The passive stiffness and strength of periscapular muscles may affect scapulohumeral control, especially in overhead athletes due to sports-specific training. This study tried to assess the relationship between the passive stiffness and strength of periscapular muscles, scapulohumeral kinematics and neuromuscular control during scaption in swimmers. Ten male adolescent competitive front-crawl swimmers were recruited. The passive stiffness and strength of periscapular muscles were measured in standard postures by a hand-held myotonometer and dynamometer, respectively. Surface electromyography and electromagnetic tracking systems were synchronized to record the muscle activities and scapulohumeral kinematics during scaption. Correlations between the passive stiffness or strength of periscapular muscles and the kinematics or muscle activity were examined by Spearman's rank correlation coefficient. The maximal strength of periscapular muscles correlated positively with the ranges of upward and external rotation of the scapula and negatively with muscle activity during scaption. Passive stiffness of periscapular muscles was associated with the downward rotation of the scapula and triggered an increase in muscle activity. Increased passive stiffness or decreased strength in the periscapular muscles may affect their role in controlling the scapular rotation and contribute to compensation from adjacent muscles. Our findings suggest that when attempting to evaluate scapular behavior, it may be beneficial to examine muscle strength and passive stiffness of periscapular muscles. This symptom is commonly referred to as the swimmer’s shoulder and may cause a decline in performance or even end a swimmer’s career. The prevalence of shoulder pain in the front-crawl style (52.44%) is higher than in other stroke types3 SD is commonly seen in the non-breathing side4 or the injured shoulder5 of competitive front-crawl swimmers. During scaption, internal rotation and anterior tilting of the scapula are more pronounced in young competitive swimmers than in non-practitioners6 Since SD is connected to abnormalities in neuromuscular control related to weakness or stiffness of periscapular muscles10 muscle activity should be comprehensively evaluated within training programs.The scapula provides a stable support for arm movements during swimming. Once scapula dyskinesis (SD) appears in athletes, the risk of developing shoulder pain is increased by 43%11 This effect is possibly caused by sport-specific training for competitive swimmers focusing on technique, which may ignore the strength balance between muscles. Long term, the strength of the LT in teenage elite swimmers decreases after three years of swim training, whereas the strength of other shoulder muscles increases12 Unbalanced periscapular muscle strength may disrupt the function of force coupling on the scapular rotations in overhead athletes. Approximately 85% of swimmers with asymptomatic shoulders have altered scapula movement immediately after a single swim training session8 When compared to non-swimmers, elite swimmers seem to have more scapular protraction during scaption13 However, it remains controversial whether SD is the cause or the result of symptomatic shoulders. Middle trapezius (MT) strength in swimmers with symptomatic shoulders was found to be lower than in swimmers with asymptomatic shoulders, even though no difference in SD seems to occurr between groups14.As mentioned, SD may be accompanied by a decline in the strength of periscapular muscle. Overhead athletes with SD were found to have reduced strength of the lower trapezius (LT) compared to athletes without SD15 The stiffness of the shoulder rotator muscles affects not only how flexible the shoulder is, but also how the muscles control the movement of the scapulohumeral joint. One study showed that swimmers had an increase in external rotation and posterior tilt of the scapula during arm elevation after reducing pectoralis minor stiffness through manual stretching16 In another study, although the muscle does not insert directly into the scapula, the latissimus dorsi stiffness was associated with an increased upward rotation and decreased posterior tilt of the scapula during scaption in swimmers9 It is still uncertain whether the stiffness of periscapular muscles is associated with the neuromuscular controls of scapulohumeral kinematics in swimmers.Muscle passive stiffness commonly appears after sports training and restricts the active range of movement. It was found that the range of shoulder external rotation was reduced after a two-hour swim training session due to the repetitive force production of the shoulder internal rotator musclesTherefore, this study aimed to investigate the relationship between the stiffness and strength of periscapular muscles and the neuromuscular control of scapulohumeral movement in competitive adolescent swimmers. The passive stiffness and maximal strength were evaluated in periscapular muscles, and the overall shoulder kinematics and the neuromuscular control of periscapular muscles during scaption were measured.For this correlational study, ten male competitive adolescent swimmers  from a high school team were recruited. All recruited swimmers had practiced middle- (200 and 400 m) and long- (800 and 1500 m) distances and participated in swimming competitions for at least two years. The front-crawl style was their main stroke pattern during training and competitions. They received swim training for three hours per day and five days per week. To eliminate other symptoms that could affect shoulder neuromuscular control, the author (P.-T. C.), who was a physical therapist with nineteen years of experience, performed physical examinations to rule out shoulder disorders . Swimmers were excluded if any musculoskeletal or neurological symptoms on the upper extremity were noticed. After explanation of study purpose and procedures, the participants signed the informed consent before enrolment. This study was conducted in accordance with the Declaration of Helsinki and approved by the review board of Hualien Tzu Chi Hospital.After ensuring the eligibility of participants, basic information  was obtained. The passive stiffness and strength of periscapular muscles and the neuromuscular control of scapulohumeral kinematics of the participant’s non-breathing shoulder were measured. To avoid immediate soreness after muscle exertion, passive stiffness measurements were performed first, and the maximal strength of the periscapular muscles was determined at the end of the experiment. Following the stiffness measurement, the scapulohumeral kinematics and the muscle electromyographic activity of periscapular muscles during scaption were recorded. Finally, the maximal strength of the periscapular muscles was tested.17; MT, the midpoint between the spinous process of the 2nd thoracic vertebra and the medial edge of scapula; LT, the midpoint of a line inclined at 55° from the point 5 cm below the root of spine of scapula down to the intersection with the spinous process of thoracic vertebrae; and SA, the midaxillary line at the level of the inferior angle of the scapula along the rib18 The mean passive stiffness from three trials was obtained.Participants were asked to lie prone on a treatment table with arms by the side in a relaxed position for measurements. A hand-held myotonometer  was used to measure the muscle passive stiffness (N/m) by placing the sensing probe on the upper trapezius (UT), middle trapezius (MT), LT and serratus anterior (SA). The measuring location of each muscle was determined as the following: UT, a point 2 cm lateral to the midpoint between the spinous process of the 7th cervical vertebra and the lateral edge of the acromion19 and the height was adjusted at the level of each participant’s shoulder joint. Then, ten bony landmarks recommended by the International Society of Biomechanics20 were captured by the digitizer in synchrony with the receiver’s position and orientation to construct the local coordinate system (LCS) of each segment. The bony landmarks consisted of incisura jugularis, processus xiphoideus, the spinous process of the 7th cervical vertebra, the spinous process of the 8th thoracic vertebra, trigonum scapulae, angulus inferior of scapulae, angulus acromialis, processus coracoideus, and lateral and medial epicondyles of humerus. After digitization, the participants performed scaption at a comfortable speed (two seconds for elevation and two seconds for lowering) over five trials. A color tape was applied to the ground  as the guiding mark of the scapular plane for the scaption movement. The kinematic data were collected at a sampling rate of 120 Hz and synchronized with the surface electromyography.For the kinematics measurement, three receivers of an electromagnetic motion tracking system  were attached to the sternum notch, lateral plateau of acromial process, and lateral side of distal humerus to record the motion of the thorax, scapula and humerus, respectively. Participants were asked to sit erect in front of the electromagnetic transmitter to minimize the effects of spine posture21. The data of sEMG was sampled at a frequency of 1000 Hz.A wireless sEMG  was utilized to record the muscle activities of UT, MT, LT, SA and middle fiber of the deltoid (MD) during scaption and maximal muscle strength measurements. The surface skin of each muscle was prepared and cleaned with an alcoholic swab and hair shaving. The location for electrode placement of each muscle was palpated and identified: UT, halfway between the spinous process of the 7th cervical vertebra and the acromion; MT, halfway on the horizontal line between the thoracic spine and the root of the scapular spine; LT, obliquely upward and laterally along a line between the intersection of the scapular spine with the vertebral border of the scapula, and the spinous process of 7th thoracic vertebra; SA, anterior to the latissimus dorsi and posterior to the pectoralis major; IS, approximately 4 cm below the spine of the scapula on the lateral aspect over the infrascapular fossa; and MD, on the lateral aspect of the upper arm and approximately 3 cm below the acromion23. For UT, while sitting upright with head in neutral and arm by the side, resistance was applied to the superior aspect of the acromial plateau. For MT, with shoulder abduction at 90° with thumb toward the ceiling in a prone position, resistance was given on the lateral aspect of the spine of the scapula. For LT, with shoulder abduction at 145° with thumb toward the ceiling in a prone lying posture, resistance was applied at the same location as for MT. For SA, with both shoulder and elbow flexion at 90° with shoulder internal rotation in a supine position, resistance was applied on the proximal site of the ulnar near olecranon. For MD, while sitting upright with the shoulder in neutral and arm by the side, resistance was given on the lateral side of the distal humerus. Participants were asked to perform two trials of five-second MVICs against the dynamometer for each muscle. For the subsequent normalization processing, the electromyographic activity of each muscle was recorded during the respective MVIC measurement.The strength of UT, MT, LT, SA and MD during maximal voluntary isometric contraction (MVIC) were assessed using a handheld dynamometer . The posture and resistance for each muscle assessment were according to previous studies20. By transforming the LCS of the distal segment to the LCS of the proximal segment, the rotational matrix of humerus-thorax (HT) and scapula-thorax (ST) can be determined. Then, the three-dimensional HT and ST movement can be obtained from the rotational matrices by the Euler sequences Y-X’-Y’’ and Y-X’-Z’’, respectively40. Finally, the angle of abduction  of HT and the internal rotation , downward rotation  and posterior tilting  of ST were calculated.The spatial relationship between the position of bony landmarks and the orientation of the receiver on the same segment in static status was used to derive virtual bony landmarks during scaption. The LCS of each segment was determined by the virtual landmarks for every time point, where the Y axis was in the inferior-superior direction, the X axis was in the posterior-anterior direction and the Z axis was in the medial–lateral direction Fig. 20. By trAll raw sEMG signals recorded during scaption and MVIC were rectified and filtered with a band-pass filter of 20–500 Hz (4th order Butterworth). Then the signal was smoothed by the root mean square value with a 50 ms moving window. The electromyographic activity of each muscle during scaption was normalized to the peak value during the MVIC test (%MVIC).24 The rotation angles of the ST movement were extracted at every 30° of HT elevation  and lowering phases  of scaption. Each muscle activity was averaged for 30° increments of the scaption, using intervals of 0–30° (ele0–30°), 30–60° (ele30–60°), 60–90° (ele60–90°) and 90–120° (ele90–120°) of HT elevation, and of 120–90° (low120–90°), 90–60° (low90–60°), 60–30° (low60–30°) and 30–0° (low30–0°) lowering.Scapular kinematics was analyzed within the range of scaption 0° to 120° suggested by the validity studys), and the significance level was set at 0.05.The relationships between muscle stiffness and strength with regard to the scapulohumeral kinematics and muscle activity were examined by Spearman's rank correlation coefficient  and ele90°  , low90°  and low60°   and low60°  and negatively correlated with the posterior tilting of the scapula at low120°     and SA  with regard to the internal rotations of the scapula. The maximal strength of UT  also correlated negatively with the downward rotations of the scapula from ele30° to ele90°   , MT  and LT  but not in SA. Muscle strength not only affected the activity of the muscle in question, but also negatively correlated with the activity of adjacent muscles. The maximal strength of UT negatively correlated with the activity of MD , MT  and LT   and LT   and MT   and MT   and the muscle activity of the MD during scaption. MD may increase its contribution to compensate for the poor control of the scapula during arm elevation. Higher MD activity was noticed in protracted shoulder posture than in neutral shoulder posture due to the altered length-tension relationshipThe present results may only be generalized to the competitive adolescent swimmers practicing middle- and long-distance with front-crawl stroke. Swimmers using other stroke styles possibly present different patterns of stiffness and strength of periscapular muscles. In addition, these correlation results can not imply the causation among variables.This observational study found that the passive stiffness and maximal strength of periscapular muscles were associated with muscle activity and shoulder kinematics during the elevation or lowering phase of scaption in competitive adolescent swimmers. The training program for competitive swimmers with SD may consider the strength and stiffness of periscapular muscles in addition to the scapular kinematics. Future studies should investigate the effects of strengthening for imbalanced strength among periscapular muscles or relaxation for stiff muscles on scapular kinematics and neuromuscular controls in competitive swimmers.Supplementary Information 1.Supplementary Information 2.


Saddle nose deformities are typically reconstructed with cartilage grafts; however, conchal cartilage grafts are and associated with a risk of damage to the posterior auricular ligament and insufficient amounts, and costal cartilage grafts require invasive surgery under general anesthesia. We proposed a double-layer dermofat graft as an alternative to these methods.Two patients with type IV saddle nose deformity underwent reconstruction with nasal augmentation with a double-layer dermofat graft harvested from the gluteal sulcus.After operation, photogrammetric analysis demonstrated an improvement in the dorsal depression area, which corresponded to the angle between the sellion, most depressed point, and pronasale. Rhinoplasty Outcome Evaluation questionnaire was assessed.The graft was divided into 2 sections; the first section was implanted transversely into the depressed nasal framework, and the second section was inserted vertically from the nasion to the supratip break for augmentation.Both patients reported high satisfaction with the Rhinoplasty Outcome Evaluation questionnaire. The mean preoperative angle between the sellion, most depressed point, and pronasale was 157.8°, and the mean postoperative angle at 6 months was 176.9°.The simple method double-layer dermofat graft technique demonstrated excellent outcomes in saddle nose deformity correction, did not require cartilage, and was easily performed under local anesthesia. Saddle nose deformities are often corrected with autologous grafts,,5–9 such as septal, conchal, and costal cartilage and the calvarial and iliac bones, because these have the lowest infection and extrusion rates. Autologous tissue is easily harvested without critical donor site morbidity.Saddle nose deformity is characterized by poor structural support of the nasal mid-vault. It may be caused by trauma, infection, or previous septorhinoplasty surgery.; however, it is difficult to acquire adequate cartilage from these sites. Harvesting conchal cartilage is also associated with a risk of injury to the posterior auricular ligament, whereas harvesting costal cartilage is invasive, requires general anesthesia, and associated with a risk for pneumothorax and chest wall deformity.,10–12Different surgical methods are recommended depending on the severity of the deformity. Daniel and Brenner proposed that conchal and costal cartilage should be used for mild (type 2 and below) and severe (require reconstruction of the L-strut) saddle nose deformity, respectivelyDermofat is an autologous tissue that is easily harvested in large amounts. Dermofat grafts are frequently utilized in rhinoplasty procedures, but few studies have been examined their role in saddle nose deformity correction, particularly in severe saddle nose deformity.This study presented a double-layer dermofat graft technique for saddle nose deformity reconstruction.This retrospective case series examined 2 patients who underwent surgical correction of type IV saddle nose deformity with a double-layer dermofat graft at the Pusan National University Hospital. The study was approved by the Institutional Review Board of Busan University Hospital and complied with the 1975 Declaration of Helsinki.All procedures were performed by a single surgeon (H.J.S.). A skin and soft tissue envelope was elevated with a transcolumellar inverted V-shaped and bilateral marginal rim incisions. Dissection was continued in the supraperichondrial plane and extended in a cephalad direction. Through wide dissection, all structural deformities including deformed septum, upper lateral cartilages and lower lateral cartilages were exposed visually. The sizes and shape of depressed area were evaluated finally.An elliptical dermofat graft measuring 7–8 cm × 1.5–2 cm was harvested from the gluteal region. The gluteal region was sufficiently thick, and the resulting scar was easily hidden under clothes. The skin was incised until the subcutaneous layer with a no. 15 blade, and the graft was de-epithelialized to prevent epidermal inclusion cyst formation.The dermofat was divided into 2 parts, one is to correct the depressed area in transverse direction , and the other is for the additional correction of depressed area with nasal augmentation in vertical direction  sutures. The skin envelope was redraped to assess contour. The VD graft was sculpted to the desired dorsal width and height, similarly as in augmentation rhinoplasty. We targeted mild overcorrection to account for postoperative graft absorption. A 4-0 Nylon suture was placed at the cephalic end of dermofat and penetrated into nasal skin at the glabella midline through the 22G straight spinal needle. Next, the vertical dermofat was inserted into the prepared pocket with pulling out the suture. Finally, the other caudal side of dermofat was fixed with Ethilon 5-0 at the supratip area appropriately. The skin envelope was redraped to assess the final nasal shape and profile. The skin and mucosa were closes with simple interrupted sutures using 6-0 Ethilon (Ethicon) and 5-0 Vicryl. Merocele  packing was performed in both nostrils, and a Joseph dressing with a thermoplastic external splint was applied for protection.Surgical outcomes were assessed with preoperative and postoperative (at least 6 months) photographs, which were taken with a digital single-lens camera  with 2 flashlights. The dorsal depression angle corresponded to the angle between the sellion, most depressed point, and pronasale on lateral view  questionnaire assessed patient preferences, perceptions, functional outcomes, and social appearance and was utilized to measure patient satisfaction. The ROE consisted of 6 categories. Each question was answered on a scale of 0 to 4, and the total score ranged from 0 to 24.A 53-year-old woman with type IV saddle nose deformity had a preoperative dorsal depression angle of 159.6°. The patient declined reconstruction with costal cartilage but consented to saddle nose correction with dorsal augmentation with a double-layer dermofat graft. The operation was performed under general anesthesia due to endoscopic sinus surgery as a co-operation with the otorhinolaryngology department (Fig A 59-year-old woman with type IV saddle nose deformity presented with endoscopic findings of chronic inflammation and large septal perforation from long-term steroid spray use and repeated intranasal procedures. Her preoperative dorsal depression angle was 156.0°. Saddle nose correction with nasal augmentation was performed under local anesthesia, because the patient had stage 5 chronic kidney disease and refused invasive surgery under general anesthesia Fig. .No major complications, such as infection or hematoma, were observed during the 6-month follow-up period. Neither patient had cosmetic complications that required secondary surgery.The preoperative dorsal depression angles of both patients measured 159.6° and 156.0°, and the postoperative dorsal depression angles of both patients at 6 months measured 176.5° and 177.3°. There was a significant improvement in the mean degree of dorsal depression from 157.8° preoperatively to 176.9° 6 months postoperatively. Both patients had ROE scores of 24, which indicated high satisfaction.,15 Grade 3 or higher severe saddle nose deformity requires L-strut reconstruction with costal cartilage;,17 however, this requires an additional surgical site and general anesthesia. The double-layer dermofat graft technique is a promising alternative because it is less invasive and relatively simple.The surgical procedure for saddle nose deformity correction is determined by the severity of the deformity and other patient factors.Dermofat is an autologous tissue and has a relatively low risk for postoperative infection compared with artificial silicon blocks. Patients with saddle nose deformities commonly have a history of facial trauma or chronic inflammation, which increases the risk for thinning of the nasal skin or subclinical infection. Both of the patients in this study demonstrated chronic inflammation on endoscopy, but no complications, such as infection, were observed during the follow-up period.The double-layer dermofat graft technique has several advantages. First, it provided excellent esthetic results because the TD graft addressed the depressed saddle deformity, and the VD graft provided simultaneous augmentation rhinoplasty. Performing the procedure with the TD graft alone would have resulted in a wide nasal vault, which led to patient satisfaction, while the VD graft effectively narrowed the dorsal width. but the procedure often damages the posterior auricular ligament. The coronavirus disease 2019 era has made wearing masks indispensable, and insufficient structural support of the ear secondary to posterior auricular ligament injury worsens patient discomfort and causes limitations in daily life. Augmentation rhinoplasty is also difficult to perform with conchal cartilage because of inadequate tissue, whereas adequate dermofat can be harvested from a single donor site and utilized in dorsal augmentation and reinforcement of the thinned nasal skin.Second, the technique did not require conchal or costal cartilage. Grade 2 or lower mild saddle nose deformities are reconstructed with conchal cartilage,Third, the technique was less invasive, required shorter operating times, and can be performed under local anesthesia. This technique is an option for patients who cannot be cleared for general anesthesia, such as in case 2. Performing the technique under local anesthesia also reduced financial costs. Dermofat graft absorption may be reduced with the folding technique or overaugmentation. Kim et al proposed that minimal graft manipulation, meticulous tissue dissection with minimal reconstruction, and good bleeding control minimize dermofat absorption. We performed overaugmentation and meticulous dissection in both of the patients in this study, and neither required a repeat dorsal augmentation or revision rhinoplasty. If significant absorption was noted, a second-day fat graft was an option.Previous studies have shown that 15% to 45% of dermofat grafts are absorbed over time.There were limitations in this study. Relative small sample size and insufficient follow-up period of <1 year should be improved though further studies. Despite these limitations, our study provides additional surgical option for the correction of saddle noseIn conclusion, the double-layer dermofat graft for saddle nose deformity correction was easily performed under local anesthesia. It did not require cartilage grafts, which risk injury to surrounding structures, and provided excellent esthetic outcomes and high patient satisfaction.Ho Yoon Jeong was responsible for data collection, data interpretation, manuscript writing, and approval of the final version. Kyu-Sup Cho was responsible for data collection and data interpretation. Yong Chan Bae was responsible for data collection and data interpretation. Hyung Joon Seo was responsible for study conception and design, manuscript critical revisions, and approval of the final version.


Both the stent-graft and the delivery system exhibited satisfactory mechanical properties and operability. The safety and efficacy of this stent-graft and the corresponding delivery system were demonstrated in vivo. In nine canine experiments, the blood vessels of the animals implanted with the stent-grafts were of good patency, and there were no thrombus and obvious stenosis by angiography after implantation for 6 months. Furthermore, all of the nine clinical cases experienced successful implantation using the stent-graft and its postrelease delivery system, and the 1-year follow-ups indicated the preliminary safety and efficacy of the trilayer stent-graft with an asymmetric Z-wave design for interventional treatment. The aortic dissection (AD) is a life-threatening disease. The transcatheter endovascular aortic repair (EVAR) affords a minimally invasive technique to save the lives of these critical patients, and an appropriate stent-graft gets to be the key medical device during an EVAR procedure. Herein, we report a trilayer stent-graft and corresponding delivery system used for the treatment of the AD disease. The stent-graft is made of nitinol stents with an asymmetric Z-wave design and two expanded polytetrafluoroethylene (ePTFE) membranes. Each of the inner and outer surfaces of the stent-graft was covered by an ePTFE membrane, and the two membranes were then sintered together. The biological studies of the sintered ePTFE membranes indicated that the stent-graft had excellent cytocompatibility and hemocompatibility   The aortic dissection (AD) is a serious disease that can occur in any position along the aorta. Unlike an aortic aneurysm, AD is caused by some lesions in the intima of the aorta, and common risk factors include hypertension and atherosclerosis . The intThe Stanford classification is the most widely used method for the evaluation of AD disease . StanforThe conventional treatment for aortic aneurysms and ADs is surgery. Doctors need to open the patient’s chest and cease the heart beating prior to establish a cardiopulmonary bypass with an extracorporeal circulation machine. The surgeons need to find the target lesion vessel and remove it and then sew the ends of the artificial graft around the cut sites to replace the diseased aorta. This procedure has high risk and is time-consuming, and even a successful operation is frequently associated with serious complications such as heart, brain and renal insufficiency . The suret al. [et al. [In 1991, Parodi et al.  first re [et al.  reported [et al. . The ste™ and Valiant™ stent-graft (Medtronic vascular), Zenith™ TX and TX2 stent-graft  [™ is another thoracic aortic stent-graft, which is composed of an expanded polytetrafluoroethylene (ePTFE) membrane and supporting nitinol stents [Since the first implantation of stent-graft into the human descending thoracic aorta, doctors and engineers have been trying to modify the stent-graft structure to better adapt to aortic lesions anatomy of the patients , 20–22. al Inc.) . The fabl stents . The steThe interventional procedure of a stent-graft is schematically presented in Different from the common Dacron stent-graft in which a poly graft is sewed on to the stents with sutures, we aim to design an ePTFE stent-graft with a trilayer structure, in which the metal stent is sandwiched between two ePTFE membranes. This special structure is expected to prevent blood leakage from the sewing holes. Moreover, from the stent design standpoint, we designed an asymmetric Z-wave stent, which has long and short Z-waves on the greater curvature side and lesser curvature side of the stent-graft, respectively. This design as schematically presented in As a necessary and supplementary medical accessory for a stent-graft in interventional treatment, the delivery system enables loading a stent-graft in a sheath, delivering it to the target lesion and deploying it at the accurate setting position. In this study, we designed a ‘post-release’ delivery system, which can fix the proximal end of the stent-graft in the right position during stent-graft deployment. Displacement of stent-graft may occur in a challenging procedure due to the flush of the blood and complex anatomy; in contrast, we can, taking advantage of the ‘post-release’ delivery system, place the stent-graft more accurately.in vivo properties of this new type ePTFE stent-graft, nine animal studies were carried out in canine models. Key issues of the interventional device and corresponding design strategies are presented in the lower part of The proposed ePTFE stent-graft consists of two ePTFE membranes and one nitinol skeleton. The nitinol skeleton is made of nitinol wire with a diameter of 0.3–0.5 mm, in which the mass content of nickel is 54.5–57%, and the rest are titanium and trace elements. The nitinol skeleton or stent was manufactured by connecting a series of Z-shape wave rings in a welding process. The ePTFE membrane was covered on a cylinder SUS304 mold bar for 5–6 layers, then the stents were capped onto the inner ePTFE membrane surface and the outer ePTFE membrane was also covered with 5–6 layers. Finally, this assembled device was placed in an oven and underwent a heat treatment process to make the ePTFE membrane sintering. After that, the trilayer stent-graft was formed. The stent-graft detached from the mold bar and the excessive ePTFE membranes at both ends of the stent-graft were trimmed neatly.We made the characterizations of the polymer membranes from several aspects. The sintered ePTFE membrane was prepared by the method described in the supporting information.The membrane morphology was observed with a field-emission scanning electron microscope . The specimens were sputter-coated with gold under 10 mA for 90s and scanning electron microscopy (SEM) images were taken at accelerating voltage of 2 kV.The tensile testing of the ePTFE membrane was carried out with an SANS CMT4104 tester. The specimens were prepared by cutting an ePTFE membrane into strips of 15 mm × 200 mm along the direction of axial direction and circumferential direction, respectively. The ePTFE membrane was stretched at a speed of 200 mm/min on the testing machine until the strips broke. The stress-strain curves were analyzed following the conventional way .The crystalline phase structure of ePTFE was detected with X-ray diffraction  in the range of 10–80°. The ePTFE membrane was cut into a size of 2 × 2 cm and laid flat in the sample slot of the device for testing.Thermal properties of the ePTFE membrane were detected with differential scanning calorimetry . The thermal behavior of ePTFE membrane was analyzed by the DSC curve in an operated temperature range from 40°C to 400°C at a heating rate of 10°C/min.Surface wettability was also examined. Water contact angle (WCA) was tested with an optical device . Briefly, 10 µl of deionized water was dropped on the membrane surface, then an image was taken with a video camera. The WCA was determined on the screen of the monitor using the SCA 20 software.n = 5. The characterization follows the standard of acceptable cytotoxicity according to ISO 10993-5:2009.The cytotoxicity evaluation is based on a cell counting kit-8 (CCK8) assay as previously reported . In briein vitro hemolysis assay. The test method is as follows. Took 8 ml of fresh anticoagulant rabbit blood and diluted it with 10 ml of 0.9% NaCl solution. The diameter of the ePTFE membrane was 3 cm. Upon addition of 5 ml of normal saline, the system was incubated at 37°C for 30 min, and the contact area between the sample and the physiological saline was strictly in accordance with ISO 10993-12:2012. After that, 0.1 ml of diluted anticoagulant blood was added and the incubation continued for 60 min at 37°C. Next, we carried out centrifugation at 800 g for 5 min, and the supernatant was taken to measure the absorbance at 540 nm with a multifunction microplate detector . Distilled water and normal saline were used as positive and negative controls, respectively (n = 5 for each group). The characterization follows the standard of acceptable hemolysis according to ISO 10993-4:2002.We also carried out the In order to detect radial force of a stent-graft, two semicircular concave blocks were assembled onto the electronic mechanics tester, and then the stent-graft was fixed between the two blocks. Then, we compressed the stent-graft from the diameter direction by 20%, and the maximal value of compression force is defined as the radial force of the stent-graft.We specifically examined the release force and postrelease force in order to check the feasibility of our delivery system for interventional treatment. Firstly, we loaded the stent-graft in the delivery system and assembled the handle tension meter with the grip or the postrelease button of the delivery system separately. Then, the grip or postrelease button was pulled back to release the stent-graft or release the proximal end of the stent-graft. The maximal force values during the above-mentioned two processes are defined as release force and postrelease force.All protocols were approved by the Lifetech Institutional Animal Care and Use Committee. Following ISO 25539-2:2012 Cardiovascular implants Endovascular device-Part 2: Vascular stents and 3R Principle , nine adult dogs were included into the animal study of the stent-graft. After a conventional anesthesia and disinfection, the delivery system loaded with stent-graft was introduced to the animal aorta through the femoral artery under the guidance of digital subtraction angiography . When the proximal end of the stent-graft was positioned near the second branch artery of the aortic arch, the stent-graft was deployed by operating the handle of the delivery system. After implantation, the puncture sites were closed and the animals were allowed to recover.All experimental animals received postoperative intramuscular injections of ceftiofur sodium (3–5 mg/kg) for 3 days. The animals were kept for 6 months and received food and water three times daily. Oral aspirin (100 mg) was administered for 30 consecutive days.The dietary habits, health, mental status and bowel activities of the animals were observed after procedure and during a 6-month follow-up. At the end of the study point, all animals were euthanized and dissected. After the aorta with implantation device was extracted from the animal body, it was fixed with formalin and then imbedded in methyl methacrylate. Afterwards, then the specimens were sliced and stained with hematoxylin and eosin (HE).After the approval of the hospital ethics committee, nine patients with Type B ADs were recruited in the Second Xiangya Hospital of Central South University, following the informed consents of the patients and good clinical practice guidelines. Before operation, each patient under computed tomography angiography (CTA) examination to verify that the patient was suitable for stent-graft implantation. The stent-grafts were implanted into the patients under the standard interventional operation procedure. Each patient underwent CTA examination at 1, 6 and 12 months after the procedure to evaluate the ADs and stent-graft performance.The trilayer ePTFE stent-graft was manufactured by a ‘composite sintering’ process as shown in The raw material of ePTFE membranes is composed of vertically and horizontally interwoven fibers and nodes . This spPhysical and biological properties of the sintered ePTFE membranes of the stent-graft were characterized. Hm) of the ePTFE was 53.2 J/g. Based on the ΔHm of pure PTFE 71.0 J/g [Hm values. The results are consistent with the previous reports that the ΔHm of PTFE decreased with the manufacture process, such as sintering, melt shrinkage and fiber formation [The crystalline phase of ePTFE was characterized with XRD. As shown in 71.0 J/g , the ePTormation .in vitro.As shown in The stent-graft was designed with an asymmetric Z-wave nitinol stent. The mini Z-waves are, as indicated by the triangle in We also determined the radial forces of the proximal end and the distal end of the stent-graft, which read about 0.32 and 0.18 N in We particularly measured the release force and postrelease force during a stent-graft deployment with a delivery system using our designed test methods. As shown in The delivery system mainly consists of a sheath tube and an integrated handle . The steThe operational procedures to implant a stent-graft in human body with our postrelease delivery system are illustrated in The delivery system with postrelease mode is more accurate than the normal release mode of delivery system when deploying the stent-graft, as shown in The performance of the novel-designed stent-graft and delivery system were evaluated in a canine model. As shown in During the 6-month follow-up, all animals were alive and in good health condition. After 6 months, angiography was performed on all animals, and the DSA images indicated that the vessels were of patency and there was neither deformation nor fracture of the stents.All the animals were sacrificed at the end of the 6-month follow-up. The aorta with stent-graft was removed from each studied animal and fixed for histopathological inspections. The gross inspection of other important organs was also inspected. No abnormalities were found in the lungs, liver, spleen, kidneys or heart.We summarized the comprehensive performances of the delivery system and stent-graft in the animal experiments in We also examined thrombus formation on the sintered ePTFE membranes in canine models. Preparation of the ePTFE membranes and the methods of animal implantation as described in the We applied this new stent-graft system in nine clinical cases with Stanford B ADs. In particular, a typical case is shown in During the procedure, the sheath loaded with the stent-graft was delivered from the femoral artery to the target lesion in the thoracic aorta. Under the guidance of DSA, the proximal end of the stent-graft was aligned with the position near the opening of LSA branch. Then the stent-graft was deployed and the proximal end of the stent-graft was released by operating the handle. The final angiography shows that the stent-graft was well-positioned, and the FL was not visible as the intimal tear entry was occluded by the stent-graft. The detailed DSA images of the procedure for this patient are provided in the We also record the follow-ups. The CTA images for pre- and after implantation of stent-graft in the same person were reconstructed, with some results shown in We did an analysis of the data of the FL diameters preoperation and postoperation of the nine patients. The data of each patient are summarized in The statistical results of the comprehensive performance evaluation of the delivery system and stent-graft in the nine human clinical cases are shown in The follow-up imaging evaluation results are summarized in The latest decade has witnessed rapid progress in biomaterials and medical devices, in both fundamental research  and applA good stent-graft should have enough flexibility to adapt to the curved aorta anatomy, such as aortic arch, to reduce the straightening force on the proximal end and distal end during the stent-graft bending . This abin vitro (The sintered ePTFE membrane had sufficient strength to resist the blood pressure when the stent-graft was implanted in the human body . It is win vitro .The radial force of the proximal part of the stent-graft was larger than that of the distal part, and the non-uniform radial force design better meets the requirements of clinical needs. The proximal part of the stent-graft with a slight larger radial force is responsible for anchoring with the vessel wall and sealing the AD, while the distal part of the stent-graft with a slight less radical force is mainly responsible for being compliant with the aorta and relieving the stimulation to the vessel wall to avoid stent-graft-induced new entry at the distal part of the aorta . This spThe safety of the stent-graft and the good controllability of the new delivery system were confirmed from nine cases of large animal studies in canine models. All devices were successfully implanted in the accurate target positions and all animals survived during follow-ups. Histopathological inspections illustrated that the stent-graft had good conformability with the aorta . In addiThe nine clinical cases strengthened the good safety and efficacy of the stent-graft and delivery system  and 8. NWhile the limited clinical results have preliminarily indicated the promising efficacy and safety of the stent-graft system, more clinical evidence needs to be collected to verify this result. The aortic arch dissection involving the branches of the arch is still a challenge for a physician to perform a full endovascular repair in the aorta. This problem may be solved by the fenestration or chimney technique with a proper stent-graft with high performance in fatigue resistance to withstand the fenestration destruction or branch stent compression and conformability to the aortic arch.We developed a trilayer ePTFE stent-graft for interventional treatment of AD. The stent-graft with sintered ePTFE membranes sandwiched a metal stent with an asymmetric Z-wave design exhibited good mechanical and biological properties. Furthermore, we designed a delivery system with a postrelease mode, which improved the accuracy of the deployment of the stent-graft. Both the animal study and clinical trial indicated the accurate deployment of stent-graft by using the novel delivery system. The follow-ups demonstrated the safety and efficacy of the trilayer ePTFE stent-graft. The stent-graft and delivery system developed here provide an advanced medical device to treat AD patients in a mini-invasive way.REGBIO online.rbac049_Supplementary_DataClick here for additional data file.


Infectious diseases, caused by various pathogens in the clinic, threaten the safety of human life, are harmful to physical and mental health, and also increase economic burdens on society. Infections are a complex mechanism of interaction between pathogenic microorganisms and their host. Identification of the causative agent of the infection is vital for the diagnosis and treatment of diseases. Etiological laboratory diagnostic tests are therefore essential to identify pathogens. However, due to its rapidity and automation, the serological diagnostic test is among the methods of great significance for the diagnosis of infections with the basis of detecting antigens or antibodies in body fluids clinically. Epitopes, as a special chemical group that determines the specificity of antigens and the basic unit of inducing immune responses, play an important role in the study of immune responses. Identifying the epitopes of a pathogen may contribute to the development of a vaccine to prevent disease, the diagnosis of the corresponding disease, and the determination of different stages of the disease. Moreover, both the preparation of neutralizing antibodies based on useful epitopes and the assembly of several associated epitopes can be used in the treatment of disease. Epitopes can be divided into B cell epitopes and T cell epitopes; B cell epitopes stimulate the body to produce antibodies and are therefore commonly used as targets for the design of serological diagnostic experiments. Meanwhile, epitopes can fall into two possible categories: linear and conformational. This article reviews the role of B cell epitopes in the clinical diagnosis of infectious diseases. Although the mass production of vaccines and widespread use of antibiotics have decreased the burden of infectious diseases in recent decades, infectious diseases remain a major cause of death at present . In partEpitopes data have accumulated rapidly in the past decade. It is of high importance to analyze epitopes for numerous medical, immunological, and biological applications including infectious diseases’ prevention, diagnostics, and treatment ,7.This paper reviews the research progress of B cell epitope recognition and prediction, and summarizes the application of B cell epitopes in the diagnosis of infectious diseases.Epitopes (antigenic determinants) are a chemical group with special structure and immune activity on the surface of antigenic molecules, and represent an immunoreactive region on antigenic molecules. They are able to stimulate the body to produce antibodies or sensitized lymphocytes and can be recognized by the generated antibodies or sensitized lymphocytes, which typically account for only 5–7 amino acid residues, with no more than 20 amino acid residues at most. Based on their spatial structure, epitopes have been divided into continuous and discontinuous epitopes. Discontinuous epitopes, also known as conformational epitopes (CEs), are composed of a group of residues that are not contiguous in sequences, and are brought together by the folding of a protein to its native structure in space. Continuous epitopes, also known as linear epitopes (LEs), consist of a stretch of residues that are sequentially connected in the polypeptide sequence ,8. AlterThe identification of B cell epitopes holds enormous potential for disease prevention, diagnosis and treatment. This ultimately falls into two types: epitope mapping in experiments and in silico epitope prediction .As bioinformatics technology and multifarious databases have continued to mature and develop, approaches and software for epitope analysis have been massively boosted . At present, the B cell epitope is the hotspot in epitopes research, as the binding of it with BCRs can stimulate B cells to produce antibodies. Continuous studies have focused on prediction, and methods for computational prediction have also made great progress. Primary sequence-based information on antigens was used for linear epitope prediction in early in silico methods. Initially, the prediction approaches for epitopes were based on amino acid propensity scales depicting the physicochemical features of B cell antigenic determinants. The most classic and earliest prediction parameter is the hydrophilic parameter proposed by Hopp and Woods in the 1980s based on the assumption that hydrophilic amino acids, considered as potentially antigenic, are generally located on the surface of antigen molecules, while hydrophobic amino acids are embedded inside . Over tiWith the emergence of a variety of machine learning (ML) algorithms, researchers have implemented them in the field of epitopes prediction. Among ML-based methods, the fundamental processes are to first collect a comprehensive epitope dataset; then, extract the features of the antigen sequences; and finally, obtain a trained model by using ML algorithms to train on the collected sample dataset. Thus, a new epoch for epitopes prediction has been established. For example, one prediction method, named BepiPred , was firSince it is generally accepted that B cell CEs account for the majority of B cell epitopes, many studies have been focused on B cell CEs prediction in recent years. However, CEs are discontinuous, making prediction difficult. Existing methods for conformational epitope prediction can be broadly classified into four categories: structure-based methods, sequence-based methods, mimotope-based methods and antibody-based methods. CEP software was the first server, the algorithm of which utilizes the accessibility of residues and spatial distance cutoff of solvent-exposed residues, to predict B cell CEs based on the 3D structure of antigens . DiscoToSimilar to LEs prediction, the antigen sequence-based method has also been introduced into the prediction of CEs; due to the expensive and time-consuming determination process, the inadequate number of currently available structures has limited the development of structure-based methods ,25. The In addition, a combinatorial approach, also known as the mimotope-based method, has been proposed that requires both the 3D structure of the antigen and the antibody affinity peptide sequence as input . The priMoreover, there is another kind of method that takes antibody information into account. One of the earlier methods was proposed by Soga et al., in which the antibody-specific epitope propensity (ASEP) index was proposed and exploited to narrow down candidate epitope residues for individual antibodies. The advantages of it are that the paratopes are easy to predict and the ASEP index can eliminate part of the base residues . EpiPredA strategy to identify epitopes in experiments would be equally welcomed in studies of immunology. Several common methods used for the identification of B cell epitopes are introduced below.X-ray crystallography: Due to it being able to provide an unequivocal, atomic resolution picture of the antibody–antigen interaction by means of analyzing the atomic structure of the antigen–antibody complex, this method is believed to be the gold standard to identify the B cell epitopes. However, the apparatus is expensive and the operation process is much more complicated because of the need for high-purity antigens and antibodies and the final eutectic formation .Nuclear magnetic resonance spectroscopy (NMR): In a high-intensity magnetic field, the nuclei of different molecules can absorb electromagnetic radiation of appropriate frequency and produce different resonance spectra. The nuclear magnetic resonance signal of the antigen itself is different from that of the antigen–mAb complex. Recording and dealing with this spectrum can accurately determine the amino terminal sequence of antigenic epitopes. This method can quickly and accurately determine the amino acid motif of the B cell epitope. However, this technique for B cell epitopes’ identification in the conventional laboratory is limited by its complicated operation process and expensive equipment ,39,40. ISurface plasmon resonance (SPR): Incident light illuminates the prism side of the surface, in total internal reflection conditions, which generates an evanescent wave . EvanescPeptide scanning (Pepscan): According to the amino acid sequence of the specific protein molecule, consecutive overlapping peptides are synthesized; then, these peptides react with the corresponding specific antibodies one by one to determine the epitopes through enzyme-linked immunosorbent assay (ELISA), Western blotting (WB), protein chip or other immunological techniques. This technique requires a clear primary structure of the antigen, and the detection result is a linear epitope of the antigen ,47. UnfoAmino acid site-directed mutagenesis: A mutation takes place in a particular gene by using correlating techniques to produce the desired mutant protein; then, the antigenic epitopes of it can be identified by analyzing the binding of antigens and antibodies. In this method, both CEs and LEs can be studied ,49. HoweSurface display technology: Based on the distinct expression system, the technology has almost been developed for display technologies of phage, bacteria and yeast surfaces, which have similar methodology. The gene of the target fragment is introduced into the genome of the expression system, and then, the desired protein is expressed on the surface of the expression vector. Finally, the epitopes of the protein are depicted and identified using associated techniques ,52. PhagIn addition to the methods mentioned above, there are many other technologies that are applied to identify epitopes, such as mass spectrometry, chemical cleavage or biological enzymatic hydrolysis, flow cytometry peptide microarray, etc. Among methods, some can be used to identify both B cell epitopes and T cell epitopes. Moreover, the accuracy of epitopes identification has clearly improved when in combination with two or more methods. For instance, the combined application of surface display technology and flow cytometry technology can shorten the test cycle and reduce the cost; an intuitive and high-throughput transformation of the process can be achieved through flow cytometry technology. Each of the above methods has its own advantages and disadvantages; researchers can choose the most suitable method according to their own preferences and experimental conditions .Mycobacterium tuberculosis) and the use of antibiotics before culture. Fortunately, serological diagnosis can alleviate the above problems to a large extent. This is due to its reaction not being affected by the morphology and load of bacteria, as well as the fact that generally IgM reflects recent infections and IgG reflects past infections. However, the sensitivity and specificity of serological laboratory diagnosis are limited by certain factors such as cross-reaction and variation in pathogens; thus, the emergence of epitope-based serological experiments is helpful to improve the accuracy of tests. Currently, with the development of techniques for screening and identification of epitopes, the complexity and cost of the determination of epitopes have been partially reduced. Accordingly, epitopes have received more and more attention in pathogen diagnosis in recent years.The diagnosis of infectious diseases has long been a difficult problem in clinics. Particularly over the last decade, outbreaks caused by pathogens such as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), Ebolavirus (EBOV), and influenza virus have increased in frequency, emphasizing the urgency for the rapid development of diagnostics [M. tuberculosis can invade every organ of the body, with pulmonary tuberculosis (TB) being the most common. The period of isolation and culture is long, and the sensitivity of smear microscopy is low. At the end of the last century, one research study compared the genome of pathogenic M. tuberculosis, M. bovis, and Bacille Calmette–Guérin vaccine strains by whole-genome DNA microarray, and found 16 genomic regions of difference (RD) [Bacterial infection has always accounted for a relatively large proportion of infectious diseases. In recent years, the importance of epitopes in bacterial diagnosis has been proven in many studies. nce (RD) . Some ofnce (RD) . Sarcoidnce (RD) . The latnce (RD) . OverallStaphylococcus aureus is an opportunistic pathogen capable of causing a wide variety of infections, ranging from skin infections to severe invasive infections and even necrotizing pneumonia and sepsis [S. aureus, but it is time-consuming. Particularly for blood-borne infections caused by S. aureus, early administration of antibiotics has been reported to have a positive influence on survival. Therefore, rapid and inexpensive detection of S. aureus infection is a necessity in clinics. One of the directions is convenient serological tests based on epitopes. As early as the last century, experiments have shown that ELISA methods for quantitative detection of toxins have been developed by selecting epitopes of S. aureus enterotoxins to synthesize peptides that can be used to produce mAbs [S. aureus has high antigenic specificity. The covalently coupling peptide haptens, therefore, were synthesized by several scholars in view of the two specific representative polypeptide epitopes, PSau5 and PSau7. Then, a competitive ELISA was successfully established with high specificity and detectability to simplify the detection process and shorten the detection time [In addition, d sepsis ,60,61. Ruce mAbs . Besidesion time .Leptospira is double membrane bacterium whose outer membrane proteins (OMPs) are potential targets for bacterial-host interactions and are therefore the focus of current epitope studies [brucella and Borrelia burgdorferi, which have been studied carefully in recent years and have yielded good diagnostic results, which have been put into the table below  are highly infectious, causing a serious economic burden on society. Thus, early diagnosis and effective treatment are the main measures to prevent the spread and aggravation of the virus. Though it is universally accepted that the isolation and culture of viruses are the standard methods for virus identification, the limitation that viruses are difficult to culture in vitro hinders widespread application in clinics. Additionally, PCR is ever-increasingly applied in diagnosing and determining the replication status of viral infections by detecting viral nucleic acid. However, it is expensive due to its instruments and is easily contaminated. In fact, the serological test has become one of the main methods for the fast diagnosis of viral infection in laboratories because of its rapidity and simplicity. False-positives caused by cross-reaction between viruses and the insensitivity of some reactions are still weaknesses of serological methodology; thus, the in-depth study of epitopes is a suitable choice to solve these problems.Regarding viruses, the first that comes to mind is SARS-CoV-2. The SARS-CoV-2 pandemic rages on in some parts of the world, constituting a global public health emergency . It is cEBV, known as human herpesvirus 4, can lead to many diseases, including infectious mononucleosis, lymphoma, nasopharyngeal carcinoma (NPC), etc. It is a virus commonly susceptible to almost all populations and can cause serious physical damage to patients ,99,100. DENV, an arbovirus of high concern, causes a range of symptoms, from mild fever to severe hemorrhagic fever, and even shock. Its non-specific clinical symptoms, especially the early mild fever, prevent timely and accurate diagnosis of dengue infection, eventually resulting in the progression of infection . NeverthViral hepatitis, caused by a group of viruses that primarily attack the liver, is a major global public health problem. Hepatitis B virus (HBV) and hepatitis C virus (HCV) are also major contributors of leading to chronic infections and cirrhosis. The serological test has long been a mainstay of viral hepatitis testing, an important method for the clinical judgment of infection status is to detect the antigen or antibody. Even if available commercial kits have been widely applied in medical settings, it is still the continuous pursuit of the laboratory to improve the accuracy of detection. The value of epitopes has gradually been uncovered in the diagnosis of hepatitis virus. For the detection of anti-HBc antibodies, a study selected HBcAg epitopes to design a multi-epitope recombinant protein, being stable and inexpensive, whose detection ability was validated to be consistent with commercial antigens . AdditioFiloviruses cause rare but deadly viral hemorrhagic fevers—EBOV and Marburgvirus (MBV) are two formidable agents among them. Even though the detection of RNA is more accurate, the two viruses are mostly spread in regions with limited resources, where local laboratories are lacking the relevant equipment. Consequently, rapid diagnostic tests (RDTs) are critical for early detection of them. The EBOV glycoprotein (GP) mediates virus attachment and entry into host cells, while nucleoprotein is abundant in virus particles, both of which can induce strong immune responses and, therefore, are ideal antigens for vaccine and diagnostic studies. Many studies have identified the epitopes of these proteins, providing data for the development of Ebolavirus detection reagents ,122,123.(G72-D110) and SHNP(P251-D264), which were highly conserved among species associated with HFRS—were found and validated in experiments. They are expected to be the diagnostic epitopes for HFRS-associated orthohantaviruses [Orthohantaviruses cause hemorrhagic fever with renal syndrome (HFRS) and orthohantavirus cardiopulmonary syndrome (HCPS). It is difficult to distinguish HFRS and HCPS from other tropical hemorrhagic diseases when relying solely on clinical manifestations, so serological assays are still necessary diagnostic tests. Whether HFRS or HCPS will appear after infection depends on the viral genotype and the reservoir species of different locations. The nucleocapsid proteins (NPs) have been found to be highly conserved among different hantavirus genotypes and are thus considered to be the ideal protein for diagnosing hantaviruses of different genotypes. In 2019, a publication reported that the Seoul orthohantavirus nucleoprotein (SHNP) sequence was used as a model to predict the linear B cell epitopes on SHNP and its immunogenicity was evaluated by in silico methods. In the end, two of nine potential immunogenic linear B cell epitopes—SHNPaviruses . In addiaviruses . Anotheraviruses . All of Toxoplasma gondii is capable of infecting an extremely wide range of mammals and birds, and is a major opportunistic pathogen with a high mortality rate in immunocompromised individuals. To date, the serological test remains the primary method to characterize Toxoplasma gondii identification, while most diagnostic kits use crude Toxoplasma antigens, with difficulties such as standardization and cost-effectiveness. A synthetic gene, named USM.TOXO1, encoding nine immunodominant epitopes of T. gondii antigens, was constructed by some researchers who cloned it into a pET32a expression vector. Then, a purified antigenic protein was obtained with 85.43% and 81.25% for diagnostic sensitivity and specificity, respectively, in IgG ELISA [Leishmania is the causative agent of a complex neglected tropical disease—leishmaniasis—and is widespread in a large number of areas [Leishmania, the most reliable and classical method relies on the microscopic visualization of the amastigote form of the parasite from tissue specimens after Giemsa staining, the sensitivity of which depends on the tissue [Leishmania infection have a promising future. Schistosomiasis remains a serious public health problem, the causative agent of which is the genus Schistosoma. One of the essential means to effectively control and monitor schistosomiasis is to select diagnostic methods for early diagnosis [S. japonicum, S. japonicum saposin protein SjSAP4. Meanwhile, an epitope of the protein has also been identified. Although the sensitivity of the epitope is inferior to the antigen protein by comparison, the specificity of the epitope was found to be 100%, and the result of dual-epitope ELISA showed superior diagnostic performance than the single-epitope version. Considering the complex immunology and host–parasite interactions, the role of epitopes in the diagnosis of schistosomes can be made more significant with a rational choice of epitopes [Most parasitic diseases are diagnosed on the basis of finding parasite eggs or parasite bodies, but the serological test is still an important part of the diagnostic process for parasitic diseases. Some examples of epitopes being used in the diagnosis of parasitic diseases are described below. gG ELISA . FurthergG ELISA ,130. In of areas ,132. It of areas ,133. Fore tissue . The sere tissue . Furthere tissue ,137,138.iagnosis ,140. Intepitopes .  and secreted aspartic protease (Sap) are two immunodominant antigens of C. albicans. Phage display technology is used to display epitopes of these proteins, but the study found that the diagnostic performance of a single epitope is not as good as its recombinant protein, which may be due to several epitopes in the recombinant protein. However, phage display technology is simpler and cheaper than traditional methods of chemical synthesis or separation and purification of antigens [Paracoccidioides brasiliensis [C. albicans [Cryptococcus spp. [Pneumocystis jirovecii [In addition, some studies directly selected a single epitope in the antigen for testing, such as antigens ,166. Similiensis ,168, oneiliensis . There aalbicans , Cryptoccus spp. , and Pneirovecii ,173.The culture examination of clinical specimens is usually used as the standard diagnosis of systemic mycosis, and the detection of fungal antigen or antibody is also one of the means for early diagnosis of invasive fungal infection, but for most fungal diseases, it is often unavailable . All in Serology remains an important diagnostic tool for infectious diseases, but an open question is a lack of sensitivity and specificity for the detection of certain diseases when using recombinant proteins as antigens. The existence of cross antigens is the main reason. As the smallest immune unit to stimulate the immune response, epitopes cannot be overestimated in the study of their structure and function. Given the specificity of epitopes, the search for epitope-based vaccine development, disease diagnosis and therapy has been ongoing for decades. Before, B cell epitopes were studied more compared with T cell epitopes for the serology tests, because the former are involved in humoral immunity. Moreover, although X-ray crystallography and NMR techniques, used in earlier years, are still reliable methods for determining epitopes at present, they are time-consuming, laborious, and costly . CarefulThe work on applications of epitopes in the diagnosis has accelerated with the help of immunoinformatics technology. The use of epitopes in serological diagnostic tests aims to the improvement of detectability, sensitivity and specificity. Therefore, theoretically, many problems have the opportunity to be solved by the rational design of epitope combinations. The selection of multiple strongly immunogenic epitopes may enhance the sensitivity and specificity to distinguish between positive and negative sera . In addiAs described before, the application of epitopes in diagnosis has a few advantages, but a number of challenges remain. Firstly, the performance of existing B-cell epitope prediction methods is not satisfactory that may be related to the datasets, features and classifier, which indicates that methods need to be further improved to acquire a more accurate result . SecondlIn conclusion, although research on antigenic determinants has obtained encouraging results, there are still many difficulties ahead of us. It is suggested that with the progress of technologies and the continuous efforts of scientists, epitopes will be more suitable to apply in the diagnosis of infectious diseases.


Dpe2/phs1 has been reported to have only one starch granule number per chloroplast when grown under diurnal rhythm. For this study, we analyzed dpe2/phs1 in details following the mutant development, and found that it showed three distinct periods of granule numbers per chloroplast, while there was no obvious change observed in Col-0. In young plants, the starch granule number was similar to that in Col-0 at first, and then decreased significantly, down to one or no granule per chloroplast, followed by an increase in the granule number. Thus, in dpe2/phs1, control over the starch granule number is impaired, but it is not defective in starch granule initiation. The data also indicate that the granule number is not fixed, and is regulated throughout plant growth. Furthermore, the chloroplasts revealed alterations during these three periods, with a partially strong aberrant morphology in the middle phase. Interestingly, the unique metabolism was perpetuated when starch degradation was further impaired through an additional lack of Isoamylase 3 (ISA3) or Starch excess 4 (SEX4). Transcriptomic studies and metabolic profiling revealed the co-regulation of starch metabolism-related genes and a clear metabolic separation between the periods. Most senescence-induced genes were found to be up-regulated more than twice in the starch-less mature leaves. Thus, dpe2/phs1 is a unique plant material source, with which we may study starch granule number regulation to obtain a more detailed understanding.An Arabidopsis mutant lacking both the cytosolic Disproportionating enzyme 2 (DPE2) and the plastidial glucan Phosphorylase 1 (PHS1) revealed a unique starch metabolism.  Each chloroplast in Arabidopsis thaliana (L.) Heynh. wild-type leaves contains three to seven discoid starch granules, and the starch stored during the day is nearly completely depleted by dawn  have a lower starch degradation rate , for which an uneven starch distribution has been revealed, with nearly starch-free mature leaves and young leaves having mostly one large and spherical starch granule per chloroplast when grown under circadian periodicity  in dpe2/phs1/sex1-8 , accumulated a high amount of maltose inside the chloroplasts and showed severe chloroplast degradation in mature leaves (pgm1 (Phosphoglucomutase 1 [EC 5.4.2.2]) and adg1 (ADP-glucose pyrophosphorylase 1 [EC 2.7.7.27]) revealed higher Rubisco-containing bodies, leading to higher percentages of aberrant chloroplasts  medium with 0.8% [w/v] agar and 1% [w/v] sucrose, pH 5.7. The plates were vernalized in darkness at 4°C for three days. Seedlings were transplanted to soil after 7–10 days and grown in a light/dark regime .dpe2/phs1 was addressed previously (The crossing of mutant eviously . To deteSex4-3 (SALK_102567) (isa3 (GABI_280G10)  , and isa_280G10)  in the CThe activities of phosphorylases and DPE2 were detected by native gels and zymograms, as described previously . AnalysiLCSM imaging was performed as described by Shoots were stained as described by Starch was enzymatically quantified according to Starch granules (10 mg) were used and digested as described by For quantitation of the maltodextrin, 100 mg of leaves were frozen in liquid nitrogen and homogenized with a mortar. Then, 500 µL cold 20% [v/v] ethanol was added to each sample and mixed by vigorously shaking. After centrifugation at 4°C, 14,000 rpm for 10 min, the supernatant was collected and heated to 99°C for 10 min. Followed by centrifugation, the supernatant containing the maltodextrin was recovered and concentrated by drying in a speed vacuum centrifuge (Thermo Savant SPD111V-115).The starch chain length distribution and the maltodextrins were analyzed by capillary electrophoresis equipped with laser-induced fluorescence detection (CE-LIF), as described by dpe2/phs1 and Col-0 were harvested after 9 h illumination for each period. Total RNAs were extracted using TRIzol reagent , according to the manufacturer’s instructions. A NanoPhotometer® spectrophotometer was used to assess the purity of the RNA . The Bioanalyzer 2100 system’s RNA Nano 6000 Assay Kit was used to examine RNA integrity and for quantification . Library construction and sequencing with an Illumina novaseq 6000 sequencer were performed by Novogene Corporation (UK). Annotation files for the reference genome and gene models were downloaded directly from the genome website browser (NCBI/UCSC/Ensembl). The HISAT2 software was used to map paired-end clean reads to the reference genome. The collected RNA-seq data were deposited in the NCBI GEO repository, under the accession number GSE201804. The DESeq2 R software was used to perform differential expression analysis between two conditions/groups . The False Discovery Rate (FDR) was controlled by adjusting P-values using Benjamini and Hochberg’s method. Genes discovered by DESeq2 with an adjusted P-value < 0.05 were labeled as differentially expressed. The statistical enrichment of differentially expressed genes in KEGG pathways was tested using the clusterProfiler R program.The leaves of The RNAs isolated with TRIzol reagent  were reverse transcribed into cDNA using a first cDNA synthesis kit , following the manufacturer’s instructions. qRT-PCR was performed with SYBR Green PCR Master Mix (Applied Biosystems), following the manufacturer’s instructions. Reactions were carried out in triplicate. Relative expression levels were determined by the comparative threshold cycle (ΔΔCt) method, using ACTIN as a reference gene. The used primers are given in The same leaf materials from the three periods (50 mg each) used for RNA-seq were used for metabolite profiling, using Gas Chromatography-Mass Spectrometry (GC-MS). The following data analysis was conducted according to dpe2/phs1 has been described as having a single starch granule per chloroplast in immature leaves, with virtually no starch in mature leaves when grown under a normal light/dark cycle  by the end of the day (EOD), and consumed almost all of it by the end of the night (EON) Table 1.dpe2/phs1 in all three phases (Dpe2/phs1 showed the largest long axis and thickness in young leaves in RGN (Scanning electron microscopy (SEM) revealed that the starch granule morphology was strongly altered in One way to investigate the reasons underlying the variation in the starch granules through the three phases is to analyze the associated gene expression by RNA-seq. Therefore, leaves from P, where the chloroplasts contained the normal starch granule number; young Y and mature M leaves from RGN, where most of the chloroplasts had only a single, large, and round starch granule and most chloroplasts were aberrant, respectively; and leaves from R, where the chloroplasts and starches recovered, were analyzed. As a control, Col-0 samples of the same age were used.P-value < 0.05) revealed only a moderate number of genes specifically expressed in the various phases , galactose (ath00052), fructose and mannose (ath00051), pentose phosphate (ath00030), pentose and glucuronate interconversions (ath00040), inositol phosphate (ath00562), citrate cycle (ath00020), glycolysis (ath00010), and oxidative phosphorylation (ath00190). Overall, 720 genes were assigned to six clusters . The higher expression level of genes involved in synthesis may reflect an attempt to overcome the limitation in starch granule number observed in young leaves in RGN.Interestingly, most genes involved in starch initiation, including SS4, SS3, PTST, PTST2, PTST3, MFP1, and MRC, also revealed a higher expression in Y . ConsideMoreover, the homologous genes did not follow the same clustering; for instance, starch synthases SS2–5 and GBSS1 increased from P to young leaves in RGN, but not SS1. However, the observed alteration was in agreement with the chain length distribution profile of the starch granules, as fewer short chains (related to SS1) and more medium–long chains (related to SS2) were observed , an important phosphatase in the phosphorylation/dephosphorylation cycle at the starch granule surface, and Isoamylase 3 , a hydrolase involved in starch degradation, both genes revealed an increase in Y. Both triple mutants revealed reduced shoot growth and uneven starch distribution, in addition, a decreased starch granule number, similar to that of dpe2/phs1 rather than Col-0  and Disproportionating enzyme 1  it was not possible to fully recover the wild-type starch phenotype in the background of the dpe2/phs1 mutant  by comparing P with Y, and Y with R, respectively. GSEA between P and Y showed that 111 gene sets were enriched, with 62 sets up-regulated and 49 sets down-regulated in P. Nine gene sets were significant at FDR < 25% Table S2mex1 mature leaves  were observed in all periods in dpe2/phs1, except a lower DP 11 in R , allowing for determination of the starch granules in leaves in different periods. Previously, dpe2/phs1 has been reported as having a severely impaired growth rate and heterogeneous starch distribution, with a single starch granule per chloroplast in young leaves and almost starch-less mature leaves when grown under diurnal rhythm. Our investigation revealed that starch metabolism in dpe2/phs1 is inhomogeneous not only among leaves, but also during development. Thus, three clearly distinguishable periods were described. By analyzing relevant leaf material from these three stages through RNA-seq and metabolic assays, we determined the co-regulated genes involved in starch metabolism, showing a clearly separated metabolism, suggesting a regulation of the starch granule number throughout the plant growth. This unique plant material, thus genetically identical—as is not the case when working with mutants—allowed us to understand the important step within the starch metabolism, regulation, and initiation of starch granules in Arabidopsis leaves.This study was inspired by a detailed investigation of dpe2/phs1 was homogeneous in both starch distribution and starch granule number over the entire growth period  may reduce chloroplast degradation. Thus, it has been reported that the total blocking of starch degradation due to a lack of GWD resulted in the complete restoration of chloroplasts  can be found in the article/XL and JF conceived and designed the experiments; XL, AA, JFC, JC, and SM conducted the experiments; XL and JF wrote the manuscript; JF revised the manuscript. All authors read and approved the final manuscript.This work was supported by the Deutsche Forschungsgemeinschaft DFG-FE 1030/5-1, and 6-1.This research was further supported by INST 336/114-1 FUGG for large equipment, granted to Prof. Markus Grebe and co-applicants.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. 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In this review paper, graft failure rates of different graft types  that are used for surgical reconstruction of the anterior cruciate ligament are compared and statistically analysed.Literature search was conducted in PubMed according to Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) criteria. A total of 194 studies, which reported graft failure rates of at least one of the anterior cruciate ligament reconstruction methods mentioned above, were included in this systematic review. To be able to compare studies with different follow-up periods, a yearly graft failure rate for each reconstruction group was calculated and then investigated for significant differences by using the Kruskal–Wallis test.Overall, a total of 152,548 patients treated with an anterior cruciate ligament reconstruction were included in the calculations. Comparison of graft types showed that hamstring tendon autografts had a yearly graft failure rate of 1.70%, whereas the bone–patellar tendon–bone autograft group had 1.16%, the quadriceps tendon autograft group 0.72%, and the allografts 1.76%.The findings of this meta-data study indicate that reconstructing the anterior cruciate ligament using quadriceps tendon autografts, hamstring tendon autografts, patellar tendon autografts or allografts does not show significant differences in terms of graft failure rates.The online version contains supplementary material available at 10.1007/s00402-021-04147-w. Isolated tear of the anterior cruciate ligament (ACL) is a common orthopaedic injury, as the annual incidence of 68.6 per 100,000 person-years of the US population shows . Recent Several factors can influence the outcome of an ACLR and should therefore be taken into consideration when graft survival is investigated. Age has been identified as a risk factor for ACL graft rupture. The odds of an ACL graft rupture decreases with every yearly increase in patient age . HoweverIn recent traumatology surgery, several ways of reconstructing a ruptured ACL were established. In a global perspective, the most common grafts used for reconstruction of the ACL are hamstring tendon autografts (HTA), bone–patellar tendon–bone autografts (BPTBA), different allografts and quadriceps tendon autografts (QTA), respectively. As a matter of fact, single-bundle reconstruction is used more frequently than double-bundle technique. The purpose of this systematic review was to compare graft failure rates of different graft options used for ACLR by taking the most recent studies with a very high volume of patients into calculation. Such an approach to this topic has not yet been chosen in recent literature. A recent review paper including far less trials than our systematic review suggested that there is no significant difference between autograft subgroups in terms of graft failure rates . BetweenA systematic literature search on common methods for ACLR using autografts and allografts was conducted in PubMed based on the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) criteria. Search was performed throughout May 2020 using the following terms for search: “ACL reconstruction” in combination with “quadriceps tendon” OR “quadriceps graft” OR “hamstring” OR “bone tendon bone” OR “patellar tendon” OR “allograft” in <All Fields> .All obtained papers’ titles and abstracts were screened, and relevant studies were identified. To further assess these studies for eligibility, certain selection criteria were defined.Publication date within the last 10 years (May 2010–May 2020).Trials only conducted on human species (level of evidence I-IV).A follow-up (FU) period of at least 2 years.Isolated ACL rupture as primary ligamentous injury with or without meniscal damage.Study population with a minimum mean age of 18 years.All tibial and femoral graft fixation techniques.Studies including transtibial or anteromedial tibial tunnel drilling.All kinds of hamstring techniques (any number of strands).All kinds of sterilisation and preservation methods for allografts.Inclusion criteria:Studies published in languages other than English.No documentation of graft failure or re-rupture rate.Revision ACLR.Studies investigating only highest-risk populations such as professional athletes or skeletally immature.All double bundle or hybrid graft techniques.Multiple ligamentous injuries of the knee joint.Follow-up duration unavailable or only available as median.Exclusion criteria:A total of 3895 citations were found, and 194 studies were selected to be eligible for the systematic review according to the inclusion and exclusion criteria, as Fig. To create homogenous data about study characteristics and patient demographics, information was collected by systematically extracting year of publication, authors, study design and level of evidence as well as number of subjects, mean age, mean follow-up time, cases of graft failure and used grafts from the studies.From this dataset, the main patient demographics containing number of operated patients, the mean age and mean follow-up time were elaborated. These main patient demographics are displayed in Table The methodological index for non-randomized studies (MINORS) was used to assess methodological quality of both non-randomized comparative studies and non-randomized non-comparative studies that were included in this systematic review . The MINThe quality of the included randomized controlled trials was assessed using the modified Jadad scale . The oveFor statistical calculation of the results, IBM SPSS Statistics 23  was used. At first, descriptive statistics were carried out: the number of patients included, the average patients’ age at surgery and overall mean follow-up was calculated per graft type and for the overall study population.p value of less than 0.05.The follow-up duration of the 194 studies, which were included in this systematic review, ranged from 2 years up to 25 years. A yearly graft failure rate (percentage) was calculated for each graft type to make studies with different follow-up periods comparable. This variable (yearly graft failure rate) was then examined for normal distribution using the Kolmogorov–Smirnov test. To examine if the grafts differ in yearly graft failure rates, the Kruskal–Wallis test was conducted. In this testing method, the null hypothesis assumed no significant difference in yearly failure rates between the reconstruction groups. The level of significance was set at a 194 studies of different study types and different levels of evidence (I–IV) were included in this systematic review. Some of the studies included more than one of the four graft types, resulting in 296 cohorts in the dataset, each cohort containing information about one of the reconstruction methods. Of these 296 cohorts, the distribution to graft types was 148 (50.0%), 87 (29.4%), 12 (4.1%) and 49 (16.5%) for hamstring autograft, bone–patellar tendon–bone autograft, quadriceps autograft and allografts, respectively . 173 studies were assessed this way. 81 of them were regarded as high-quality studies.The included randomized controlled trials  reached an average of 6.4 out of 8 points (80% of maximum). All of the 21 assessed randomized controlled trials were regarded as high-quality trials. Detailed tabular results of the methodological quality assessment, study type and level of evidence of the included studies can be found in the appendix.p value of 0.072 which is considered as not significant (p > 0.05). Thus, the yearly graft failure rate of the four investigated ACLR groups did not differ significantly and the null hypothesis was retained.The Kruskal–Wallis test of yearly failure rates for the four main graft groups showed a The mean of the yearly failure rate was calculated for each graft type and is displayed in Table In terms of follow-up time, the BPTB autograft group had an average of 86.9 months follow-up duration, the hamstring autograft group 53.3 months, the allograft group 47.5 and the QTA group was followed up with a mean of 36.7 months Table .The most important finding of this systematic review is that all types of grafts that are commonly used for ACLR did not differ significantly in terms of yearly graft failure rates. These main findings confirm the results of a recent meta-analysis by Mouarbes et al. in which HTA, BPTBA and QTA were compared and no significant difference in graft failure rates could be found .Recent findings of Nyland et al. displayed that QTA produces better results in terms of graft failure rates than other common grafts . HoweverFurthermore, a lot of quality research has been done on the comparison of graft failure rates of HTA with BPTBA in the last decade . Graft fGraft failure may be the most important indicator for measuring success of an ACLR, but there are still other considerable factors such as harvest site pain and functional results. A well-known downside of BPTBA is that anterior knee pain and kneeling pain often occur due to the harvest site defect . This esIn terms of functional outcomes and harvest site morbidity, the QTA is regarded as a promising alternative for ACLR compared to current gold standard methods (HTA and BPTBA). Slone et al. stated that stability outcomes , functional outcomes , range of motion, overall patient satisfaction and complications were similar when comparing quadriceps tendon autograft to other graft options –24. ContIn addition, graft selection should always be done thoroughly considering the patient’s age, needs, activity level and concomitant injuries. By implication, having many valid graft options available is advantageous and surely does enrich the quality of ACL surgery as all of them deliver predominantly satisfying results.autograft with Quadriceps tendon allografts [The promising results achieved by quadriceps tendon autograft inevitably lead to the implication that the quadriceps tendon could be used for allograft ACLR as well. Up to now, hardly any high-quality studies including the QT allografts have been carried out. Kwak and colleagues published a matched case control study comparing Quadriceps tendon lografts . This stThere are several limitations to this study. Firstly, different study types of different levels of evidence (I-IV) were included and equally weighted in this review. Overall, 102 of 194 included studies were high-quality studies. Secondly, the studies that were included in the present systematic review contained different graft fixation methods such as suspensory fixation methods, fixation with interference screws or a combination of both methods. Thirdly, the cohort size of the included studies varied from 8 to 17.096 patients (e.g. nationwide cohort studies). Furthermore, the definition of graft failure was inconsistent as some studies defined the need for ACL revision as graft failure, while others considered pathological magnetic resonance imaging (MRI) or clinical deficits as graft failure. Another important aspect that should be kept in mind in this respect is the question whether traumatic re-rupture of the reconstructed ACL should be considered as graft failure or re-injury . As a fiAs a matter of fact, this systematic review has certain strengths. To our knowledge, this is by far the largest systematic review paper: a great number of studies (194) and ACL reconstructions  were included. Only studies published in the past 10 years were selected for this review in order to ensure actuality of surgical procedures to a certain extent. By calculating a yearly graft failure rate, the results of 194 different clinical studies were combined and compared in an effective and comprehensible way.The findings of this extensive systematic review showed no significant differences in yearly graft failure rates of HTA, BPTBA, QTA and allografts. Based on these data, all these graft options deliver comparable results in terms of graft failure rates and therefore every graft type could be rightly considered as reliable option for ACLR. An increased use of QTA that will ultimately be documented in publications of more long-term studies will show whether a positive or negative impact on failure rates of this graft can be detected.Supplementary file1 (XLSX 32 KB)Supplementary file2 (SAV 14 KB)Below is the link to the electronic supplementary material.