DICER1 somatic mutations strongly impair miRNA processing even in benign thyroid lesions

The alteration of miRNA processing is a driver event in several tumors including thyroid cancer. In particular, somatic DICER1 mutations, reported in follicular-patterned lesions, are shared by benign as well as malignant tumors. In the present study, we investigated the effects of alterations in the miRNA processing genes on the miRNA profile. The study included 19 follicular adenomas (FAs) and 22 follicular variant of papillary thyroid carcinomas (FVPTCs). The mutational status in the hot spot regions of DICER1, DROSHA, TARBP2, DGCR8 and the most commonly affected genes in thyroid tumors was investigated on both tumor and paired normal tissues. The miRNA profile and the mRNA expression levels of DICER1, DROSHA, TARBP2, DGCR8 and XPO5 were also evaluated. Two DICER1 RNase IIIb domain mutations were found in FAs. These lesions presented a considerable loss of 5p miRNAs. Fifteen miRNAs were specifically deregulated in DICER1-mutant lesions compared to FAs and FVPTCs. These miRNAs regulate crucial pathways in cancer such as Hippo, p53 and TGF-beta signalling. DICER1 somatic mutations in the RNase IIIb domain are not specific for malignancy, but the miRNA imbalance that they cause is remarkable, especially with regard to the loss of 5p miRNAs. DICER1-mutant lesions have a characteristic miRNA deregulation, which is different from that of FVPTCs; nevertheless, this impairment is consistent with malignant transformation. Further studies providing the real risk of malignancy associated with DICER1 mutations and the evolution of DICER1-mutant lesions are needed to make them useful in the clinical practice

mRNA and miRNA expression profiling of follicular variant of papillary thyroid carcinoma with and without distant metastases

Follicular Variant of Papillary Thyroid Carcinoma (FVPTC) is usually associated with a good outcome. Nevertheless, in rare cases, it develops distant metastases (1–9%). Our goal was to investigate whether mRNA and miRNA expression profiles may help distinguish between metastatic versus non-metastatic FVPTCs. Twenty-four primary FVPTCs, 12 metastatic and 12 non-metastatic, with similar clinicopathological features were selected and analyzed by nanoString nCounter technology using two distinct panels for expression analysis of 740 mRNA and 798 miRNAs. Data analysis was performed using the nanoString nSolver 3.0 software. Forty-seven mRNA and 35 miRNAs were differentially expressed between the two groups. Using these mRNA and miRNAs, metastatic and non-metastatic FVPTCs were clearly divided into two distinct clusters. Our results indicate that FVPTCs with metastatic abilities have different expression profiles compared to the non-metastatic. A prospective validation is needed to evaluate the usefulness of this molecular approach in the early identification of high-risk FVPTCs

A multilayer biomaterial for osteochondral regeneration shows superiority vs microfractures for the treatment of osteochondral lesions in a multicentre randomized trial at 2 years

Purpose: The increasing awareness on the role of subchondral bone in the etiopathology of articular surface lesions led to the development of osteochondral scaffolds. While safety and promising results have been suggested, there are no trials proving the real potential of the osteochondral regenerative approach. Aim was to assess the benefit provided by a nanostructured collagen-hydroxyapatite (coll-HA) multilayer scaffold for the treatment of chondral and osteochondral knee lesions. Methods: In this multicentre randomized controlled clinical trial, 100 patients affected by symptomatic chondral and osteochondral lesions were treated and evaluated for up to 2 years (51 study group and 49 control group). A biomimetic coll-HA scaffold was studied, and bone marrow stimulation (BMS) was used as reference intervention. Primary efficacy measurement was IKDC subjective score at 2 years. Secondary efficacy measurements were: KOOS, IKDC Knee Examination Form, Tegner and VAS Pain scores evaluated at 6, 12 and 24 months. Tissue regeneration was evaluated with MRI MOCART scoring system at 6, 12 and 24 months. An external independent agency was involved to ensure data correctness and objectiveness. Results: A statistically significant improvement of all clinical scores was obtained from basal evaluation to 2-year follow-up in both groups, although no overall statistically significant differences were detected between the two treatments. Conversely, the subgroup of patients affected by deep osteochondral lesions (i.e. Outerbridge grade IV and OCD) showed a statistically significant better IKDC subjective outcome (+12.4 points, p = 0.036) in the coll-HA group. Statistically significant better results were also found for another challenging group: sport active patients (+16.0, p = 0.027). Severe adverse events related to treatment were documented only in three patients in the coll-HA group and in one in the BMS group. The MOCART score showed no statistical difference between the two groups. Conclusions: This study highlighted the safety and potential of a biomimetic implant. While no statistically significant differences were found compared to BMS for chondral lesions, this procedure can be considered a suitable option for the treatment of osteochondral lesions

Results of isolated posterolateral corner reconstruction

BACKGROUND: Isolated posterolateral corner (PLC) tears are relatively rare events. Various surgical techniques to treat posterolateral knee instability have been described; because surgical results are linked to cruciate reconstructions it has been difficult to date to define whether one surgical procedure has better prognosis than another. The goal of this study is to determine the clinical outcome of PLC reconstruction following fibular-based technique. MATERIALS AND METHODS: We retrospectively evaluated a case series of patients who received isolated PLC reconstruction between March 2005 and January 2007. Ten patients were surgically treated for isolated injuries and were available for follow-up; average patient age was 27.4 years (range 16-47 years). All patients were treated following the fibular-based technique: double femoral tunnel was performed in six patients, while in the remaining four patients, the reconstruction of the PLC was performed with a single femoral tunnel. Six patients had semitendinosus allograft and four had semitendinosus autograft. All patients had the same evaluation and the same rehabilitation protocol. RESULTS: Mean follow-up was 27.5 months (range 18-40 months). Mean range of motion (ROM) was 143.5 degrees for flexion (range 135-150 degrees) and 0.5 degrees for extension (range 0-3 degrees). Three patients showed 1+ on varus stress test, while on Dial test another three patients showed 10 degrees reduction of external rotation compared with contralateral knee. The average Lysholm score was 94 points (range 83-100), and the mean International Knee Documentation Committee (IKDC) subjective result was 88.48 (range 74-96.5). Based on Lysholm score, the results were excellent in eight knees and good in two knees. On IKDC evaluation, two patients were grade A and eight were grade B. No significant difference in clinical results was observed between single and double femoral tunnel. CONCLUSION: Fibular-based technique showed good results in terms of clinical outcome, restoring varus and rotation stability of knees in treatment of chronic isolated PLC injury

Analysis of gene and microRNA expression profiles in Follicular Variant of Papillary Thyroid Carcinomas with and without distant metastases

Introduction: Follicular variant (FV) is one of the most frequent subsets of papillary thyroid carcinoma (PTC), accounting for 15 to 30% of all PTC cases. Usually FVPTC is associated with a good outcome. Nevertheless, in rare cases, it can have distant metastases (1-9%). Aim: Firstly, the primary goal of this study was to investigate the gene and miRNA expression profiles in two groups of homogeneous FVPTCs that were distinguished only by the presence of distant metastasis. The ultimate goal, however, would be to include these expression profiles in a larger project and use them as useful tools to detect precociously those carcinomas with extremely aggressiveness and metastatic behavior. Methods/Case Presentation: Twelve primary FVPTCs with distant metastasis and 12 non-metastatic FVPTCs with similar clinicopathological features were selected. All tumors were analyzed with the nanoString nCounter system for gene and miRNA expression using two distinct panels, composed of 740 genes, involved in cancer progression, and 798 miRNAs, respectively. Expression data were analyzed using the nanoString nSolver 3.0 software, and an unsupervised hierarchical clustering analysis was performed. In addition, we performed the analysis to evaluate the potential interactions between the differentially expressed miRNAs and genes using DIANA - miRPath v3.0, a web-based computational tool. Results/Discussion: Forty-seven out of the 740 genes were differentially expressed between the metastatic and non-metastatic lesions. Considering only these differentially expressed genes, the clustering analysis divided all the cases into two distinct groups: one included mainly the metastatic cases (82%), and the other one the majority of non-metastatic FVPTCs (75%). Additionally, 35 out of the 798 miRNAs were differentially expressed between the two groups, and the clustering analysis performed using these 35 miRNAs generated two main clusters. The first one included 9 metastatic FVPTCs only, whereas the second one included all the non-metastatic tumors and two metastatic FVPTCs. In addition, the 47 genes and 35miRNAs differentially expressed between Met and NonMet lesions were used to study the potential interactions miRNA-mRNA using DIANA-miRPath v3.0. We identified 34 enriched pathways. Three out of the 34enriched pathways, the ECM-receptor interaction (hsa04512), the TGF-β signaling (hsa04350) and the Cell Cycle (hsa04110) and pathways, were included in the PanCancer Progression Panel. In 2 out of 3 pathways, some miRNAs targeted genes differentially expressed between the two groups, determining an interaction miRNA-mRNA. Conclusions: Our results clearly indicate that FVPTCs with metastatic abilities have different gene and miRNA expression profiles compared to their non-metastatic counterparts. A prospective validation is needed to evaluate the usefulness of this molecular approach in the early identification of high-risk FVPTCs

ANALISI CITOGENETICHE IN COLTURE DI LINFOCITI UMANI DOPO TRATTAMENTO CON NANOPARTICELLE DI OSSIDO DI RAME

Con il termine nanoparticella si identificano normalmente delle particelle formate da aggregati atomici o molecolari con un diametro inferiore a 100 nm; possiamo ritrovarle nei prodotti di combustione, nei prodotti di attività fotochimica e vulcanica, nelle gomme da masticare, nelle emissioni veicolari, nei prodotti personali, quali creme solari, talco, cosmetici, nei prodotti finali del processo di smaltimento dei rifiuti ad opera di inceneritori. Diversi studi tossicologici presenti in letteratura hanno riscontrato effetti potenzialmente dannosi di tali particelle; queste ultime possono entrare nel corpo umano tramite l’apparato respiratorio, il tratto alimentare o la cute per poi penetrare nelle cellule e successivamente nel nucleo. Una volta raggiunto il nucleo possono interagire con il DNA o con le proteine associate ad esso e indurre un danno al materiale genetico. Recenti studi in letteratura indicano che le nanoparticelle inducono citotossicità, stress ossidativo con conseguente danno al DNA ed aumento della risposta infiammatoria. Lo scopo di questo lavoro di tesi è quello di valutare il potenziale genotossico di nanoparticelle sferiche di ossido di rame (CuO) su cellule monocitarie estratte da sangue periferico (PBMC) prelevato da donatori volontari. Tali colture in vitro vengono utilizzate al fine di testare il potenziale effetto tossico sull’organismo umano. Il potenziale genotossico viene valutato mediante l’utilizzo del test del micronucleo e del test della cometa. Il test del micronucleo consente di valutare il danno alla doppia elica del DNA e il danno cromosomico che ne deriva: l’indicatore della presenza di un danno cromosomico è rappresentato dai micronuclei. Il test della cometa, invece, consente di rilevare e quantificare le lesioni al DNA nucleare mediante l’elettroforesi di lisati cellulari sospesi in agarosio. Questo test è stato utilizzato per valutare il danno primario al DNA indotto dal trattamento, mentre il danno ossidativo viene valutato tramite il test della cometa con gli enzimi ENDO III (endonucleasi III) e FPG (formamidopirimidina glicosidasi). Per effettuare i test della cometa e del micronucleo viene utilizzato il range di concentrazioni determinate, tenendo in considerazione gli studi presenti in letteratura e i precedenti lavori effettuati presso questo laboratorio su colture cellulari differenti ( 1-100 μg/ml )

Management of anterior cruciate ligament revision in adults: the 2022 ESSKA consensus part III-indications for different clinical scenarios using the RAND/UCLA appropriateness method

Purpose: The aim of the ESSKA 2022 consensus Part III was to develop patient-focused, contemporary, evidence-based, guidelines on the indications for revision anterior cruciate ligament surgery (ACLRev). Methods: The RAND/UCLA Appropriateness Method (RAM) was used to provide recommendations on the appropriateness of surgical treatment versus conservative treatment in different clinical scenarios based on current scientific evidence in conjunction with expert opinion. A core panel defined the clinical scenarios with a moderator and then guided a panel of 17 voting experts through the RAM tasks. Through a two-step voting process, the panel established a consensus as to the appropriateness of ACLRev for each scenario based on a nine-point Likert scale (in which a score in the range 1-3 was considered 'inappropriate', 4-6 'uncertain', and 7-9 'appropriate'). Results: The criteria used to define the scenarios were: age (18-35 years vs 36-50 years vs 51-60 years), sports activity and expectation (Tegner 0-3 vs 4-6 vs 7-10), instability symptoms (yes vs no), meniscus status (functional vs repairable vs non-functional meniscus), and osteoarthritis (OA) (Kellgren-Lawrence [KL] grade 0-I-II vs grade III). Based on these variables, a set of 108 clinical scenarios was developed. ACLRev was considered appropriate in 58%, inappropriate in 12% (meaning conservative treatment is indicated), and uncertain in 30%. Experts considered ACLRev appropriate for patients with instability symptoms, aged ≤ 50 years, regardless of sports activity level, meniscus status, and OA grade. Results were much more controversial in patients without instability symptoms, while higher inappropriateness was related to scenarios with older age (51-60 years), low sporting expectation, non-functional meniscus, and knee OA (KL III). Conclusion: This expert consensus establishes guidelines as to the appropriateness of ACLRev based on defined criteria and provides a useful reference for clinical practice in determining treatment indications. Level of evidence: II

Alternatives to meniscus transplantation outside the United States

4noneOver the years, the surgical treatment of meniscal injury or damage has changed from total meniscectomy to partial meniscectomy or repair. The goal with arthroscopic methods is to preserve as much functional meniscal tissue as possible, rather than to remove it. During recent years, efforts have increasingly focused on the promotion of meniscal healing, as well as the replacement of damaged menisci with allografts, scaffolds, meniscal implants, or substitutes. This chapter will focus on three types of meniscal substitutes, their biological and biomechanical properties, and their clinical relevance. None are available for patient use in the USA at the time of this writing. These substitutes are: the biological MenaflexTM or collagen meniscal implant (CMI), the biomimetic ActifitTM meniscal scaffold, and the nonbiological NUsurface® meniscal substitute. In addition to the surgical techniques, clinical outcomes for the respective devices will be reviewed.Condello, Vincenzo; Ronga, Mario; Linder-Ganz, Eran; Zorzi, ClaudioCondello, Vincenzo; Ronga, Mario; Linder Ganz, Eran; Zorzi, Claudi

Autologous micro‐fragmented adipose tissue injection provides significant and prolonged clinical improvement in patients with knee osteoarthritis: a case‐series study

Abstract Purpose Among the conservative strategies to manage patients with symptomatic knee osteoarthritis (OA), an innovative approach exploiting the regenerative capability of adipose tissue and its resident MSCs (Mesenchymal Stem Cells or Medicinal Signalling Cells) has been proposed with encouraging results. This study aims to demonstrate the benefits of autologous micro‐fragmented adipose tissue (MAT) injection in the conservative treatment of knee osteoarthritis and whether any variables may affect the outcome. This is a case series single‐centre study in which patients underwent intraarticular MAT injection without any associated procedures. Methods Based on inclusion and exclusion criteria, 49 patients (67 Knees) were included and retrospectively analysed with a mean follow‐up of 34.04 ± 13.62 months (minimum 11 – maximum 59). Patients were assessed through the WOMAC and KOOS questionnaires at baseline (pre‐treatment) and 1‐, 3‐, 6‐, 12‐, 24‐ and 36‐month follow‐up. A minimal clinically important difference (MCID) of at least 7.5 points for the WOMAC pain scale and 7.2 for the WOMAC function scale compared to the baseline value was used. Results WOMAC and KOOS scores improved after treatment compared to baseline at all follow‐ups with p < 0.001. Male gender and Kellgren‐Lawrence (KL) grade 2 were associated with smaller improvement in WOMAC and KOOS scores (with respect to females and to KL grade 1, respectively) up to 24 months. The percentage of patients who reach the MCID for WOMAC pain is generally lower than that of patients who reach the MCID for WOMAC function (around 80% at all time points), but it increases significantly over time. Moreover, the baseline score of the WOMAC pain and function influence the outcome. Patients with worse symptoms are more likely to reach the MCID. Conclusions Intra‐articular knee injection of MAT for the treatment of knee osteoarthritis (KOA), recalcitrant to traditional conservative treatments, proved to be effective in a high percentage of cases. The positive association between a worse pre‐operative score and a better clinical response to the treatment would support the idea that intra‐articular administration of MAT could be considered in patients with very symptomatic KOA in which joint‐replacement surgeries are not indicated (or accepted). Level of Evidence IV, case series