcato anche su campioni da pazienti multiresistenti. L’attività dell’isotiocianato sui campioni da pazienti affetti da LLA è decisamente più marcata sul campione da paziente affetto da LLA a cellule B, mentre sul campione di Leucemia Acuta Bifenotipica l’effetto proapoptotico del SFN si registra dopo tempi di trattamento brevi piuttosto che dopo tempi di trattamento più lunghi. In conclusione, i risultati ottenuti evidenziano che il SFN possiede un’interessante attività antileucemica in vitro e, dato di particolare rilevanza, anche ex vivo.

Analisi clinica ed ex-vivo dei punti isometrici durante tecnica tightrope nella ricostruzione del legamento crociato craniale del cane

La rottura del Legamento Crociato Craniale (LCCr) rappresenta una delle patologie ortopediche di maggiore riscontro clinico nella specie canina. In seguito a rottura del LCCr si presenta un continuo slittamento craniale della tibia il quale esita in un processo osteoartrosico. La risoluzione chirurgica rappresenta la migliore soluzione terapeutica. Le tecniche chirurgiche extra-articolari con sfruttamento dei punti isometrici del ginocchio si presentano come delle procedure molto diffuse e utilizzate. Questa tesi propone di validare l’uso di un nuovo sistema di navigazione computerizzato-assistito per la valutazione cinematica durante la ricostruzione del LCCr nel cane, ma soprattutto di studiare e confrontare il comportamento e l’efficacia dopo ricostruzione TightRope (TR) in due diverse coppie di punti isometrici. Abbiamo effettuato due analisi in parallelo. La prima eseguendo interventi chirurgici con tecnica TR su 18 casi clinici e sfruttando il punto isometrico del femore (F2) e due diversi punti isometrici della tibia (T2 o T3). L’analisi prevedeva dei controlli postoperatori a 1, 3 e 6 mesi. Ad ogni controllo veniva effettuata una visita ortopedica, esami radiografici, un questionario di valutazione clinico e di soddisfazione del proprietario. Mentre nella ricerca Ex-Vivo abbiamo eseguito dei test su 14 preparati anatomici con l’utilizzo di un sistema di navigazione computerizzato per la rilevazione dei dati. L’analisi prevedeva la valutazione dell’articolazione in diversi stadi: LCCr intatto; LCCr rotto; dopo ricostruzione con TR in F2-T2 e tensionato a 22N, 44N e 99N; dopo ricostruzione con TR in F2-T3 e tensionato a 22N, 44N e 99N. Ad ogni stadio si eseguivano cinque test di valutazione, tra cui: Test del Cassetto, Test di compressione tibiale (TCT), Rotazione Interna/Esterna, Flesso/Estensione e Varo/Valgo. Lo scopo di tale studio è quello di confrontare tra loro i punti isometrici del ginocchio e di analizzare l’efficacia della tecnica TR nelle due differenti condizioni di isometria (F2-T2 e F2-T3).

The porcine animal model goes through the 3Rs: development of in vitro and ex vivo system to study vascular biology

Since the publication of the book of Russell and Burch in 1959, scientific research has never stopped improving itself with regard to the important issue of animal experimentation. The European Directive 2010/63/EU “On the protection of animals used for scientific purposes” focuses mainly on the animal welfare, fixing the Russell and Burch’s 3Rs principles as the foundations of the document. In particular, the legislator clearly states the responsibility of the scientific community to improve the number of alternative methods to animal experimentation. The swine is considered a species of relevant interest for translational research and medicine due to its biological similarities with humans. The surgical community has, in fact, recognized the swine as an excellent model replicating the human cardiovascular system. There have been several wild-type and transgenic porcine models which were produced for biomedicine and translational research. Among these, the cardiovascular ones are the most represented. The continuous involvement of the porcine animal model in the biomedical research, as the continuous advances achieved using swine in translational medicine, support the need for alternative methods to animal experimentation involving pigs. The main purpose of the present work was to develop and characterize novel porcine alternative methods for cardiovascular translational biology/medicine. The work was mainly based on two different models: the first consisted in an ex vivo culture of porcine aortic cylinders and the second consisted in an in vitro culture of porcine aortic derived progenitor cells. Both the models were properly characterized and results indicated that they could be useful to the study of vascular biology. Nevertheless, both the models aim to reduce the use of experimental animals and to refine animal based-trials. In conclusion, the present research aims to be a small, but significant, contribution to the important and necessary field of study of alternative methods to animal experimentation.

Radiolabeling of defined polymer architectures with fluorine-18 and iodine-131 for ex vivo and in vivo evaluation : visualization of structure-property relationships

Nuclear medicine imaging techniques such as PET are of increasing relevance in pharmaceutical research being valuable (pre)clinical tools to non-invasively assess drug performance in vivo. Therapeutic drugs, e.g. chemotherapeutics, often suffer from a poor balance between their efficacy and toxicity. Here, polymer based drug delivery systems can modulate the pharmacokinetics of low Mw therapeutics (prolonging blood circulation time, reducing toxic side effects, increasing target site accumulation) and therefore leading to a more efficient therapy. In this regard, poly-N-(2-hydroxypropyl)-methacrylamide (HPMA) constitutes a promising biocompatible polymer. Towards the further development of these structures, non-invasive PET imaging allows insight into structure-property relationships in vivo. This performant tool can guide design optimization towards more effective drug delivery. Hence, versatile radiolabeling strategies need to be developed and establishing 18F- as well as 131I-labeling of diverse HPMA architectures forms the basis for short- as well as long-term in vivo evaluations. By means of the prosthetic group [18F]FETos, 18F-labeling of distinct HPMA polymer architectures (homopolymers, amphiphilic copolymers as well as block copolymers) was successfully accomplished enabling their systematic evaluation in tumor bearing rats. These investigations revealed pronounced differences depending on individual polymer characteristics (molecular weight, amphiphilicity due to incorporated hydrophobic laurylmethacrylate (LMA) segments, architecture) as well as on the studied tumor model. Polymers showed higher uptake for up to 4 h p.i. into Walker 256 tumors vs. AT1 tumors (correlating to a higher cellular uptake in vitro). Highest tumor concentrations were found for amphiphilic HPMA-ran-LMA copolymers in comparison to homopolymers and block copolymers. Notably, the random LMA copolymer P4* (Mw=55 kDa, 25% LMA) exhibited most promising in vivo behavior such as highest blood retention as well as tumor uptake. Further studies concentrated on the influence of PEGylation (‘stealth effect’) in terms of improving drug delivery properties of defined polymeric micelles. Here, [18F]fluoroethylation of distinct PEGylated block copolymers (0%, 1%, 5%, 7%, 11% of incorporated PEG2kDa) enabled to systematically study the impact of PEG incorporation ratio and respective architecture on the in vivo performance. Most strikingly, higher PEG content caused prolonged blood circulation as well as a linear increase in tumor uptake (Walker 256 carcinoma). Due to the structural diversity of potential polymeric carrier systems, further versatile 18F-labeling strategies are needed. Therefore, a prosthetic 18F-labeling approach based on the Cu(I)-catalyzed click reaction was established for HPMA-based polymers, providing incorporation of fluorine-18 under mild conditions and in high yields. On this basis, a preliminary µPET study of a HPMA-based polymer – radiolabeled via the prosthetic group [18F]F-PEG3-N3 – was successfully accomplished. By revealing early pharmacokinetics, 18F-labeling enables to time-efficiently assess the potential of HPMA polymers for efficient drug delivery. Yet, investigating the long-term fate is essential, especially regarding prolonged circulation properties and passive tumor accumulation (EPR effect). Therefore, radiolabeling of diverse HPMA copolymers with the longer-lived isotope iodine-131 was accomplished enabling in vivo evaluation of copolymer P4* over several days. In this study, tumor retention of 131I-P4* could be demonstrated at least over 48h with concurrent blood clearance thereby confirming promising tumor targeting properties of amphiphilic HPMA copolymer systems based on the EPR effect.

Detection of dentinal cracks after root-end resection: an ex vivo study comparing microscopy and endoscopy with scanning electron microscopy

Dentinal cracks are occasionally observed at the cut root face after root-end resection in apical surgery. The objective of this ex vivo study was to evaluate and compare the efficiency of visual aids to identify root-end dentinal cracks.

Steerable intravitreal inserts for drug delivery: in vitro and ex vivo mobility experiments

The progress of wet age-related macular degeneration can now be controlled by intravitreal drug injection. This approach requires repeated injections, which could be avoided by delivering the drug to the retina. Intraocular implants are a promising solution for drug delivery near the retina. Currently, their accurate placement is challenging, and they can only be removed after a vitrectomy. In this paper, we introduce an approach for minimally invasive retinal drug delivery using magnetic intraocular inserts. We briefly discuss the electromagnetic-control system for magnetic implants and then focus on evaluating their ability to move in the vitreous humor. The mobility of magnetic intraocular implants is estimated in vitro with synthesized vitreous humors, and ex vivo with experiments on cadaver porcine eyes. Preliminary results show that with such magnetic implants a vitrectomy can be avoided.

Impact of corneal cross-linking on drug penetration in an ex vivo porcine eye model

To analyze the influence of corneal cross-linking (CXL) using ultraviolet-A and riboflavin on corneal drug penetration of topically applied drugs.

Effect of topical anesthetic agents and ethanol on corneoepithelial wound healing in an ex vivo whole-globe porcine model

To assess the impact of topical anesthetic agents and ethanol on ocular surface wound healing using an ex vivo whole-globe porcine model.

Effects of Echinaforce® treatment on ex vivo-stimulated blood cells

The herb Echinacea purpurea, also called purple coneflower, is regarded as an immune modulator. This study examined changes in cytokine production in blood samples from 30 volunteers before and during 8-day oral administration with an ethanolic extract of fresh Echinacea purpurea (Echinaforce(®)). Daily blood samples were ex vivo stimulated by LPS/SEB or Zymosan and analysed for a series of cytokines and haematological and metabolic parameters. Treatment reduced the proinflammatory mediators TNF-α and IL-1β by up to 24% (p<0.05) and increased anti-inflammatory IL-10 levels by 13% (p<0.05) in comparison to baseline. This demonstrated a substantial overall anti-inflammatory effect of Echinaforce(®) for the whole group (n=28). Chemokines MCP-1 and IL-8 were upregulated by 15% in samples from subjects treated with Echinaforce(®) (p<0.05). An analysis of a subgroup of volunteers who showed low pre-treatment levels of the cytokines MCP-1, IL-8, IL-10 or IFN-γ (n=8) showed significant stimulation of these factors upon Echinaforce(®) treatment (30-49% increases; p<0.05), whereas the levels in subjects with higher pre-treatment levels remained unaffected. We chose the term "adapted immune-modulation" to describe this observation. Volunteers who reported high stress levels (n=7) and more than 2 colds per year experienced a significant transient increase in IFN-γ upon Echinaforce(®) treatment (>50%). Subjects with low cortisol levels (n=11) showed significant down-regulation of the acute-phase proteins IL1-β, IL-6, IL-12 and TNF-α by Echinaforce(®) (range, 13-25%), while subjects with higher cortisol levels showed no such down-regulation. This is the first ex vivo study to demonstrate adapted immune-modulation by an Echinacea preparation. While Echinaforce(®) did not affect leukocyte counts, we speculate that the underlying therapeutic mechanism is based on differential multi-level modulation of the responses of the different types of leukocytes. Echinaforce(®) thus regulates the production of chemokines and cytokines according to current immune status, such as responsiveness to exogenous stimuli, susceptibility to viral infection and exposure to stress.

Prospective ex-vivo study on thermal effects in human skin phototypes II, IV and VI: a comparison between the 808, 1064, 1210 and 1320-nm diode laser

Laser Assisted Skin Healing (LASH) was first introduced in 2001 by Capon and Mordon to prevent keloids and hypertrophic scars. LASH requires homogenous heating throughout the full thickness of the skin around the wound. However, LASH therapy with 808-nm diode laser is deemed to be only applicable for phototype I-IV due to melanin absorption. This prospective ex-vivo study aims to evaluate the thermal effects of different wavelengths (808, 1064, 1210 and 1320 nm) on human skin phototype II, IV and VI.