CT angiography-based radiomics as a tool for carotid plaque characterization: a pilot study.

PURPOSES: Radiomics is a quantitative method able to analyze a high-throughput extraction of minable imaging features. Herein, we aim to develop a CT angiography-based radiomics analysis and machine learning model for carotid plaques to discriminate vulnerable from no vulnerable plaques. MATERIALS AND METHODS: Thirty consecutive patients with carotid atherosclerosis were enrolled in this pilot study. At surgery, a binary classification of plaques was adopted ("hard" vs "soft"). Feature extraction was performed using the R software package Moddicom. Pairwise feature interdependencies were evaluated using the Spearman rank correlation coefficient. A univariate analysis was performed to assess the association between each feature and the plaque classification and chose top-ranked features. The feature predictive value was investigated using binary logistic regression. A stepwise backward elimination procedure was performed to minimize the Akaike information criterion (AIC). The final significant features were used to build the models for binary classification of carotid plaques, including logistic regression (LR), support vector machine (SVM), and classification and regression tree analysis (CART). All models were cross-validated using fivefold cross validation. Class-specific accuracy, precision, recall and F-measure evaluation metrics were used to quantify classifier output quality. RESULTS: A total of 230 radiomics features were extracted from each plaque. Pairwise Spearman correlation between features reported a high level of correlations, with more than 80% correlating with at least one other feature at |ρ|> 0.8. After a stepwise backward elimination procedure, the entropy and volume features were found to be the most significantly associated with the two plaque groups (p < 0.001), with AUCs of 0.92 and 0.96, respectively. The best performance was registered by the SVM classifier with the RBF kernel, with accuracy, precision, recall and F-score equal to 86.7, 92.9, 81.3 and 86.7%, respectively. The CART classification tree model for the entropy and volume features model achieved 86.7% well-classified plaques and an AUC of 0.987. CONCLUSION: This pilot study highlighted the potential of CTA-based radiomics and machine learning to discriminate plaque composition. This new approach has the potential to provide a reliable method to improve risk stratification in patients with carotid atherosclerosis.

Loss of Prosthetic Aortic Valve during TAVI Procedure: Endovascular Treatment in Emergent Setting.

BACKGROUND: Transcatheter aortic valve implantation (TAVI) has proven over the years to be a viable alternative to open surgery. A rare but severe complication is represented by the valve migration. We report a case of TAVI complication due to the loss of the prosthetic valve in the abdominal aorta treated by endovascular approach. METHODS: An 88-year-old patient with severe aortic valve stenosis, symptomatic for dyspnea was proposed for a TAVI because considered at high risk for surgery. During the TAVI procedure, the undeployed device (Edwards SAPIEN 3 - Edwards Lifesciences, Irvine, CA, USA) detached from its delivery system. Several attempts to withdraw the valve fluctuating in the aorta into its supporting system were performed without success. An emergency endovascular treatment was promptly planned to obtain the exclusion from the flow of the embolized valve. Under local anaesthesia, through the percutaneous femoral access already present, a tube aortic endograft (EndurantTM II, Medtronic, Santa Rosa, CA; ETTF2828C70EE) was successfully introduced and deployed in the infrarenal aorta without any related complications. The embolized valve was completely covered by the endgraft and thus fixed to the aortic wall. The first postoperative computer tomography angiography (CTA) confirmed the correct placement of the endograft, the exclusion of the valve from the flow and the patency of the great vessels. No perioperative or postoperative complications were recorded. The patient was discharged on the ninth postoperative day with the indication to a new attempt of TAVI, through transapical access. CONCLUSIONS: In case of intraprocedural loss of an undeplyed valve during TAVI, the valve fixing through endograft deployment in infrarenal aorta is a possible solution.

Complete Relining in Type 3 Endoleak with AFX Endograft Billowing and Severe Kinking: A Case Report.

BACKGROUND: Type 3 endoleak (T3E) is usually treated by endovascular relining. The procedure can be technically complex in cases of endografts with kinking of innermost stents. We report a case of T3E in an AFX (Endologix, Irvine, CA, USA) endograft with sac enlargement, billowing, and severe kinking of the main body stents, managed with a complete relining endovascular procedure. METHODS: A 69-year-old man with severe comorbidities and prior aorto-bi-iliac AFX endograft completed by an Endurant II cuff (Medtronic, Santa Rosa, CA, USA) for a 63-mm asymptomatic infrarenal aneurysm was admitted to our department for a T3E with 7-mm sac enlargement. The computed tomography angiography (CTA) showed perfusion of the aneurysmal sac, AFX fabric disconnection from its stent (billowing), and severe stent kinking of the main body without a residual lumen. A digital subtraction angiography confirmed the T3E. A complete relining was performed by deploying a bifurcated Endurant II through the AFX stents. RESULTS: The 1-year CTA proved the resolution of the endoleak with a stable aneurysmal sac diameter. CONCLUSIONS: In case of T3E with severe main body stent kinking and graft billowing, an endovascular procedure with a complete aorto-bi-iliac relining through inner stents may be considered.

Intraoperative contrast-enhanced ultrasound for early diagnosis of endoleaks during endovascular abdominal aortic aneurysm repair.

BACKGROUND: The aim of this study was to evaluate the feasibility and utility of intraoperative contrast-enhanced ultrasound (CEUS) for early detection of endoleaks (ELs) during endovascular abdominal aortic aneurysm repair (EVAR) compared with completion digital subtraction angiography. METHODS: Patients undergoing elective EVAR from January 2017 to April 2018 were consecutively enrolled in this prospective study. After endograft deployment, two-digital subtraction angiography (2DSA) with orthogonal C-arm angulations (anteroposterior and sagittal view) were routinely performed. After the endovascular treatment of clear, high-flow type I/III ELs detected by 2DSA, intraoperative CEUS was carried out in sterile conditions on the surgical field before guidewire removal. Presence and type of EL were evaluated with 2DSA and CEUS. CEUS was performed with the vascular surgeon blinded to the 2DSA findings. The primary end point was the level of agreement between 2DSA and CEUS to detect any type of EL and type II EL. Agreement between two diagnostic methods was calculated using Cohen's kappa. The secondary end point was utility of CEUS for intraoperative adjunctive procedure guidance. RESULTS: Sixty patients were enrolled (mean age, 78 ± 6 years; 90% male). 2DSA revealed 11 ELs (18%; 1 type IA, 10 type II), and CEUS 25 ELs (42%; 2 type IA, 23 type II). 2DSA and CEUS were in agreement in 39 cases (65%; 32 no ELs, 7 type II ELs). CEUS detected 17 ELs not identified by 2DSA (28%; 2 type IA, 15 type II); 2DSA detected three ELs not identified by CEUS (5%; 3 type II). In one case, 2DSA and CEUS detected type II and type IA ELs, respectively. For EL and type II EL detection, Cohen's kappa was 0.255 and 0.250, respectively (both "fair agreement"). Intraoperative adjunctive sac embolization was performed under CEUS control in 4 cases and technical success was 100%. CONCLUSIONS: Intraoperative CEUS during EVAR is feasible and can detect a greater number of ELs than 2DSA, in particular type II ELs. Further studies are necessary to assess the reliability of this intraoperative diagnostic examination. In type II ELs, CEUS may represent an additional, useful tool for intraoperative sac embolization guidance.

Systematic Review and Meta-Analysis of the Outcome of Different Treatments for Type 1a Endoleak After EVAR.

BACKGROUND: Insufficient evidence is available to recommend a particular strategy for the treatment of type 1a endoleaks (T1aELs) after endovascular abdominal aneurysm repair (EVAR). The aim of this study was to report outcomes of the different treatment modalities proposed for persistent and late-occurring T1aEL after EVAR. METHODS: A systematic review of the literature (database searched: PubMed, Web of Science, Scopus, Cochrane Library) was undertaken until August 2018. Studies about treatment of T1aEL after EVAR (excluding intraoperative treatments during the first EVAR) presenting a series of 5 or more patients with extractable outcome data (at least intraoperative and/or early results) were included. Meta-analyses of proportions were performed using a random-effects model. RESULTS: A total of 39 nonrandomized studies were included (714 patients; 88.1% males, 95% confidence interval [CI] 84.5-91.7; weighted mean age 75.76 years, 95% CI 74.11-77.4). Overall estimated technical success (TS) and clinical success (CS) rates were 93.2% (95% CI 90.5-95.8) and 88.2% (95% CI 84.5-91.9), respectively. Two hundred eighteen patients underwent proximal extension (98.1% TS, 95% CI 96.3-99.8), 131 chimney EVAR (93.9% TS, 95% CI 89.9-97.9), 97 fenestrated EVAR (86.2% TS, 95% CI 77.3-95.1), 90 open conversion (96.5% TS, 95% CI 93-100), 71 embolization (95.2% TS, 95% CI 90.4-100), 35 endostapling (57.2% TS, 95% CI 14.1-100), and 72 conservative treatment (75.4% CS, 95% CI 56.4-94.5). Estimated overall 30-day mortality was 3.2% (95% CI 1.7-4.7), and it was higher for patients undergoing open surgery (6.6%, 95% CI 1.7-11.5). Overall, endoleak resolution during the mean follow-up of 19.4 months (95% CI 15.45-23.36) was maintained in 91% of the patients (95% CI 87.7-94.3). CONCLUSIONS: T1aEL repair appeared generally feasible, with good early to midterm outcomes. Different treatments are available, and the choice should be based on endoleak characteristics, aortic anatomy, and the patient's surgical risk. Conservative treatment and endoleak embolization should be considered only in selected cases, such as low-flow endoleaks and unfit patients.

Interleukin-21 (IL-21) synergizes with IL-2 to enhance T-cell receptor-induced human T-cell proliferation and counteracts IL-2/transforming growth factor-ß-induced regulatory T-cell development.

Interleukin-2 (IL-2) is a mainstay for current immunotherapeutic protocols but its usefulness in patients is reduced by severe toxicities and because IL-2 facilitates regulatory T (Treg) cell development. IL-21 is a type I cytokine acting as a potent T-cell co-mitogen but less efficient than IL-2 in sustaining T-cell proliferation. Using various in vitro models for T-cell receptor (TCR)-dependent human T-cell proliferation, we found that IL-21 synergized with IL-2 to make CD4(+) and CD8(+) T cells attain a level of expansion that was impossible to obtain with IL-2 alone. Synergy was mostly evident in naive CD4(+) cells. IL-2 and tumour-released transforming growth factor-ß (TGF-ß) are the main environmental cues that cooperate in Treg cell induction in tumour patients. Interleukin-21 hampered Treg cell expansion induced by IL-2/TGF-ß combination in naive CD4(+) cells by facilitating non-Treg over Treg cell proliferation from the early phases of cell activation. Conversely, IL-21 did not modulate the conversion of naive activated CD4(+) cells into Treg cells in the absence of cell division. Treg cell reduction was related to persistent activation of Stat3, a negative regulator of Treg cells associated with down-modulation of IL-2/TGF-ß-induced phosphorylation of Smad2/3, a positive regulator of Treg cells. In contrast to previous studies, IL-21 was completely ineffective in counteracting the suppressive activity of Treg cells on naive and memory, CD4(+) and CD8(+) T cells. Present data provide proof-of-concept for evaluating a combinatorial approach that would reduce the IL-2 needed to sustain T-cell proliferation efficiently, thereby reducing toxicity and controlling a tolerizing mechanism responsible for the contraction of the T-cell response.

Different methods of bone marrow harvesting influence cell characteristics and purity, affecting clinical outcomes.

Background: Bone marrow (BM)-derived stem cells were implanted to induce angiogenesis in patients with no-option critical limb-threatening ischemia. Considering the potential for this therapy, conflicting results related to BM harvesting methods have been reported that could affect stem cell concentrations and quality. Methods: A total of 75 patients with no-option critical limb-threatening ischemia were treated with BM implantation. For 58 patients, BM was harvested using a BM aspirate concentrate system (Harvest Technologies; group HT) with a standard aspiration needle, followed by an automated centrifugation process, to produce BM aspirate concentrate. For 17 patients, BM was harvested using the Marrow Cellution system (Aspire Medical Innovation; group MC). CD34+ cells/mL, CD117+ cells/mL, CD133+ cells/mL, CD309+ cells/mL, hematocrit, and BM purity were compared between the two BM preparations. Results: The retrospective analysis of a subset group after adjustment for age shows that the quality of BM obtained using the Marrow Cellution system is better, in terms of purity, than the classic harvesting method before centrifugation. Harvested BM before centrifugation is characterized by a higher percentage of CD133+ cells compared with BM after centrifugation. In contrast, the MC aspirate had a larger amount of very small embryonic-like cells, as indicated by the higher percentage of CD133+, CD34+, and CD45- cells. These differences translated into an increased occurrence of leg amputations in group HT than in group MC and an increase in transcutaneous oxygen pressure in patients treated with BM aspirated using MC. Conclusions: BM manipulation, such as centrifugation, affects the quality and number of stem cells, with detrimental consequences on clinical outcomes, as reflected by the different amputation rates between the two groups.

Twenty-Five Year Multicentre Experience of Explantation of Infected Abdominal Aortic Endografts.

We report a multicenter experience of open conversions (OC) for aortic endograft infections (AEI). We retrospectively analyzed all patients who underwent OC for AEI after endovascular aneurysm repair (EVAR), from 1997 to 2021 in 12 Italian centers. The endpoints were as follows: mortality (30-days, in-hospital), major postoperative complications. Follow-up data included: survival, aortic-related complications, infection persistence or reoccurrence. Fifty-eight patients (mean age: 73.8 ± 6.6 years) were included. Median time from EVAR to OC was 14 months (interquartile range 7-45). Thirty-five patients (60.3%) were symptomatic at presentation. Aortic reconstruction was anatomic in 32 patients (55.2%), extra-anatomic in 26 (44.8%). Thirty-day mortality was 31% (18/58). Six additional patients died after 30 days during the same hospitalization (in-hospital mortality: 41.4%). Most common post-operative complications included respiratory failure (38.6%) and renal insufficiency (35.1%). During 28.1 ± 4 months follow-up, 4 aneurysm-related deaths were recorded. Infection re-occurred in 29.4% of the patients. Estimated survival was 50% at 1 year, and 30% at 5 years, and was significantly lower for patients who underwent extra-anatomic reconstructions (37 vs 61% at 1 year, 16 vs 45% at 5 years; log-rank P = .021). OC for AEI is associated with high early mortality. The poor mid-term survival is influenced by aortic complications and infection re-occurrence.

Evidence of an anti-inflammatory effect of PCSK9 inhibitors within the human atherosclerotic plaque.

BACKGROUND AND AIMS: Preclinical evidence suggests that proprotein convertase subtilisin-kexin type 9 (PCSK9) inhibitors hold anti-inflammatory properties independently of their ability to lower LDL-cholesterol (C). However, whether PCSK9 inhibitors exert anti-inflammatory effects within the atherosclerotic plaque in humans is unknown. We explored the impact of PCSK9 inhibitors, used as monotherapy, compared with other lipid-lowering drugs (oLLD) on the expression of inflammatory markers within the plaque, assessing also the subsequent incidence of cardiovascular events. METHODS: In an observational study, we recruited 645 patients on stable therapy for at least six months and undergoing carotid endarterectomy, categorizing patients according to the use of PCSK9 inhibitors only (n = 159) or oLLD (n = 486). We evaluated the expression of NLRP3, caspase-1, IL-1ß, TNFα, NF-kB, PCSK9, SIRT3, CD68, MMP-9, and collagen within the plaques in the two groups through immunohistochemistry, ELISA, or immunoblot. A composite outcome including non-fatal myocardial infarction, non-fatal stroke, and all-cause mortality was assessed during a 678 ± 120 days follow-up after the procedure. RESULTS: Patients treated with PCSK9 inhibitors had a lower expression of pro-inflammatory proteins and a higher abundance of SIRT3 and collagen within the plaque, a result obtained despite comparable levels of circulating hs-CRP and observed also in LDL-C-matched subgroups with LDL-C levels <100 mg/dL. Patients treated with PCSK9 inhibitors showed a decreased risk of developing the outcome compared with patients on oLLD, also after adjustment for multiple variables including LDL-C (adjusted hazard ratio 0.262; 95% CI 0.131-0.524; p < 0.001). The expression of PCSK9 correlated positively with that of pro-inflammatory proteins, which burden was associated with a higher risk of developing the outcome, independently of the therapeutic regimen. CONCLUSIONS: The use of PCSK9 inhibitors is accompanied by a beneficial remodelling of the inflammatory burden within the human atheroma, an effect possibly or partly independent of their LDL-C lowering ability. This phenomenon might provide an additional cardiovascular benefit.

Surgical and Endovascular Management of Isolated Internal Iliac Artery Aneurysms: A Systematic Review and Meta-Analysis.

OBJECTIVES: The purpose of this paper is to report the different modalities for the treatment of isolated internal iliac artery aneurysms (IIIAA), as well as their outcomes. METHODS: We performed a systematic review of the literature (database searched: PubMed, Web of Science, Scopus, Cochrane Library; last search: April 2020). We included articles reporting on the outcomes for IIIAA interventions comprising at least 5 patients. Studies were included when presenting extractable outcome data regarding intraoperative and/or early results. We performed meta-analyses of proportions for different outcomes, using random effects model. RESULTS: Thirteen non-randomized studies were included (192 patients with 202 IIIAA). IIIAA were symptomatic in the 18.1% (95%CI 9.3-26.9; I2 54.46%, P = .019). Estimated mean IIIAA diameter was 46.28 mm (95%CI 39.72-52.85; I2 88.85%, P < .001). Open repair was performed in 21/202 cases. Endovascular treatments were: embolization (81/181), embolization and hypogastric artery coverage (79/181), hypogastric artery coverage by stent-grafting (15/181), stent-grafting in the hypogastric artery (6/181). Overall estimated technical success (TS) rate was 91.6% (95% CI 86.8-95.5; I2 45.82%, P = .031). TS rate was 94.5% for open surgery (95%CI 85.3-100; I2 0%, P = .907), and 89.7% for endovascular repair (95%CI 83.8-95.6; I2 55.43%, P = .006). Estimated overall 30-day mortality was 3.1% (95%CI 0.8-5.4; I2 0%, P = .969). Mortality rates after open surgery and endovascular repair were 8.2% (95%CI 3.4-19.8; I2 0%, P = .545) and 2.8% (95%CI 0.5-5.1; I2 0%, P = .994), respectively. Estimated mean follow-up was 32.63 months (95%CI 21.74-43.53; I2 94.45%, P < .001). During this timeframe, IIIAA exclusion was preserved in 92.8% of the patients (95%CI 89.3-96.2; I2 0%, P = .797). Buttock claudication occurred in 13.9% of the patients (95%CI 8.7-19.2; I2 0%, P = .622). CONCLUSIONS: IIIAA are frequently large, and symptomatic at presentation. Several treatments are proposed in literature, open and endovascular, both with good results. The endovascular treatment is the preferred method of treatment in literature, since it offers good short- to mid-term results and low early mortality. Buttock claudication after hypogastric artery exclusion is a common complication.

The Utility of Intraoperative Contrast-enhanced Ultrasound for Immediate Treatment of Type Ia Endoleak during EVAR: Initial Experience.

Objectives Type Ia endoleak (EL) after endovascular abdominal aortic repair (EVAR) may be misdiagnosed at completion angiography. Intraoperative contrast-enhanced ultrasound (CEUS) may play a role in early detection and immediate treatment of type Ia EL. Methods From January 2017 to April 2018, patients treated with EVAR underwent intraoperative CEUS. After endograft deployment and ballooning, digital subtraction angiography (DSA) and intraoperative CEUS were performed in a blinded fashion. All cases of type Ia EL at DSA or CEUS were considered. Results Type Ia EL detected at intraoperative CEUS and undetected at DSA was defined in 2 patients. The former was solved with intraoperative re-ballooning; in the latter case, a Palmaz stent deployment was required. The resolution of type Ia EL was detected at intraoperative CEUS control and post-operative computed tomography angiography (CTA). In another patient, the DSA detected a type Ia EL, but intraoperative CEUS reveal a type II EL from lumbar arteries. Post-operative CTA confirm the type II EL. Conclusions The reported cases prove the clinical utility of the intraoperative CEUS, permitting the early identification of 2 type Ia EL. In addition, the intraoperative CEUS is useful in case of dubious type Ia EL at DSA, avoiding unnecessary intraoperative adjunctive procedure or post-operative CTA.

Atherosclerotic Plaque Fissuration and Clinical Outcomes in Pre-Diabetics vs. Normoglycemics Patients Affected by Asymptomatic Significant Carotid Artery Stenosis at 2 Years of Follow-Up: Role of microRNAs Modulation: The ATIMIR Study.

Atherosclerotic plaque instability and rupture in patients with asymptomatic carotid artery stenosis (ACAS) is a leading cause of major adverse cardiac events (MACE). This could be mainly evidenced in patients with pre-diabetes. Indeed, the altered glucose homeostasis and insulin resistance could cause over-inflammation of atherosclerotic plaque, favoring its conversion to unstable phenotype with rupture and MACE. Notably, metformin therapy reducing the metabolic distress and the inflammatory burden could reduce MACE in ACAS patients with pre-diabetes. In this setting, the microRNAs (miRs) could be used as molecular biomarkers of atherosclerosis progression, plaque rupture, and worse prognosis in normoglycemics (NG) versus pre-diabetics metformin users (PDMU) versus pre-diabetics non-metformin users (PDNMU). However, our study aimed to investigate a wide miRNA panel in peripheral blood exosomes from patients with ACAS divided in NG versus PDMU versus PDNMU, and to associate the circulating miRNA expression profiles with MACE at 2 years of follow-up after endarterectomy. The study included 234 patients with ACAS divided into NG (n = 125), PDNMU (n = 73), and PDMU (n = 36). The miRs' expression profiles of circulating exosomes were determined at baseline and at 2 years of follow-up by Affymetrix microarrays from the patients' plasma samples from any study cohort. Then we collected and analyzed MACE at 2 years of follow-up in NG versus PDMU versus PDNMU. Prediabetics versus NG had over-inflammation (p < 0.05) and over expressed miR-24 and miR-27 at baseline. At 2 years of follow-up, PDNMU versus NG, PDMU versus NG, and PDNMU versus PDMU over-expressed inflammatory markers and miR-24, miR-27, miR-100, miR-126, and miR-133 (p < 0.05). Finally, at the end of follow-up, we observed a significant difference about MACE comparing PDNMU versus NG (n = 27 (36.9%) versus n = 8 (6.4%); p < 0.05), PDNMU versus PDMU (n = 27 (36.9%) versus n = 6 (16.6%); p < 0.05); and PDMU versus NG (n = 6 (16.6%) versus n = 8 (6.4%); p < 0.05). Admission glucose values (HR (hazard ratio) 1.020, CI (confidence of interval) 95% (1.001-1.038), p = 0.029), atheromatous carotid plaque (HR 5.373, CI 95% (1.251-11.079), p = 0.024), and miR-24 (HR 3.842, CI 95% (1.768-19.222), p = 0.011) predicted MACE at 2 years of follow-up. Specific circulating miRs could be over-expressed in pre-diabetics and specifically in PDNMU versus PDMU after endarterectomy. MiR24, hyperglycemia, and atheromatous plaque could predict MACE at 2 years of follow-up.

In vivoorganized neovascularization induced by 3D bioprinted endothelial-derived extracellular vesicles.

Extracellular vesicles (EVs) have become a key tool in the biotechnological landscape due to their well-documented ability to mediate intercellular communication. This feature has been explored and is under constant investigation by researchers, who have demonstrated the important role of EVs in several research fields ranging from oncology to immunology and diagnostics to regenerative medicine. Unfortunately, there are still some limitations to overcome before clinical application, including the inability to confine the EVs to strategically defined sites of interest to avoid side effects. In this study, for the first time, EV application is supported by 3D bioprinting technology to develop a new strategy for applying the angiogenic cargo of human umbilical vein endothelial cell-derived EVs in regenerative medicine. EVs, derived from human endothelial cells and grown under different stressed conditions, were collected and used as bioadditives for the formulation of advanced bioinks. Afterin vivosubcutaneous implantation, we demonstrated that the bioprinted 3D structures, loaded with EVs, supported the formation of a new functional vasculaturein situ, consisting of blood-perfused microvessels recapitulating the printed pattern. The results obtained in this study favour the development of new therapeutic approaches for critical clinical conditions, such as the need for prompt revascularization of ischaemic tissues, which represent the fundamental substrate for advanced regenerative medicine applications.

Sodium-glucose co-transporter2 expression and inflammatory activity in diabetic atherosclerotic plaques: Effects of sodium-glucose co-transporter2 inhibitor treatment.

OBJECTIVE: We evaluated sodium-glucose co-transporter2 (SGLT2) expression and the effect of SGLT2 inhibitor (SGLT2i) therapies on carotid plaques of asymptomatic diabetic and non-diabetic patients. METHODS: Plaques were obtained from 296 non-diabetic patients and 227 patients with type 2 diabetes undergoing carotid endarterectomy. 97 patients with type 2 diabetes were treated with SGLT2 inhibitors for 16 ± 4 months before endarterectomy. After propensity score matching analysis, patients with type 2 diabetes were categorized without (n = 87) and with SGLT2i therapy (n = 87). To investigate SGLT2 expression levels' effects on major adverse endpoints (MACE = stroke, transient ischemic attack, myocardial infarction, and death), we evaluated MACE outcomes at a 2-year follow-up. RESULTS: Compared to plaques from patients without diabetes, plaques from patients with diabetes had higher SGLT2 expression, inflammation, and oxidative stress, along with lower SIRT6 expression and collagen content. Compared with plaques from patients with diabetes, SGLT2i-treated patients with type 2 diabetes presented increased SIRT6 expression and collagen content and lowered inflammation and ion and oxidative stress, thus indicating a more stable plaque phenotype. These results supported in vitro observations on human aorta endothelial cells (EC) (TeloHAEC-cells). Indeed, EC treated with high glucose (25 mM) in the presence of SGLT2i (100 nM canagliflozin) presented higher SIRT6 expression and decreased mRNA and protein SGLT2 levels, nuclear factor-kappa B (NF-B(NF-κB), and matrix metallopeptidase 9 (MMP-9) expression compared to cells treated only with high glucose. After two years following endarterectomy, a multivariable Cox regression analysis showed significantly higher 2-year overall survival from MACE in patients without diabetes (P < 0.01). Among patient with diabetes, the current SGLT2i users presented a significantly lower rate of MACE through 2 years compared to non-SGLT2i users (P < 0.05). CONCLUSIONS: These findings unveil a critical involvement of the SGLT2/SIRT6 pathway in the inflammatory process of diabetic atherosclerotic lesions and suggest its possible favorable modulation by SGLT2i.

Non-catalytic region of tyrosine kinase adaptor protein 2 (NCK2) pathways as factor promoting aggressiveness in ovarian cancer.

BACKGROUND: In this study we investigated the function of the non-catalytic region of tyrosine kinase adaptor protein 2 (NCK2) and its correlation with ITGB1 and ITGB4 integrins in driving ovarian cancer (OvCa) aggressiveness. We also evaluated whether NCK2 may influence prognosis in OvCa patients. METHODS: Nanofluidic technology was used to analyze expression of NCK2 in 332 OvCa patients. To evaluate mRNA expression of NCK2, integrins and VEGFA in OvCa cell lines, qRT-PCR was performed. Stable NCK2 overexpression was obtained in OVCAR3. qRT-PCR and Western blot were performed to evaluate expression changes of VEGFA, vimentin, ITGB1, ITGB4, MMP2 and MMP9 under normoxia and hypoxia conditions. Coimmunoprecipitation (Co-IP) was performed in the A2780 cell line to study the interaction between NCK2 and proteins of interest. To investigate whether NCK2 can influence anchorage-independent growth, a soft agar assay was completed. Transwell invasion assay was performed on stable-transfected OVCAR-3 cell lines. RESULTS: Nanofluidic data showed NCK2 can play an important role as a factor promoting tumor aggressiveness and survival in OvCa. This role was also linked to the behaviors of ITGB1 and ITGB4. Moreover, in cells overexpressing NCK2, the expression of vimentin, MMP2, MMP9, VEGFA and ITGB1, but not of ITGB4 was induced by hypoxia. Co-IP showed that NCK2 can directly bind ITGB1, but not VEGFA. NCK2 may be involved in mediating cell-extracellular matrix interactions in OvCa cells by influencing tumor aggressiveness. CONCLUSIONS: This study provides evidence of a possible role of NCK2 as biomarker of OvCa progression.

Preoperative Anti-Class III ß-Tubulin Antibodies As Relevant Clinical Biomarkers in Ovarian Cancer.

Class III ß-tubulin (TUBB3) overexpression in ovarian cancer (OC) associates with poor prognosis. We investigated whether TUBB3 overexpression elicited anti-TUBB3 antibody production in OC patients and whether these antibodies may have diagnostic and prognostic impact. The presence of serum anti-TUBB3 antibodies was investigated in 49 untreated OC patients and 44 healthy individuals by an in-house developed ELISA that used recombinant TUBB3 as the antigen. Receiver operating characteristic (ROC) curves were generated to assess the diagnostic accuracy of the assay. Anti-TUBB3 antibodies discriminated OC patients and healthy individuals with excellent sensitivity and specificity (91.8% and 90.9%, respectively). In multivariate analysis, anti-TUBB3 antibody level emerged as an independent prognostic factor for progression free and overall survival. The ELISA was then optimized using a biotin-labeled TUBB3 C-terminal peptide424-450 instead of recombinant TUBB3 as the antigen and streptavidin-coated plates. The diagnostic role of the anti-TUBB3 antibodies was studied in an independent series of 99 OC patients and 80 gynecological benign disease patients. ROC-curve analysis showed a valuable diagnostic potential for serum anti-TUBB3 antibodies to identify OC patients with higher sensitivity and specificity (95.3% and 97.6%, respectively). Overall, our results provide evidence that preoperative anti-TUBB3 antibody level is a promising diagnostic and prognostic biomarker for the management of OC patients.

One-year experience in carotid endarterectomy combining general anaesthesia with preserved consciousness and sequential carotid cross-clamping.

BACKGROUND AND AIM OF THE WORK: We report 1-year single-centre experience in carotid endarterectomy (CEA) combining general anaesthesia with preserved consciousness (GAPC) and standardized carotid sequential cross-clamping, for our protocol effectiveness evaluation in reduction of perioperative stroke, death or cardiologic complications. METHODS: We considered all patients who underwent CEA in 2016. All patients underwent superficial cervical plexus block and GAPC with Remifentanil. The surgical technique consisted of common carotid artery (CCA) cross-clamping, carotid bifurcation isolation, external (ECA) and internal carotid artery (ICA) cross-clamping. After CCA cross-clamping, we performed a neurological tolerance test (NTT); this allowed selective shunting only for positive NTT. Primary end-points were: transient ischemic attack (TIA)/stroke, myocardial infarction, death in perioperative period. Secondary end-points were: carotid shunting, peripheral cranial nerves injuries (PCNI), GAPC intolerance, other complications, reintervention in perioperative period, length of hospital stay. RESULTS: 104 consecutive patients underwent CEA with this protocol in the considered period. Twenty-seven (25.9%) patients were symptomatic. Mean clamping time was 48±13.5 minutes. Five cases (4.8%) requested internal carotid artery shunting. No TIA/stroke, myocardial infarction or death were recorded in the perioperative period. PCNI were observed in 19 cases (18.2%) in the immediate post-operative period; 16 of them (84.2%) showed complete or partial resolution at discharge. Only one patient (0.9%) showed GAPC intolerance. No other complication occurred. Three patients (2.9%) underwent reintervention for neck haematoma drainage. Mean hospital stay were 3±0.9 days. CONCLUSIONS: GAPC associated with sequential carotid cross-clamping appeared to be safe and effective in prevention of major neurological and cardiologic complications during CEA.