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Background: Tibiofemoral components rotational congruency affects the total knee arthroplasty (TKA) success. The smart insert sensor (I-S) helps to establish tibial component rotation reciprocally to a fixed femoral rotation. We aimed (1) to validate the use of I-S as a possible tool to reach reproducible reciprocal femorotibial rotation (RftR) in TKA independently from anatomic landmarks, reducing outliers in combined and mismatched femorotibial rotation (CftR and MMftR, respectively) positioning and (2) to validate the "curve-on-curve" method for a specific type of asymmetrical tibial component., Methods: From February 2018, we conducted a prospective case-control study including 106 patients undergoing TKA. Patients were divided into 2 groups based on the method used to establish tibial component rotation: with the I-S use (group A, n = 53) and with the standard "curve-on-curve" technique (group B, n = 53). Rotational alignment was calculated using the Berger protocol with postoperative computed tomography scanning. Alignment parameters measured were tibial and femoral component rotations (tR, fR), the CftR, the MMftR, and the RftR., Results: Intraoperative rotation measured by I-S correlated the best with RftR (r = 0.84; P < .001) at the post-operative CT scanning. No significant differences were found between groups A and B regarding all types of rotation (fR: P  = .774; tR: P  = .467; CftR: P  = .847) except for MMftR ( P  = .036) and RftR ( P  = .023). There were no outliers in group A but 27 and 12 outliers in group B for MMftR and CftR respectively ( P < .001; P  = .032)., Conclusions: The TKA components' rotation established using a smart I-S intraoperatively is ascribable to the RftR at postoperative computed tomography scan. The I-S helps reduce outliers in the CftR and MMftR. An asymmetrical tibial base plate implanted using the curve-on-curve technique does not create a neutral reciprocal femorotibial rotation significantly increasing the number of cases with mismatched femorotibial internal rotation., (© 2024 The Authors.)

Purpose: Despite modern advancements in the treatment of late stages of wrist joint degeneration, few reliable options exist for patients requiring motion-preserving reconstruction of their radiocarpal and midcarpal joints. Vascularized composite allotransplantation (VCA) could be considered an option for wrist reconstruction in the future. The goal of this study was to describe the relevant anatomy and design a potential surgical technique for wrist VCA., Methods: Anatomic studies were performed on 17 human upper extremities. The arterial system of each cadaver was injected with latex dye or radiographic contrast. After injecting a contrast medium visible on a computerized tomography (CT) scan, the initial three specimens were examined using microCT. This confirmed joint vascular patency and allowed for the dissection of the other specimens that were injected with latex for the study of joint vascularization and the design of the wrist VCA. We then outlined a donor and recipient surgical technique for transplant based on recipient CT scans. Customized cutting guides were designed for the transplant procedure. After the procedure, we performed angiography of the VCA to determine the vascularity of the transplant., Results: Using a combined volar and dorsal approach, we were able to perform a complete wrist VCA procedure. After the completed transplant procedure, angiographic imaging of the specimens demonstrated that the flap dissection and transplantation preserved the nutrient endosteal supply to the distal end of the radius and ulna, as well as to the carpal bones and the metacarpal bases., Conclusions: The dissection of the donor, recipient, and the entire vascularized joint transplant procedure served to illustrate the anatomical feasibility of the cadaveric surgical technique. This establishes an anatomic basis for the possibility of future human clinical applications., Clinical Relevance: This study helps investigate the anatomical feasibility of a wrist VCA., (Copyright © 2024 American Society for Surgery of the Hand. Published by Elsevier Inc. All rights reserved.)

A method for the determination of the intermetallic diffusion coefficient in the Cu-Au system is described based on energy dispersive X-ray techniques. XRF and EDS analysis were used to measure the thickness of the electroplated gold coating and the copper diffused through it, respectively. This information was used to obtain the diffusion coefficient through an equation based on Fick's law. Colour measurements and metallographic section analysis of the samples were also performed to evaluate alternative methods for a qualitative determination of diffusion rate. The thickness of the gold layer was chosen in agreement with what is used in decorative and functional applications (<1 μm). The measurements were performed on samples heated in a range of temperatures between 100 °C and 200 °C from 12 to 96 h. The results obtained follow a linear trend between the logarithm of the diffusion coefficient and the inverse of the temperature and are in line with the values found in the literature., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

The main goal of this work is to open new perspectives in the field of electrodeposition and provide green alternatives to the electroplating industry. The effect of different anions (SO 4 2- , ClO 3 - , NO 3 - , ClO 4 - , BF 4 - , PF 6 - ) in solution on the electrodeposition of copper was investigated. The solutions, containing only the copper precursor and the background electrolyte, were tailored to minimize the environmental impact and reduce the use of organic additives and surfactants. The study is based on electrochemical measurements carried out to verify that no metal complexation takes place. We assessed the nucleation and growth mechanism, we performed a morphological characterization through scanning electron microscopy and deposition efficiency by measuring the film thickness through X-ray fluorescence spectroscopy. Significant differences in the growth mechanism and in the morphology of the electrodeposited films, were observed as a function of the background electrolyte., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Background: Although predictable implant longevity in total knee replacement (TKR) is now established, work continues to satisfy the demands of patients who seek full restoration of the painless function of the native knee following TKR. This prospective study examines the early clinical outcomes of 156 patients implanted with a novel 'kinematic-retaining' (KR) implant., Methods: 156 Physica KR TKRs were implanted for primary osteoarthritis at three European centres. Patients were reviewed up to two years using radiographic, clinical and functional evaluations., Results: Of the 137 patients retained at two years' follow up, none had been revised. Within 6 post-operative months, 51.7% and 79.9% had excellent clinical and functional KSS values respectively, increasing to 81.8% and 88.3% beyond two years. Mean KSS improvement was 34.8 (from 48.6 to 83.4). All KOOS sub-scores improved significantly with total KOOS improving from a mean of 35.5 (SD ±13.0) to 86.5 (±13.7) at two years post-operatively. Pain and sports KOOS sub-scores improved rapidly during the early post-operative periods, with sustained improvements beyond this. Mean OKS improved by 44.1 (±5.1) at two years. VAS satisfaction scores improved significantly at all time points beyond six weeks. Mean FJS-12 was 75.7 at two years, with no significant effects of age or gender. No progressive adverse radiographic features were noted., Conclusions: Early clinical and radiographic outcomes of this kinematic-retaining knee prosthesis are promising, with improvements in clinical parameters similar to, or exceeding those published in other contemporary TKR designs., Level of Evidence: II, Multicentre Prospective cohort study., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 The Author(s). Published by Elsevier B.V. All rights reserved.)

Background: Total wrist fusion (TWF) is indicated for longstanding degenerative, posttraumatic and/or post-oncological conditions to provide pain relief and wrist stability at partial expense of wrist motion., Patients and Methods: A total of 11 consecutive patients who had completed TWF with Vascularized Fibula Graft (VFG) for massive distal radius defects were identified retrospectively from our center using inpatient records. We evaluated bone fusion times and long term functional outcomes following the procedure. Post-operative grip strength (GS) and prono-supination were objectively measured. The new Disabilities of the Arm, Shoulder and Hand (DASH) questionnaire was used to rate disability and symptoms; pre- and post- operative pain with the Visual Analog Scale (VAS) was assessed. A literature review of the present studies about TWF with VFG was performed, with the aim of comparing long-term functional results of the surgical techniques so far reported in the English literature., Results: Our experience with TWF using VFG appeared slightly better than that found in the literature. The procedure was successful in all the cases, achieving bone union in 4,8 months on average. Complication rate was 27,2%, no flap loss was recorded. There were no wrist instability, deformation or dislocation; mean pronation/supination (P/S) was 57,5°/61,2° Average grip strength resulted 59% of the contralateral side. Mean recorded levels of visual analog scale (VAS) for pain postoperatively were 2,32 ± 0,792, which improved significantly from the pre-operatively value of 7,90 ± 0,79. Mean overall satisfaction was good and all the patients comfortably returned to normal activities., Conclusions: Wrist arthrodesis by means of VFG resulted to be an effective and reliable option in dealing with massive defects of distal radius with involvement of radio-carpal joint. Although the cohort analyzed is relatively small and definitive conclusions cannot be drawn, the long term radiographs and the overall functional outcomes encourage to use the described surgical option over other techniques, such as prosthetic replacement and non-vascularized bone grafts., Competing Interests: Declaration of Competing Interest None., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Background: The use of proximal femoral replacements (PFR) has been recently described for catastrophic internal fixation failures. PFR is an attractive treatment option because it is technically straightforward and allows for immediate mobilization of the patient. The aim of the study was to determine the survivorship, functional outcome and complications' rate in a group of elderly patients who underwent proximal femoral replacement as a salvage treatment after femur cephalomedullary nails' mechanical failures., Methods: We evaluated 21 patients who underwent salvage of a failed cephalomedullary nail by using a single design PFR at our institution between 2014-2017. A cemented stem was used in all cases. Radiographs were assessed for fractures, sign of loosening, presence of heterotopic ossification and leg length discrepancy. Functional evaluation was performed through Harris Hip Score (HHS), FIM™ and Time Up and Go test (TUG). Kaplan-Meier estimator was used to determine the overall implants' survival., Results: The average age at the time of surgery was 83years. The mean follow-up was 3.1years. We recorded 3 dislocations of which 2 required a revision. No case of septic or aseptic failure was reported. Two patients died respectively at 11 and 14 months after surgery. At the last follow-up the mean HHS, FIM™, and the TUG improved significantly (p<0.05)., Conclusion: Immediate weight bearing, good functional outcomes, low complications' and one-year mortality rate make the proximal femur replacement with megaprostheses a potential first line treatment of intertrochanteric/subtrochanteric fixations' failures among elderly, osteoporotic, frail patients. Dislocation is the most common complication to bear in mind within the first six months after surgery., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Introduction: The femur is the most frequent involved site by post-attinic fractures. The appropriate treatment of pathological fractures after radiotherapy is still controversial as they are associated with a high risk of delayed consolidation and non-union. Authors review a single Center series of pathological fractures after radiation therapy in patients affected by soft tissue sarcomas analyzing incidence, risk factors, failure rate and proposing a flow chart of treatment of postattinic fractures of the femur., Methods: Authors selected 570 patients treated by limb salvage surgery associated to radiation therapy from 1992 to 2018. A pathological fracture during follow up was observed in 28 cases (5%). The mean time between the onset of the fracture after the prior surgery + radiotherapy was 70 months (range 3-182). The mean follow-up from the fracture was 86 months (range 9-222)., Results: The fracture treatment was performed with an intramedullary nail in 15 cases. Eight femoral fractures healed uneventfully (57%) and 6 required further surgical procedures. A total of eight patients underwent prosthetic replacement, 3 as a primary treatment and 5 as a salvage procedure after failure of internal fixation. Five patients developed a deep infection (62.5%).We observed 10 non-union (53%) in 19 patients treated with osteosynthesis. Overall amputation rate of the entire series was 18%. Authors propose a flowchart of treatment for femoral fractures., Conclusion: Intramedullary nailing is the treatment of choice in postradiation diaphyseal fractures of long bones, prosthetic replacement in meta-epiphyseal fracture site. Free vascularized grafts remain a valid salvage solution after failure of internal fixation., (Copyright © 2020. Published by Elsevier Ltd.)

Background and Objectives: Upper limb preservation after soft tissue sarcoma (STS) surgical excision is now the accepted gold standard and it often requires reconstruction with free flaps. The purpose of this review is to summarize current literature on upper limb reconstruction with free flaps after STS resection., Methods: A systematic review was performed in July 2019 in PubMed and MedLine Ovid databases according to the PRISMA guidelines., Results: A total of 17 studies were included in the final analysis, with 132 patients. The most common diagnosis was Malignant Fibrous Histiocytoma. The most frequent timing of flap coverage was immediate. The success rate was almost always 100%. The length of follow-up was reported in 11 studies with a range of 2-187 months. The most commonly reported patient-centered outcome was the MSTS Score. Based on the evidence of the literature collected, we divided the upper limb into four parts (shoulder, elbow and arm, forearm and wrist, and hand) and described the most common and functional free flaps used for reconstruction after STS resection., Conclusions: Free flaps in the treatment of STS of the upper extremity have a good overall outcome, with a low postoperative complication rate. A wide array of free flaps is available for reconstruction, and the choice of flap is based on defect size, types of tissue required, postoperative functional goal, and surgeon preference. A greater degree of standardization is needed in the reporting of patient-centered outcomes to facilitate future comparative studies., Competing Interests: Declaration of Competing Interest None declared., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

We present the thickness measurement of multilayer samples by X-ray fluorescence (XRF) using calibration curves obtained from simulated spectra through Monte Carlo (MC) algorithm. The XRF is a widespread technique for the analysis of single and multilayer films but the accuracy of quantitative analysis must be increased. Moreover, the use certified standards is not easy to implement due to the high variability of combination and/or concentration in layered samples. The results of this work were compared with fundamental parameter (FP) method and focussed ion beam scanning electron microscopy (FIB-SEM) analysis. The results show good quantitative values even without the use of any standard with known thickness. In addition to having built the calibration curves with a simple univariate approach, also multivariate data analysis was performed to consider multiple variables simultaneously. From the comparison of the obtained results, it can be inferred that the univariate analysis worked well in the case of single layer samples and in the determination of the upper layer in multilayer samples but only multivariate analysis, taking into account the matrix effect of each layer, provided maximum accuracy on each layer of multilayer samples., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Purpose: The aim of this report is to compare free perforator flaps and propeller flaps in the coverage of lower limb soft tissue defects., Patients and Methods: 179 patients (age between 5 and 92 years old), underwent soft tissue reconstruction of the lower limb between January 2009 and January 2015, either by free flap or propeller flap. The two groups were retrospectively evaluated in order to assess the outcome, complications and potential risk factors. Correlations between risk factors and presence/absence of failure or complications have been evaluated with descriptive statistical analysis and a set of logistic regression models. Finally, an economic analysis was conducted to evaluate the different tecniques., Results: In a simple descriptive statistical analysis, the overall failure rate is 6% for free flaps and 3.7% for propeller flaps; the complication rates are 14% vs 21.5% and it increases as dimension increases. The logistic models relating failure and complications with potential risk factors do not show significant differences, whereas the economic analysis show that the average expense of free flaps is 5077.5€ per patient, 1595.6€ per patient for propeller flaps., Conclusions: Our results do not demonstrate significant differences between the two groups about correlation of risk factors or flap size with complication or failure. The surgical option choice should be taken only after accurate evaluation of the soft tissue surrounding the defect. Propeller flaps should be preferred in case of small/medium size defects in otherwise healthy extremities. Free perforator flaps should be the choice in large defects due to trauma or vascular diseases. The economic analysis suggests that propeller flap should be considered when possible., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Background: Intraneural ganglia are benign mucinous cystic formations that originate within the epineurium of peripheral nerves. Different treatments have been recommended, with an overall satisfactory outcome. In this paper, we aim to evaluate the long-term outcomes of surgical treatment of peroneal intraneural ganglia by reviewing our local institutional experience., Methods: We performed a case series review of peroneal intraneural ganglia surgical treatment performed by the senior author. Demographic and surgical details were abstracted from the medical record for each patient. Electrodiagnostic studies and magnetic resonance imaging (MRI) were performed in all patients pre- and postoperatively., Results: Eight men were enrolled, with an average age at time of surgery of 47.5 years (range 28-68 years). Motor testing revealed a preoperative deficit of dorsiflexion, eversion, and toe extension in 7 patients, with a median preoperative Medical Research Council (MRC) score of 0/5. Sensory loss in the distribution of the common peroneal nerve was present in 7 patients. Mean clinical follow-up time was 113 months (range 32-189 months). Significant pain relief was achieved in all patients. Overall neurologic function was improved, more so for motor function. The median postoperative dorsiflexion, eversion, and toe extension at last follow-up were MRC score of 5/5. No complications occurred postoperatively. There was no clinical evidence of intraneural recurrence, as confirmed in postoperative MRI. In 2 patients, an extraneural cystic formation was visible in the anterior muscular compartment., Conclusions: The data from our series support excellent long-term postoperative motor outcomes with a low recurrence rate. To avoid extraneural recurrence, resection of the superior tibiofibular joint is necessary., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Objectives: To evaluate the effects of perioperative myocardial infarction in patients with hip fracture referred to multidisciplinary unit at a tertiary teaching hospital., Methods: 1030 patients with hip fracture underwent troponin measurement, electrocardiogram and echocardiogram at admission, 24 and 48 h after surgery. Exclusion criteria were age < 70 years, severe aortic stenosis, myocardial infarction <30 days, stress cardiomyopathy, renal failure, sepsis, active neoplastic disease. End-points were 30-day and 1 year mortality., Results: Troponin I levels ≥0.5 μg/l were found in 129/1030. 37 of them were excluded according to reported criteria. In the 92 patients included in the study in hospital and 1 year mortality were significantly higher than in controls (12.5% vs 3.5%, p .0012 and respectively 44% vs 16.1% at 12 months, p < .001). 18 patients underwent coronary angiography within 1 week from hip surgery. All had multivessel coronary artery disease. One patient died after angiography. At multivariate logistic analysis age (OR 1.09, 95% CI = 1.01 to 1.19, p = .044) and creatinine values (OR = 7. 55, 95% CI = 1.26 to 45.3, p = .02) were independent predictive factors of 1 year mortality whereas coronary revascularization (OR = 0.15, 95% CI = 0.03 to 0.78, p = .024) was an independent factor associated with improved survival., Conclusions: Perioperative TnI elevation is associated with a significantly increase in 30-day and 1-year mortality. Severe coronary disease may be suspected in patients with perioperative myocardial infarction after hip fracture surgery. Our study is one of the first providing data on the safety and feasibility of early (inhospital) coronary angiograpy and PCI after hip surgery. Further studies are needed to establish indication of coronary angiography in these patients., (Copyright © 2018. Published by Elsevier B.V.)

Background: Oxidized zirconium (OxZr) has demonstrated excellent mechanical properties in vitro when used against articular cartilage; less coefficient of friction and less chondral damage have been found when compared with cobalt-chromium (CoCr) implants. However, controversy exists as to whether implants with a zirconium femoral component articulate safely with a native patella in total knee arthroplasty (TKA). To answer this question, the clinical and radiographic results were analysed from a group of patients who underwent a TKA with patella retention; the OxZr versus CoCr femoral components were compared., Methods: The present study prospectively evaluated 83 knees of 74 patients from 2009 to 2010. Each patient was evaluated clinically (visual analogue scale, Knee Society score, patellar score) and radiographically (long leg standing radiograph, anterior-posterior and latero-lateral projections, axial view of the patella) pre-operatively and postoperatively with a mean follow-up of 4.47years. The patellar tilt and shift, and progression of patellofemoral osteoarthritis were calculated with the axial view., Results: There were no patient reported adverse reactions and none of the evaluated prostheses failed. Both the clinical and radiographic evaluations showed no statistically significant between-group differences., Conclusion: No adverse events were observed clinically or radiologically. These results justify pursuing the use of oxidized zirconium as an alternative bearing surface for a femoral component associated with patellar retention in TKA., (Published by Elsevier B.V.)

Background: The aging process affects the skin, muscle, and fat of the eyes in a different manner. Their individual rejuvenation would require specific surgical treatments according to their particular demands. In the present study, we analyzed the effect of an orbicularis oculi muscle flap fixed to the superior orbital rim to prevent lower eyelid dislocation during transcutaneous blepharoplasty., Methods: The study was conducted retrospectively comparing pre- and postoperative images of two different groups of patients: group A (20 patients) treated with an orbicularis oculi flap and group B (17 patients) treated without the flap. Pre- and postoperative distances between the center of the pupil and upper border of the lower lid at the midpupillary line were measured in each photo and then compared and statistically analyzed., Results: The mean age of the 37 patients was 57 years. From the estimated linear mixed model, we observed a significant effect of the presence of the flap (p-value < 0.0001). All the patients treated with the orbicularis oculi flap (group A) showed a cranial movement of the lower eyelid, although only minimally. All the patients treated without the muscle flap (Group B) showed a caudal movement of the lower eyelid. In total, 47% of the eyes in group B showed a positive variation of >10%., Conclusion: The study suggests that the anchorage of the orbicularis muscle flap to the upper orbital rim reinforces the lower eyelid's anterior lamellae. It efficiently stabilizes lower eyelid position after transcutaneous blepharoplasty, thus confirming clinical results and improving surgical outcomes even in patients with minimal or no eyelid laxity., (Copyright © 2016 British Association of Plastic, Reconstructive and Aesthetic Surgeons. Published by Elsevier Ltd. All rights reserved.)

A fast and selective capillary electrophoresis method has been developed for the simultaneous determination of the antihypertensive drugs captopril and hydrochlorothiazide and their related impurities in a combined dosage form. Method development was carried out implementing each step of Quality by Design workflow, the new paradigm of quality outlined in International Conference on Harmonisation Guidelines. Captopril is characterized by the lack of a strong chromophore and contains a proline-similar moiety, which gives rise to the presence of interconverting cis-trans isomers and leads to the possible interference between electrophoretic migration and reaction of isomerization. The scouting phase was dedicated to the investigation of several operative modes in order to overcome detection and isomerization issues. The best performances were obtained with sodium cholate-based micellar electrokinetic chromatography with the addition of n-butanol and γ-cyclodextrin. Critical quality attributes were represented by the critical resolution values and by analysis time. Critical process parameters were defined as temperature, voltage, concentration and pH of borate buffer, concentration of sodium cholate, n-butanol and γ-cyclodextrin. Screening experimental design was applied for investigating knowledge space. Response surface methodology pointed out several significant interaction effects, and with Monte-Carlo simulations led to map out the design space at a selected probability level. Robustness testing was carried out and a control strategy based on system suitability tests was defined. The selected working conditions gave a complete separation of the analytes in less than three minutes. The method was validated and applied to the analysis of a real sample of coformulation tablets., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Background: Paramedian forehead flap represents a workhorse in facial soft tissue reconstruction but always requires a second step in order to interrupt the skin bridge. Direct visualization and dissection of vascular pedicle may avoid this drawback., Methods: Twenty one patients were treated between January 2012 and January 2014: 6 underwent nasal reconstruction and 15 full thickness defect of medial canthus of an orbital region. The follow-up ranges between 6 months and 2 years. All procedures were carried out in a single stage., Result: All flaps were performed in a single-stage procedure, survived and healed uneventfully., Conclusion: The authors describe a refinement of the original technique to be reserved for cases where vessels could be identified by Doppler, consisting in the direct visualization and dissection of the initial tract of the vascular pedicle, thus providing an extremely mobile pedicle whose size allows to locate the first part of the pedicle in the nasion, tunneling under a skin bridge between the emergence of the vessels and the defect., (Copyright © 2016. Published by Elsevier Ltd.)

Industrial processing of materials containing quartz induces physico-chemical modifications that contribute to the variability of quartz hazard in different plants. Here, modifications affecting a quartz-rich sand during cast iron production, have been investigated. Composition, morphology, presence of radicals associated to quartz and reactivity in free radical generation were studied on a raw sand and on a dust recovered after mould dismantling. Additionally, cytotoxicity of the processed dust and ROS and NO generation were evaluated on MH-S macrophages. Particle morphology and size were marginally affected by casting processing, which caused only a slight increase of the amount of respirable fraction. The raw sand was able to catalyze OH and CO2(-) generation in cell-free test, even if in a lesser extent than the reference quartz (Min-U-Sil), and shows hAl radicals, conventionally found in any quartz-bearing raw materials. Enrichment in iron and extensive coverage with amorphous carbon were observed during processing. They likely contributed, respectively, to increasing the ability of processed dust to release CO2- and to suppressing OH generation respect to the raw sand. Carbon coverage and repeated thermal treatments during industrial processing also caused annealing of radiogenic hAl defects. Finally, no cellular responses were observed with the respirable fraction of the processed powder., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Background: Experimental and clinical studies have shown a strong association between hypertriglyceridemia and diabetic nephropathy. A variability of triglyceride (TG) levels has been reported in diabetes., Objectives: To investigate the relationship of TG variability with the incidence of microalbuminuria (albumin excretion rate > 20 μg/min), in patients with type 2 diabetes., Methods: A longitudinal, retrospective, observational study was performed on a consecutive series of 457 normoalbuminuric outpatients, with measurements of HbA1c, lipids and microalbuminuria thrice per year with 6.8-year follow-up. TG variability, defined as standard deviation of TG (TG-SD) and TG-SD adjusted for the number of visits was calculated. A nested case-control sensitivity analysis was performed to validate the results of the primary cohort study., Results: Incident microalbuminuria (N = 124, 27.1%) was associated with higher median TG-SD (33.6 mg/dL vs 29.0 mg/dL, P < .05) and TG-SD adjusted for the number of visits (31.4 mg/dL vs 26.7 mg/dL, P < .05). At multivariate (Cox) analysis, logTG-SD and adj-logTG-SD were significant predictors of incident microalbuminuria (hazard ratio 2.1 [1.1-4.2], P = .028 and 1.5 [1.1-3.3], P = .042, respectively). In the case-control analysis, time spent with TG > 150 mg/dL during the follow-up was significantly higher in cases vs controls (27.2 ± 19 vs 16.7 ± 12.5 months, P < .05) with hazard ratio 2.0 (1.1-5.1), P < .05, for adj-logTG-SD., Conclusions: A higher intraindividual TG variability is a predictor of incident microalbuminuria in type 2 diabetes. In addition, time of exposure to elevated TG levels (>150 mg/dL) predicts incident microalbuminuria., (Copyright © 2016 National Lipid Association. Published by Elsevier Inc. All rights reserved.)

Rosmarinus officinalis L. is a plant of relevant commercial interest because of its volatile fraction and also its phenolic constituents which are both well known for their numerous properties. Nevertheless, an extractive method suitable to recovering both the aromatic and phenolic fractions from the same fresh foliar tissue has not yet been reported. In this work we have optimized a two-step procedure able to recover first the phenolic compounds and successively the volatile terpenes from the same foliar sample. The recovery of the whole phenolic fraction, partially degraded using a traditional extractive method, was guaranteed and we observed a significant increment in the amount of volatile terpenes compared to a traditional extraction procedure. We also highlight crucial information on the enzymatic activity of the endogenous oxidases that rapidly transform the phenolic substrates, mainly the rosmarinic acid. Our results suggest that this extractive procedure could also be used for other aromatic plants, thus providing a useful tool for more complete analyses of the main phytochemicals available in fresh foliar samples and creating the possibility of incrementing yields of volatile compounds., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Background: New bearing surfaces for total knee replacement have been described in an attempt to reduce polyethylene wear and secondary osteolysis and improve longevity of implants. Oxidized zirconium is a new material that combines the strength of a metal with the wear properties of a ceramic. However, there are no reports as to who documents the long term results. We report a 10 year follow-up note of a selected series of TKAs with an oxidized zirconium femoral component., Methods: We prospectively follow 98 TKAs performed in 94 patients with an oxidized zirconium femoral component. Five patients (5 knees) had died and 6 (6 knees) were lost to follow-up at a minimum of 2 years (mean, 6.3 years; range, 2-9 years) after the operation. For the remaining 83 patients (87 knees), the minimum follow-up was 10 years (mean, 11.3 years; range, 10.0-12.6 years). In 51 cases (58.6%), a cruciate-retaining implant with a deep-dished, more conforming PE was used, and in 36 cases (41.4%), a posterior-stabilized design was used. The patellae were resurfaced in 32 cases (36.7%) and in 55 cases (63.3%) were left unresurfaced., Results: Survivorship was 97.8% at 10 years postoperatively. Two knees were revised for aseptic loosening of the femoral component. No major complication was observed clinically or radiologically. Mean Knee Society score improved from 36 to 84 and functional score from 37 to 83., Conclusions: Oxidized zirconium femoral component in TKA performs well over the first 10 years following implantation, with excellent survival rates and good clinical and radiological outcomes., Level of Evidence: Level IV, therapeutic study., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Vascularised fibular grafts (VFGs) are widely used for primary reconstruction of long bones after bone tumour resections. The biological properties of VFGs are such that they can be a useful option even in failed intercalary reconstructions. The purpose of the current study was to investigate the results and the morbidity of VFGs as a salvage procedure in failed previous reconstructions after intercalary bone tumour resection of the femur. Our series included 12 patients, treated from April 1989 to March 2005, with an average age of 23 years (range 10-43 years) at presentation. The initial diagnosis was osteosarcoma in 10 cases and Ewing's sarcoma in two cases. All patients received chemotherapy and none received radiation therapy. Seven patients received VFG as biologic augmentation in intercalary allograft non-union and in the other five patients, a combination of allograft and VFG was used to replace a cement spacer with hardware failure (four patients) and a failed intercalary prosthesis (one patient). Three patients died during follow-up, in all cases because of metastatic disease. At an average follow-up of 147 months (range 11-260 months), the remaining nine patients were continuously disease-free. Complete healing of the osteotomy of both allograft and VFG was observed in 10 patients at final follow-up. Two major complications were observed that required surgical revision, eventually healing in one case and leading to a poor functional outcome in one case. Significant hypertrophy of the VFG was detected in seven of nine evaluable patients. At final follow-up the mean Musculoskeletal Tumour Society (MSTS)'93 functional score of the nine evaluable patients was 90% (range 66-100%). These results indicate that VFG is a valid salvage procedure in failed intercalary reconstructions of the femur after bone resection., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Purpose: Soft tissue defects involving the anterior aspect of the knee are a frequent finding in a number of pathological conditions. The aim of this article is to describe a new pedicled flap consisting of a conventional medial gastrocnaemius muscle flap associated with a propeller flap based on a perforator of the medial sural artery., Material and Methods: Five males ranging in age between 26 and 72 years underwent a reconstruction of the soft tissue of the knee by means of the described procedure. Three patients sustained complex tissue loss subsequent to high-energy trauma; two losses were due to septic complications after elective knee surgery., Results: Four flaps survived allowing adequate proximal tibial metaphysis and patella coverage. One patient underwent early above-the-knee amputation due to life-threatening septicaemia., Discussion: The described chimaera flap consists of a medial gastrocnaemius flap with a skin paddle that is elevated on a perforator of the medial sural artery and then rotated according to the propeller flaps' principles. It provides effective coverage of large soft tissue defects of the knee. In the authors' experience, the propeller flap portion proved to be particularly useful to cover the patella, while the muscle flap was used to cover the proximal metaphysis of the tibia and fill the dead space if present., (Copyright © 2013 British Association of Plastic, Reconstructive and Aesthetic Surgeons. Published by Elsevier Ltd. All rights reserved.)

Background: Elevation of the joint line frequently occurs in revision total knee arthroplasty (RTKA) because of a wider flexion space than extension space. One solution to balance this flexion-extension space involves the introduction of couplers between the stem and femoral components, and the use of posteriorly offset femoral stems that we hypothesized would improve gap balancing and facilitate joint line restoration., Methods: We retrospectively reviewed a selected series of 43 RTKA. Postoperative joint line height was subtracted from intended height using postoperative lateral radiographs. The value was negative if the joint line position was lowered, and positive if raised., Results: Forty knees were followed for a mean of 3.5years. Mean postoperative joint line position change from intended position was 1.5mm (range -2.5-7.5mm). In 28 knees (70%), the joint line position was restored to within ±2mm of the intended position; in eight knees (20%), from 2-4mm; and in four knees (10%), >4mm. Joint line position was raised in 32 knees (80%) and lowered in eight (20%). In the offset stem knees, the intended joint line position was 0.9mm (range -1.2-3.4mm) as compared with 3.2mm (range -2.5-7.5mm) for the straight stem knees., Conclusions: A coupler system between the femoral stem and femoral component restored the joint line in 70% of cases. The posterior offset stem provided increased posterior condylar offset, addressed the wider flexion space, provided better positioning of the stem, and restored the joint line., Level of Evidence: Therapeutic Study Level IV., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Introduction: Haemophilic arthropathy is the most common clinical manifestation of haemophilia, secondary to recurrent haemarthrosis and chronic synovitis, and the knee represents the main target joint. Modern bleeding prevention has significatively limited the incidence of severe arthropathy, and primary approach is usually conservative. Viscosupplementation is felt as one of the most efficient treatments for the early stages of knee haemophilic arthropathy, based on short-term follow-up studies. The aim of this prospective case series study is to assess the clinical effectiveness of intra-articular administration of hyaluronic acid in the knee, evaluating long-term results, and focusing on the necessity of further treatments after viscosupplementation., Methods: Twenty-seven haemophilic patients with knee arthropathy underwent at least two cycles of injections of hyaluronians between 2003 and 2009. They were evaluated with VAS, SF-36, WFH, Pettersson score, and WOMAC, with a seven-year follow-up., Results: All patients showed improvement in pain relief and functional recovery without any complications. Considering the severity of arthropathy in haemophilic patients, only a limited number of subjects (five) underwent total knee arthroplasty for persistent pain or functional limitation., Conclusions: Viscosupplementation is a safe and effective therapeutic strategy in knee haemophilic arthropathy, with no complications, persisting good clinical results, and determining in most cases a delay of surgery., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Introduction: The use of an intramedullary fibular allograft together with a locking plate fixation has been recently described in biomechanical studies to provide additional medial support and prevent varus malalignment for displaced proximal humeral fractures with promising results. The aim of this study was to evaluate the clinical and radiographic outcome of a locking plate with fibular allograft augmentation in unstable humeral fractures., Methods: We prospectively assessed the functional outcome and complications in 17 patients with proximal humeral fractures with disrupted medial column, treated with a locking plate and a fibular strut graft. The median patient age was 62 years. Postoperative assessments included radiographic imaging, range of motion, pain according to the visual analogue scale (VAS), Short Form (SF36) Health Survey, Constant-Murley and Disabilities of the Arm, Shoulder and Hand (DASH) shoulder scores as well as return to previous occupation and complications., Results: No patients were lost to follow-up and no major complications were recorded. There was no collapse of the humeral head more than 2mm, osteonecrosis or screw penetration of the articular surface. All fractures healed clinically and radiographically. After an average follow-up of 13 months, the mean Constant score was 79 points. The mean active flexion was 149°; extension, 47°; internal rotation, 40°; external rotation, 65°; and abduction, 135°. The median VAS pain level was 1 point. The median DASH score was 33 points, and the median SF36 was 83 points., Conclusion: Locking plate with fibular graft augmentation is a safe and promising technique to support the humeral head and maintain reduction in the proximal humeral fracture with medial comminution., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

The Rosmarinus officinalis L. is widely known for its numerous applications in the food field but also for the increasing interest in its pharmaceutical properties. Two groups of compounds are mainly responsible for the biological activities of the plant: the volatile fraction and the phenolic constituents. The latter group is mainly constituted by rosmarinic acid, by a flavonoidic fraction and by some diterpenoid compounds structurally derived from the carnosic acid. The aim of our work was to optimize the extractive and analytical procedure for the determination of all the phenolic constituents. Moreover the chemical stability of the main phenols, depending on the storage condition, the different drying procedures and the extraction solvent, have been evaluated. This method allowed to detect up to 29 different constituents at the same time in a relatively short time. The described procedure has the advantage to being able to detect and quantify several classes of compounds, among them numerous minor flavonoids, thus contributing to improving knowledge of the plant. The findings from this study have demonstrated that storing the raw fresh material in the freezer is not appropriate for rosemary, mainly due to the rapid disappearing of the rosmarinic acid during the freezing/thawing process. Regarding the flavonoidic fraction, consistent decrements, were highlighted in the dried samples at room temperature if compared with the fresh leaf. Rosmarinic acid, appeared very sensitive also to mild drying processes. The total diterpenoidic content undergoes to little changes when the leaves are freeze dried or frozen and limited losses are observed working on dried leaves at room temperature. Nevertheless it can be taken in account that this fraction is very sensitive to the water presence during the extraction that favors the conversion of carnosic acid toward it oxidized form carnosol. From our findings, it appear evident that when evaluating the phenolic content in rosemary leaves, several factors, mainly the type of storage, the drying process and the extraction methods, should be carefully taken into account because they can induce partial losses of the antioxidant components., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Vascularized bone grafts have been successfully applied for the reconstruction of bone defects at the forearm, distal radius, carpus, and hand. Vascularized bone grafts are most commonly used in revision cases in which other approaches have failed. Vascularized bone grafts can be obtained from a variety of donor sites, including the fibula, the iliac crest, the distal radius (corticocancellous segments and vascularized periosteum), the metacarpals and metatarsals, and the medial femoral condyle (corticoperiosteal flaps). Their vascularity is preserved as either pedicled autografts or free flaps to carry the optimum biological potential to enhance union. The grafts can also be transferred as composite tissue flaps to reconstruct compound tissue defects. Selection of the most appropriate donor flap site is multifactorial. Considerations include size matching between donor and defect, the structural characteristics of the graft, the mechanical demands of the defect, proximity to the donor area, the need for an anastomosis, the duration of the procedure, and the donor site morbidity. This article focuses on defects of the distal radius, the wrist, and the hand., (Copyright © 2010. Published by Elsevier Inc.)

Data from the Italian Hemophilia Centres were collected to perform a retrospective survey of joint arthroplasty in patients with severe hemophilia. Twenty-nine of 49 hemophilia centers reported that 328 of the 347 operations were carried out in 253 patients with severe hemophilia A (HA) and 19 in 15 patients with severe hemophilia B (HB). When results were normalized to the whole Italian hemophilia population (1770 severe HA and 319 severe HB), patients with HA had a 3-fold higher risk of undergoing joint arthroplasty (odds ratio [OR], 3.38; 95% confidence interval [CI], 1.97-5.77; P < .001). These results were confirmed after adjustment for age, HIV, hepatitis C virus (HCV), and inhibitor in a Cox regression model (HR, 2.65; 95% CI, 1.62-4.33; P < .001). The survival analysis of time to joint arthroplasty in the subset of patients with severe HA was not affected by the severity of factor VIII (FVIII) gene mutations. A systematic review of literature articles reporting joint arthroplasties in HA and HB showed that the proportion of HA patients who had undergone arthroplasties was higher than that of HB patients, in agreement with the findings in our Italian cohort. These data suggest that the 2 inherited coagulation disorders have a different severity of clinical phenotype.

A study was carried out to compare the efficacy of two injectable formulations of ivermectin, Ivomec,(1) Merial (IVM reference) and Ivogell,(2) Intervet (IVM generic) in the treatment of psoroptic mange (Psoroptes ovis) in Charollais feedlot cattle. A total of 22 animals were ranked in order of the severity of mange and allocated to 11 replicates of 2 animals each. Within each replicate, one animal was randomly allocated to IVM reference product treatment (Group 1) and one to IVM generic (Group 2). Animals were treated on Day 0 and on Day 8 at the recommended dosage of 200 microg ivermectin/kg bodyweight. The pharmacokinetics profiles (pK) of both IVM formulations were evaluated in plasma samples taken from 6 cattle randomly chosen per group on Day 0, before treatment, and then at 6, 12, 24 hours and daily from Day 2 to Day 7 after the treatment on Day 0. Additionally, the severity of mange lesions was assessed and mites were counted in skin scrapings on Days 0, 8, 15 and 25. Animals were weighed on Day 0 and 25 and body weight and average daily gains (ADG) were evaluated. No statistical differences were found between the cattle of the two groups in any pK parameters, although the mean IVM plasma concentrations in cattle treated with the IVM reference product were consistently higher than those found in cattle treated with the generic compound. By Day 25, all animals in Group 1 had recovered clinically and parasitologically from psoroptic mange while cattle from Group 2 still had mange lesions and, in two animals, living mites were found in the skin scrapings; these differences were significant (P<0.001). The mean body weight of the two groups was significantly different on Day 25 (P<0.01) when animals in Group 1 weighed 20 kg more than those in Group 2. In conclusion, despite similarities in their pharmacokinetic profiles and formulations, the clinical efficacy of the two injectable formulations of IVM differed significantly in their therapeutic efficacy against psoroptic mange in feedlot cattle up to 25 days after treatment: this difference in response was reflected in an incomplete clinical and parasitological response in Group 2 and a slower growth rate.

We investigated the effects of rumen-protected choline (RPC) and vitamin E (VITE) administration on milk production and status of folate, vitamin B12 and vitamin E during the periparturient period of dairy goats. Forty-eight Saanen multiparous goats were selected for the 72-day experiment, being moved to a maternity pen 30 days before expected parturition and assigned to one of the four experimental groups: control (CTR), no choline or vitamin E supplementation; choline (RPC), supplemented with 4 g/day choline chloride in rumen-protected form; vitamin E (VITE), supplemented with 200 IU/day vitamin E in rumen-protected form; and choline and vitamin E (RPCE), supplemented with 4 g/day RPC chloride and 200 IU/day vitamin E. Supplements were administered individually before the morning feed to ensure complete consumption, starting 30 days before kidding and continuing for 35 days after. During the experiment, milk yield and 4% fat-corrected milk (FCM) yield were, respectively, 210 and 350 g/day higher in RPC-supplemented goats than in non-supplemented goats. Milk fat concentration and fat yield were also increased by RPC treatment. Milk yield and composition were unaffected by vitamin E supplementation. There were no significant interactions between RPC and VITE for any of the variables measured. Plasma metabolites did not differ between treatments before and after kidding except that plasma folate at parturition was higher in RPC-supplemented goats. Neither choline nor vitamin E affected vitamin B12 plasma concentrations, while a time effect was evident after the second week of lactation, when B12 levels in each treatment group started to increase. Vitamin E administration resulted in plasma α-tocopherol levels that were 2 to 2.5 times higher than in non-supplemented goats. Overall, these results suggest that greater choline availability can improve milk production and methyl group metabolism in transition dairy goats.

The aim of the present study was to compare the phenolic content in selected monocultivar extra virgin olive oils. Analyses were carried out by HPLC/DAD/MS on Picual, Picuda, Arbequina and Hojiblanca oils from Spain and Seggianese and Taggiasca oils from Italy. Oils from cultivar Picual showed similar characteristics to those of Seggianese oils, with total amounts of secoiridoids of 498.7 and 619.2mg/L, respectively. The phenolic composition of Arbequina oils is close to that of the Taggiasca variety with lignans among the main compounds. The determination of free and linked OH-Tyr, by way of an acid hydrolysis, represents a rapid and suitable method, especially when standards are not available, to determine antioxidant potentialities in terms of MPC, particularly for fresh extra virgin olive oils rich in secoiridoidic derivatives.

The results of 24 modular Genesis II (Smith & Nephew, Memphis, TN, USA) total knee arthroplasties, performed in 20 patients with haemophilia were prospectively reviewed. The hypothesis that the use of a new-generation implant and advanced, more aggressive hematological care, will result in a decrease rate of complications and better functional results for the haemophilic patients with TKA was tested. The mean age at the time of surgery was 36 years (25 to 44). All the patients were reviewed clinically and radiologically with an average follow-up of 4.4 years (2 to 7 years). At the final follow-up the knee score improved from an average of 23 points (11 to 45) to 86 points (62 to 100; p<0.001). The mean knee flexion contracture improved from 22 degrees (0 degrees to 45 degrees ) to 3 degrees (0 degrees to 10 degrees ; p<0.0001). The mean total flexion arc improved from 69 degrees (5 degrees to 130 degrees ) to 92 degrees (80 degrees to 145 degrees p<0.001). The results of our study confirm that the introduction of modular design may improve the functional results of TKA in haemophilic arthropathy which results in a better range of motion and lower flexion contracture.

Vascularised bone graft is a well accepted technique when dealing with long defects. Its role in refractory nonunion, in small defects and in the growing patient is rarely discussed. In this paper the authors review the different alternatives to deal with bone defects in the upper extremity. The indications of vascularised corticoperiosteal graft for solving small defects harbouring refractory nonunion, and the use of vascularised bone phalanx and metatarsal for complex - but small - defects in the fingers is presented. The ability of the bone to grow and remodel when a living epiphysis is included, and to maintain the cartilage viability when a composite osteochondral graft is transferred are also discussed.

The present paper describes our experience of 47 cases of atypical ductal hyperplasia (ADH) diagnosed at vacuum-assisted biopsy. From June 1999 to December 2003, 47 consecutive diagnoses of non-palpable ADH of the breast were made by 11-gauge vacuum-assisted biopsy (Mammotome). Of these, 17 were subjected to surgical excision and 11 underwent a second Mammotome at the site of the previous vacuum-assisted biopsy. Diagnostic underestimation occurred in only two cases, with a surgical diagnosis of ductal carcinoma in situ. In both patients, aged between 46 and 55 years, the radiological images showed microcalcifications of >20 mm, and the lesions were not completely removed by Mammotome. Despite the obvious limitations of the present study, it can be concluded that the probability of underestimating ADH diagnosis by Mammotome appears to be related to the radiological features of the lesion (>20 mm) and to the adequacy of specimens.

We resected the proximal femur in a 4-year-old child who had Ewing's sarcoma. We reconstructed her hip by an autotransplant of the ipsilateral proximal fibula with its vascular supply, inserted inside a massive bone allograft, and fixed to the residual femur by a plate. Serial postoperative imaging showed the progressive hypertrophy of the transplanted fibula, whose head displayed impressive tridimensional growth inside the acetabulum. 4 years after surgery, hip motility is 100 degrees in flexion, 10 degrees in extension, and 30 degrees in abduction, and the patient's limb is weight bearing.

Forty-two single-rooted lower premolars, extracted for periodontal reasons, were endodontically treated and divided into 7 groups of 6 teeth each. In five of the groups, three different types of carbon fiber posts, quartz fiber posts, and titanium posts were used in combination with All Bond 2 dental adhesive. In two groups, two types of carbon fiber posts were also cemented with Panavia 21 dental adhesive. After 3 wk storage in saline, the teeth were longitudinally sectioned; one half was observed using confocal microscopy and the other by scanning electron microscopy. The specimens were evaluated for the presence of a resin dentin interdiffusion zone for the presence of voids at post-resin-dentin interfaces and for the determination of the fiber posts' structure. Upon examination with the confocal microscope, the interfaces of teeth restored with All Bond 2 showed a higher percentage (p < 0.05) of resin dentin interdiffusion zone than those treated with panavia. The fiber size and the post structure were similar in all the fiber posts observed. Some voids were present inside the fiber post structure.

Root canal fillings performed with dentin bonding agent, gutta-percha, and epoxy resin sealer were observed by stereomicroscope. Fourteen palatal roots of extracted maxillary molars were used. Seven roots (group 1) were filled with gutta-percha, epoxy resin sealer, and dentin bonding agent. Seven (group 2) were filled with gutta-percha and epoxy resin sealer alone. The presence of these materials (gutta-percha, adhesive, cement, and a combination of these materials) and the number of voids present were evaluated in the apical, middle, and coronal thirds of the canals. In group 1, a combination of the three materials used was found to be the statistically most frequent case only in the apical third. No statistically significant difference was found between the overall number of voids present in the two groups. The resin-dentin-gutta-percha interface of roots from group 1 was observed by scanning electron microscopy: no statistically significant difference was found between apical and coronal sections regarding the presence of hybrid layer, penetration of resin into the dentinal tubules, or the presence of gaps in either the dentin-resin interface or the resin-gutta-percha interface.

Recurrent digital fibromatosis is a rare, benign fibrous growth of childhood. The condition, which features rapid growth and local recurrences, may be mistaken for a malignant tumor. A case report is presented.

A freeze-fracture study of affected and unaffected psoriatic skin has demonstrated the presence of marked modification of the plasma membrane in the psoriatic lesion. In the lower layers of the epidermis, an increase of membrane associated particles was observed in many keratinocytes, possibly representing the morphological intramembranous equivalent of changes in the outer cell membrane demonstrated with cytochemical techniques. Furthermore, in the malphighian layer, numerous gap junctions have been found, which may be interpreted as a phenomenon compensating the uncontrolled proliferation, and may represent a point of differentiation between cell proliferation in psoriasis and neoplasia. This technique confirmed the poor tendency to adhesion of keratinocytes in extrajunctional areas, which had already been shown by other morphological techniques.