The results of Fourier transform infrared spectroscopy (FT-IR) and X-ray photoelectron spectroscopy (XPS) confirmed successful grafting. The grafting ratios of MAH-g-starch, LA-g-starch, and MA-g-starch were 6.50%, 12.45%, and 0.57%, respectively. Influenced by the grafting ratio, LA-g-starch had the best relative hydrophobicity and the largest molecular weight, and those for MA-g-starch were the worst. The surfaces of grafted starches were covered with graft polymer, with obvious surface roughness and bond degree of MAH-g-starch and LA-g-starch. The crystalline structure of grafted starches showed some damage, with LA-g-starch exhibiting the greatest decrease in crystallinity, and less of a change for MA-g-starch. Overall, the grafting reaction improved thermoplasticity, with LA-g-starch the most improved, followed by MAH-g-starch, and then MA-g-starch.Three kinds of hydrophobic groups grafted starches of maleic anhydride grafted starch (MAH- In recent years, with increasing environmental awareness and the gradual consumption of fossil resources, there has been significant interest in the development of green and renewable biomass resources as a substitute for petrochemical products and as alternate energy sources. Starch is a natural polymer material with many advantages such as a wide variety of sources, low price, biodegradability, and renewability ,2,3. ThuOne strategy for modification is the use of hydrophobic groups to replace hydrophilic hydroxy groups on the starch molecule chain ,9. To doHydrophobic groups graft starches are usually produced by the wet method, organic solvent method , reactivConsidering the limitations of the current technological methods used to graft hydrophobic groups to starch, the objective of this investigation was to produce hydrophobic groups grafted to starches by an in situ solid phase polymerization method. In this process, dry starch, reaction monomers, and a catalyst are mixed in a closed hydrothermal reactor, and in situ polymerization of the starch and reaction monomers is initiated under pressure at 80 °C . This apAnhydrides, carboxylic acids, and acrylate compounds are commonly used in hydrophobic modification of starch and interfacial modification of starch/polymer matrix composites ,20,21,22Corn starch was obtained from Dacheng Corn Development Co. Ltd. , and dried in a vacuum drying oven of 50 °C for 48 h to eliminate moisture before use. Methyl acrylate was obtained from Tianjin Kwangfu Fine Chemical Industry Research Institute . Lactic acid and stannous octoate were supplied by Sinopharm Chemical Reagent Co., Ltd . Maleic anhydride, acetone, and ammonium persulfate were purchased from Tianjin Kemiou Chemical Reagent Co., Ltd. . All chemicals were AR grade.Thirty grams of corn starch were added into a hydrothermal reactor, together with 4.5 g grafting monomer and 0.9 g stannous octanoate catalyzer, and mixed and ground evenly. The hydrothermal reactor was sealed and placed in an oven at a set temperature  for in situ solid phase polymerization. Heating was stopped after a set time . When the reacted starches were cooled to room temperature, acetone was added to the mixture. The mixture was stirred and subjected to suction filtration. Finally, the mixture was washed three times with acetone and dried in an oven at 55 °C until a constant weight was achieved.At the same time, in order to compare the grafting effect of in situ solid phase polymerization, the graft copolymerization of starch was carried out by aqueous phase polymerization and organic solvent polymerization. Deionized water was the solvent in aqueous phase polymerization, and tetrahydrofuran was selected by organic solvent polymerization. Starch and solvents were mixed into starch emulsion according to the mass ratio of 1/9, and protection of flushing nitrogen into reaction system. The reaction temperature was 80 °C, the reaction time was 2.0 h, and the catalyzer and treatment of grafted starches were consistent with in situ solid phase polymerization.−1.In order to characterize the chemical changes in the grafted starch, the samples were tabletted with KBr and subjected to FTIR . To completely remove the moisture, the native starch and hydrophobic group grafted starch materials were further dried in a muffle oven (at 50 °C) for 48 h. The tested samples were obtained after grinding fully using a weight ratio of sample: KBr of 1:100. The FTIR curves for the samples were obtained at a range of 400–4000 cm3d5/2 line, these conditions produced an FWHM of 0.80 eV. XPS measurements were performed at room temperature with monochromatic AlKα radiation (1486.6 eV) using a K-Alpha X-ray photoelectron spectrometer . The X-ray beam was a 100 W, 200 mm-diameter beam raster over a 2 mm by 0.4 mm area on the sample. A high-energy photoemission spectrum was collected using a pass energy of 50 eV and resolution of 0.1 eV. For the AgGR) was calculated as follows:W is the substitution degree of the hydrophobic group, %; M is the molecular weight of the graft monomer; c is the concentration of the aqueous hydrochloric acid solution, mol/L; V0 is the consumed volume of aqueous hydrochloric acid solution by the blank sample, mL; V1 is the consumed volume of aqueous hydrochloric acid solution by the grafted starch sample, mL; n is the number of hydrophobic groups from the grafted monomer; and m is the mass of the sample, g. First, 1.00 g of dry grafted starch was weighed and placed in a 250-mL conical flask. Next, 10 mL of 75% ethanol solution in deionized water were added to the flask, followed by the addition of 10 mL of 0.5 mol/L aqueous sodium hydroxide solution. The stoppered conical flask was agitated, warmed to 30 °C, and stirred for 1 h. The excess alkali was then back-titrated with a standard 0.5 mol/L aqueous hydrochloric acid solution. A blank titration was performed using native, un-modified starch. The grafting ratio  was used to measure the contact angle of samples, using distilled water as the test solution. For each measurement, 4 µL liquid in a microsyringe were dripped on to the surface of the samples, and the contact angle values were measured until 0°.tW was the weight of the sample after water absorption for t hours and W0 was the weight of the sample when it reached a dry constant weight. To determine the water absorption of the grafted starches, 4.0 g native starch and grafted starches (dry base) were separately placed in glass dishes that contained a set amount of water. Over the test period the weight of each sample was measured every 12 h. The water absorption was calculated as follows:g-starch, LA-g-starch, and MA-g-starch were tested with a gel permeation chromatograph system . The solvent was dimethyl sulfoxide (DMSO) + 20 mmol LiBr, the column set was IGuard + 1 × I-H, the flow rate was 0.500 mL/min, the injection volume was 100 uL, and the detector and column temperature were 50 °C.The molecular weight of native starch, MAH-The hot paste viscosities of the native starch and grafted starches were measured with a Rotational Viscometer . Starch slurry with a mass percentage of 6% was prepared by dispersing starch (4.2 g dry basis) in 65.8 g distilled water and then heating in a 95 °C constant-temperature water bath. The rotational viscometer was connected to the thermostat and gradually increased to 95 °C, then maintained for 15 min. The viscosity value was recorded when the viscometer became stable.The morphology of the native starch and grafted starches was determined with a scanning electron microscope , operating at an acceleration voltage of 20 kV. Starch granules were mounted on circular aluminum stubs with double-sided adhesive tape and coated with gold before testing.The native starch and grafted starch samples were further dried in a vacuum oven at 50 °C for 48 h to remove the remaining moisture. The crystallinity index of the samples was measured by an X-ray diffractometer  with Cu target at 36 Kv and 20 mA. Samples were tested in the angular range of 2θ = 5–40° with a scanning rate of 4°/min.TGA measurements of native starch and grafted starch samples were made with a 209 F3 TGA instrument . About 5 mg of dried sample powders were placed in a platinum crucible and heated from 25 to 800 °C at the rate of 10 °C/min. Dynamic carrier nitrogen gas flowed at a rate of 30 mL/min. Thermogravimetric (TG) and derivative thermogravimetric (DTG) data were obtained for each sample.The gelatinization temperature of NS, DS and ATDS were studied by using a Differential Scanning Calorimeter , as described by Sandhu and Singh . Starch p < 0.05).The data in the current study were statistically evaluated using the statistical software package Minitab Version 15  and reported as the mean value ± standard deviation of the replicates. A single-factor analysis of variance was conducted to identify significant differences among mean values according to least significant difference criteria with a 95% confidence level  to establish the binding modes of C in the samples; the results are shown in g-starch b, LA-g-sg-starch c, and MAg-starch d shows tThe reaction variable of the in situ solid polymerization reaction between native starch and hydrophobic monomer was directly related to the variety of hydrophobic monomer, and inevitably leads to the difference in the grafting ratio. At the same time, the graft copolymerization of starch was carried out by the aqueous phase polymerization and organic solvent polymerization methods, and the grafting ratio has been tested; the results are shown in As shown in g-starch was 6.50% ± 0.42%, that of LA-g-starch was 12.45% ± 0.37%, and MA-g-starch was 0.57% ± 0.05%. The results show that the efficiency of in situ solid phase polymerization for lactic acid and starch was the highest under these conditions. The XPS analysis showed that the C=O content of LA-g-starch was the highest and that of MA-g-starch was the smallest. The in situ solid phase polymerization of lactic acid and maleic anhydride with starch occurred as esterification, while methyl acrylate occurred as a radical polymerization. The reaction conditions required for radical polymerization are more stringent, resulting in the lowest grafting ratio for MA-g-starch. Lactic acid is a liquid that can penetrate into the starch well before the reaction, increasing the chance of contact with hydroxyl groups on the starch. In contrast, maleic anhydride is solid, and does not penetrate into the starch to react with hydroxyl until the temperature exceeds its melting point (52.8 °C). Additionally, maleic anhydride needs to open the acid anhydride to react with hydroxyl groups. These two reasons cause the grafting ratio of the LA-g-starch to be significantly greater than that of the MAH-g-starch.With the in situ solid phase polymerization, the grafting ratio of MAH-The effects of reaction temperature and reaction time on the grafting ratio of three kinds of grafted starches were discussed. The results are shown in g-starch had the largest contact angle and the longest absorption time and the MA-g-starch had the smallest contact angle and the shortest absorption time, indicating that the relative hydrophobicity of LA-g-starch was the best and that of MA-g-starch was the worst. The relative hydrophobicity of the grafted starches was directly related to the number of hydrophobic groups grafted into the starch, and more hydrophobic groups resulted in better relative hydrophobicity. Thus, the measured contact angles were consistent with the grafting ratios.The molecular chain of the native starch contains many hydrophilic hydroxyl groups, and the grafting of hydrophobic groups onto the native starch would improve the relative hydrophobicity of the starch. The contact angle (CA) of water on a surface is the angle formed by a tangent line from the water droplet to the solid surface, and is an indication of the relative hydrophobicity of the sample surface. Generally speaking, the larger the CA, the higher the water resistance of the material . The surg-starch was the lowest and MA-g-starch was the highest. Thus, the LA-g-starch had the best relative hydrophobicity, and the MA-g-starch had the worst. The results were in agreement with the contact angle test results.To further confirm that graft modification can improve the relative hydrophobicity of native starch, the water absorption of the materials was determined based on their relative weight change after exposure to water. The results are shown in g-starch, LA-g-starch and MA-g-starch were characterized by gel permeation chromatography (GPC) and the results are shown in As shown in Mn) of native starch was 7.869 × 104 D, the weight average molecular weight (Mw) was 4.325 × 105 D, and the distribution index (DI) was 5.496. Compared with native starch, the Mn and Mw of grafted starches increased and the DI increased also. It was also proved that the grafting monomers were successfully grafted onto the starch molecular chain. However, the Mn, Mw and DI of grafted starches obtained by different grafting monomers was different. The Mn and Mw of from large to small was LA-g-starch, MAH-g-starch, MA-g-starch. The molecular weight distribution index also showed the same trend. This phenomenon was closely related to the grafting ratio of grafted starches. In g-starch was the largest, the polylactic acid molecular chain grafted on the starch molecular chain was the most. But the grafting ratio of MA-g-starch was only 0.57%, which leads to the minimum molecular weight and distribution index.It can be seen from g-starch, LA-g-starch and MA-g-starch were 23,600 mPa·s, 27,100 mPa·s and 17,200 mPa·s, respectively. It was proved that the molecular weight of starches increased after graft copolymerization. The pasting viscosity of LA-g-starch was the largest and MA-g-starch was the smallest, which indicated that the average molecular weight of LA-g-starch was the largest and that of MA-g-starch was the smallest. This also validates the results of the GPC test.For polymers, viscosity was proportional to molecular weight. In order to further verify the effect of the in situ solid phase polymerization on the molecular weight of modified starches, the pasting viscosity of the native starch and grafted starches were tested. The pasting viscosity of the native starch was 16,500 mPa·s. Compared with the native starch, the pasting viscosity of all grafted starches were increased. The pasting viscosity of MAH-Scanning electron microscopy, in principle, was to use a very fine focused high-energy electron beam to scan the sample and stimulate a variety of physical information, by accepting, amplifying and displaying this information, the surface morphology of the test specimen was observed. The various samples of test materials were subjected to SEM analysis to determine the extent of any changes in surface morphology of the material as a result of the grafting reaction.g-starch and LA-g-starch were more obvious, especially for the LA-g-starch. The results indicate that the surface roughness of the grafted starch was positively correlated with the grafting ratio.The FTIR, XPS, and SEM analysis of grafted starches all verified that a grafting reaction occurred as a result of the in situ solid phase polymerization method. Therefore, it was reasonable to assume that the crystalline structure of the reactants had also changed. We next analyzed the crystalline structure of the native starch and the grafted starches to further verify this conclusion. The XRD method was used to determine if there was any change in the crystal structure of the native starch as a result of the grafting reaction, and the results are shown in g-starch decreased to 23.47%, LA-g-starch decreased to 22.29%, and MA-g-starch decreased to 24.69%. The grafting ratio of LA-g-starch was the highest and more hydroxyl groups on the chain of starch reacted, with the most serious destruction of hydrogen bonds. The grafting ratio of MA-g-starch was the lowest and the destruction of its crystalline structure was the smallest. More graft polymer molecular chains attached to the starch molecular chain resulted in a more loose structure.The XRD diffraction peaks for native starch were typical of an A-type crystalline structure, with 2θ values of 15°, 17°, 18°, and 23° . After tg-starch was the lowest and the decomposition residual rate was the highest, and the MA-g-starch showed the opposite trends. The LA-g-starch was most easily decomposed, indicating the best thermoplasticity in the melting process. The most serious damage was evident for the crystalline structure of the LA-g-starch, and the MA-g-starch crystallinity was the lowest.The thermal behavior of the native starch and grafted starch were analyzed by TGA, and the results are shown in g-starch was the lowest and that of MA-g-starch was the highest.The DTG results in g-starch, LA-g-starch and MA-g-starch were tested by DSC, and the results are presented in To (onset), Tp (peak) and Tc (conclusion), and the enthalpy of melting was referred to as ΔH.DSC is a common method for researching melting properties . The staTo and Tp between 103.54 and 134.88 °C, Tp was 129.03 °C, and ΔH was 630.27 J/g. After in situ solid phase grafting modification of MAH, LA and MA, the To, Tp, Tc of grafted starches all shifted to lower temperature, and ΔH was decreased to varying degrees. It was indicated that the grafted starches was easier to gelatinization and lower in enthalpy of gelatinization. This was mainly attributable to the decrease of crystallinity and intermolecular loosening after graft modification. There were differences in gelatinization temperature and gelatinization enthalpy of three grafted starches. The To, Tp, Tc, and ΔH of LA-g-starch were the lowest, and those of MA-g-starch were the highest. The reason for this difference was that the degree of crystallinity of the three kinds of grafted starches varies with different grafting ratios. At the same time, this indicates that LA-g-starch had the best thermoplasticity and MA-g- starch the worst.As can see in g-starch, LA-g-starch, and MA-g-starch were 6.50%, 12.45%, and 0.57%, respectively. The grafted reaction resulted from replacement of the hydroxyl groups on d-anhydroglucose moieties of the starch with hydrophobic carbonyl groups form the graft monomer, which resulted in improved relative hydrophobicity of the starch. With the influence of the grafting ratio, LA-g-starch had the best relative hydrophobicity and MA-g-starch had the worst. The Mn, Mw, and DI of LA-g-starch was the largest and MA-g-starch was the smallest. The surface of the grafted starches was covered with graft polymer, and the surface roughness and the bond degree of the MAH-g-starch and LA-g-starch were more obvious. The starch crystalline structure was destroyed by the grafting reaction, the degree of crystallinity was decreased, and the hydrogen bonding between the starch molecules was weakened, resulting in improved thermoplasticity in the grafted starches. For the degree of crystallinity of grafted starches, MAH-g-starch decreased to 23.47%, LA-g-starch was 22.29%, and MA-g-starch was 24.69%. The degree of thermoplasticity improvement from large to small was LA-g-starch, MAH-g-starch, and MA-g-starch.This study demonstrated the successful preparation of three grafted starches using an in situ solid phase polymerization reaction. The grafting ratios of the MAH-The grafted starches produced by in situ solid phase polymerization exhibited increased overall relative hydrophobicity with increased interface compatibility, which allows for expanded application of thermoplastic starch and starch/plastic composites. This work comparing the influence of three grafting monomers on the grafting ratio and hydrophobic modification of starch provides reference data for the preparation of blended composites of starch and different polymers.


The optimal positioning of anterior cruciate ligament graft is still controversially discussed. We therefore wanted to determine the tunnel-to-joint (TJA), tunnel-to-shaft (TSA), and graft-tunnel divergence angles which would provide the best outcome, determined by the KOOS (Knee Injury and Osteoarthritis Outcome Score). This study evaluated the clinical influence of graft orientation as measured with the KOOS questionnaire in patients with anterior cruciate ligament reconstruction with bone-patellar tendon-bone autografts.We designed a prospective cohort study, with a 1 ¼ year recruitment phase from March 2011 to July 2012 and a minimal follow-up period of 1 year. Inclusion criteria were patients ≥ 18 years of age receiving an ACL reconstruction with bone-patellar tendon-bone autografts at our institution after having suffered an acute ACL rupture. The primary outcome was the KOOS. Independent variables were patient age, gender, laterality of rupture, mechanism of trauma, and type of femoral and tibial fixation, as well as sagittal graft-tunnel divergence, TJA, and TSA, the latter two being assessed on coronal slices of magnetic resonance imaging. Equations modeling the relationship between TJA, TSA, and graft-tunnel divergence with the KOOS overall score were fitted, and the optimum angles were mathematically determined.In total, 31 patients were included in our study. Our cohort with a median age of 28 years was predominantly male. The mathematically determined optimal placement of the implant in the coronal plane was a TJA of 74.8°, a TSA of 80.1°, and a graft-tunnel divergence angle of 8.5°.With regard to patient-reported outcome, the optimal graft orientation is provided by a coronal tunnel-to-shaft angle of 80° and tunnel-to-joint angle of 75°, respectively. Interestingly, in our series, patients reported best clinical outcomes with a sagittal graft-tunnel divergence. These results should be validated in larger studies.The online version of this article (10.1186/s13018-018-0954-3) contains supplementary material, which is available to authorized users. Anterior cruciate ligament (ACL) rupture is a frequent sports injury that often leads to post-traumatic knee instability and secondary knee damage with meniscal tears and articular cartilage injuries . The priThe self-administered easy to use Knee Injury and Osteoarthritis Outcome Score (KOOS) developed in 1998 assesses all of the aforementioned outcomes and can be used for assessment of ACL reconstruction outcome . The KOOThe objective of this study therefore was to determine the optimal surgical implantation technique with regard to surgical outcome 1 year after surgery, determined by the implant or tunnel angles associated with the minimum KOOS overall score.n = 5), additional rheumatic/musculoskeletal disorders (n = 3), and inadequate image quality in any sequence  of magnetic resonance imaging (MRI) (n = 6). MR imaging was performed using a 1.5 Tesla MR unit . Patients who did not complete the 1-year follow-up period (n = 12) or withdrew their consent to participate in the study (n = 3) were also excluded from the study, as were patients who did not complete the KOOS (n = 2). One patient, who has sustained a re-rupture within the follow-up period, was excluded due to missing assessability of the graft orientation. In total, 63 patients receiving ACL reconstruction at our center during the aforementioned timeframe were screened, of which 31 were included in the final analysis. Both clinicians and patients were blinded as to the results of the MRI assessments to avoid bias.We designed a prospective cohort study, with a 1 ¼ year recruitment phase from March 2011 to July 2012 and a minimal follow-up period of 1 year. Inclusion criteria were patients ≥ 18 years of age receiving an ACL reconstruction with bone-patellar tendon-bone autografts at our institution after having suffered an acute ACL rupture. The primary outcome was the KOOS after a minimum follow-up period of 1 year, after which patients were requested to complete the KOOS questionnaire. MR imaging was obtained on the day of filling in the KOOS questionnaire. Exclusion criteria were prior ACL reconstruction . The patient-administered KOOS questionnaire consists of five subscales . Patient age, gender, laterality of rupture, mechanism of trauma, type of femoral and tibial fixation, transplant orientation to joint (TOJ), transplant orientation to shaft (TOS), tunnel-to-joint (TJA), and tunnel-to-shaft (TSA) angles as well as sagittal graft-tunnel divergence  were included in our analysis. The angles defined are illustrated in Figs. y = k + ß1 × x + ß2 × x2 to model the relationship between the KOOS and TSA or TJA respectively. We fitted linear and quadratic functions to model the relationship between the KOOS and graft-tunnel divergence angle. The optimal TJA, TSA, and graft-tunnel divergence angles in the coronal plane were determined mathematically by resolving the curve’s equation for the minimum KOOS, i.e., where the derivative of the independent variable was 0.Data was collected in spreadsheets . All statistical analyses were performed using IBM SPSS Statistics version 22 . Distribution of independent variables was assessed with descriptive statistics. The relationship between TSA, TJA, graft-tunnel divergence angle, and the primary outcome, the KOOS, was graphically assessed. Presuming there is an optimum for which the KOOS would be the lowest , we fitted quadratic functions of the type In total, 31 patients were included in our analysis. Our cohort comprised younger, mostly male patients with a normal body mass index. The median age at injury was 28 years. All but two ACL ruptures were sports-related, the most frequent being soccer (36%) followed by alpine skiing (26%). Concomitant meniscal tears were frequently diagnosed (48%), whereas cartilage lesions (maximum grade II according to Outerbridge) were only found in two patients (6%). However, the KOOS overall score was not significantly different between patients with and without concomitant injuries. Femoral fixation was performed with screws of 6–7 mm in width and 19 or 23 mm in length and tibial fixation with screws 7–9 mm in width and 23 mm in length. An ACL implant with a thickness of 9 mm was inserted in 24 patients (77%), and an ACL implant of 8 mm thickness was used in 4 patients (13%). Further ACL implant thicknesses employed were 8.5 mm (2 patients) and 7.0 mm (1 patient). Patient characteristics are summarized in Table R squared was higher for the quadratic model , we chose the quadratic equation to model the relationship between the KOOS and graft-tunnel divergence. We also modeled the relationship between the KOOS (dependent variable) and TSA as well as TJA  with quadratic equations. The quadratic equation of coronal TOJ and TOS angles showed an insufficient curve fit  and no significant p values. Thus, the derivation of optimum values using the parameter estimates was not meaningful for TOJ and TOS angles. The summaries of the curve estimations are depicted in Table We modeled the relationship between the KOOS overall score (dependent variable) and graft-tunnel divergence (independent variable) with quadratic and linear equations. As the y’ = f’(x) = 0. In the coronal plane, the optimal angles hence were a TJA of 74.8° and a TSA of 80.1°. In the sagittal plane, the curve morphology did not admit the abovementioned calculations of optimum values due to its convexity. The optimal graft-tunnel divergence angle was calculated to be 8.5°.With the optimum outcome being the minimum KOOS, the optimal implant angles can be mathematically determined by resolving the equation for Assessing the relationship between post-surgical outcome determined by the KOOS and surgical ACL implantation technique determined by the TSA, TJA, and graft-tunnel divergence angles in a prospective cohort study with 31 patients, we mathematically determined the optimum angles to be 80.1°, 74.8°, and 8.5°, respectively.Although calculated in a small study population, the sagittal graft-tunnel divergence angle was conspicuous. Derived from the surgical technique of implantation with drilling a bone tunnel over a leading wire, we expected the optimal graft-tunnel divergence angle to lean towards zero. Especially when considering the supine position of the patient during the image acquisition, a posterior graft-tunnel divergence seems surprising. Graft-tunnel divergence at the tibial site could possibly be associated to tibial tunnel widening through a wiper effect. Eventually, this observation and its biomechanical or clinical relevance need to be verified and further investigated in larger studies.This study certainly has limitations. The small sample size substantially limited the statistical possibilities and the validity of the results. However, as this is a primarily radiological investigation, we did not want to compromise the imaging quality and excluded patients with poor post-surgical imaging. Therefore, we accepted a high dropout rate. The recruitment process is illustrated below. Despite the small sample size and weak statistics, our results did not show significant differences when compared to other series .n = 1). This might have yielded valuable information on the desirable graft orientation.The study design also contains the weakness of acquiring the imaging at different points in the course of treatment. The postoperative MRI was obtained between 11 and 23 months (median 20 months) postoperatively. Even though it is doubtful, that the graft orientation changes during this time, imaging earlier in the postoperative course might have allowed inclusion of patients that have sustained a graft re-rupture . (DOC 111 kb)


T0, Tp, and Tc), gelatinization temperature ranges (ΔTr), and enthalpies of gelatinization (ΔH) were measured on a differential scanning calorimeter analyzer (DSC) and ranged from 57.81 to 61.25°C, 64.36 to 67.57°C, 82.03 to 84.70°C, and 21.52 to 26.89°C and 7.14 to 10.66 J/g, respectively. The varying chemical and structural characteristics of HB starch isolated from different cultivars suggested the potential for broader applications of the cereal.This study aimed to isolate starch and evaluate its chemical and structural characteristics from six Chinese hulless barley (HB) cultivars. Starch isolated from naked barley displays A‐type crystalline packing and a regular granular shape. We measured peak viscosity values ranging from 237 to 264 cP, trough viscosity values from 305 to 380 cP, breakdown values from 390 to 535 cP, final viscosities from 357 to 523 cP, setback values from 245 to 354 cP and 383 to 460 cP, peak times from 5.53 to 5.73 min, and pasting temperatures from 93.10 to 94.65°C by RVA. Transition temperatures ( The currently selected variety is a newly developed variety that has a large cultivation volume, high yield, and strong adaptability in Tibet in the past 10 years, and basically covers the main varieties of the current barley in Tibet. Among them, black barley is a newly developed variety containing anthocyanins in Tibet, and the others are white barley. It is worth noting that the starch of the new main cultivar has not been studied and compared, and it has research value, and it is easy to collect in Tibet, which is conducive to experimentation. Moreover, the differences in the nature of these six species are also obvious. The results of this study have great guiding significance for the deep processing of barley. After studying and comparing their properties, they can have a clear understanding of the advantages and disadvantages of each species and use their unique advantages to apply to their suitable fields.22.1Seeds of HB  were collected in 2018 from the major production areas of China by the Agricultural Research Institute of Tibet Academy of Agricultural and Animal Husbandry Sciences. All samples were obtained from a single field and harvested prior to analysis.g for 12 min, and the supernatants were discarded. The upper precipitate (pigment layer) was removed, and the starch was separated by resuspending in water. Samples were centrifuged, the starch paste was washed in ethanol (95%), and samples were centrifuged three times. Supernatants were discarded, and the starch was oven‐dried at 50°C for 12 hr.Hulless barley starch was obtained according to methods described by Bello‐Pérez group with minor modifications .Morphology was assessed by scanning electron microscope . The crystallinity was evaluated on MDI Jade 6 software to fit the measured XRD peak. R% crystallinity is used to measure the ratio of peak area, background area, and peak area . A two‐way ANOVA test was used for statistical analysis using ANOVA and Tukey's test. Analysis was performed by SPSS 16.0 statistical software. p < 0.05 indicates that the difference is statistically significant.The data obtained were the average values of the six HB varieties , while Zangqing 2000 had the highest (95.50°C). The majority of HB starch samples had a pasting temperature of ~93.87°C.3.5T0), conclusion temperature (Tc), peak temperature (Tp), gelatinization temperature range (ΔTr), and enthalpies of gelatinization (ΔH) significantly differed among samples. The transition temperature , ΔTr, and ΔH ranged from 57.81 to 61.25°C, 64.36 to 67.57°C, 82.03 to 84.70°C, and 21.52 to 26.89°C and 7.14 to 10.66 J/g, respectively. Dongqing 17 showed the lowest onset and conclusion temperatures. The maximum onset temperature was Dongqing 18. Higher gelatinization temperatures indicate a greater energy requirement for starch gelatinization. Gelatinization temperatures among the starch samples differed according to crystallinity, granule shape and size, % amylose, amylopectin length, and the arrangement of starch fractions within granules. Dongqing 17 displayed the highest ΔTr value (26.89°C), with Dongqing 18 starch showing the lowest (21.52°C)  Table . The ΔT4We demonstrate the diversity of HB starch from different cultivars regarding physicochemical and morphological, pasting, thermal, and structural properties. This suggests that starch suits an array of applications. The starch from HB cultivars has comparable crystalline packing and granule shape, but the degree of crystallinity significantly differs. Experimental results have shown that among the six species, the extraction rate of Dongqing 18 and Zangqing 2000 starch is high, which is most suitable for food applications.Hulless barley starch has the following potential applications: firstly, convenient food is processed. General foods are expected to obtain highly gelled and unaged product slag. Highly gelatinized foods are soft, palatable, easy to rehydrate, and easily digested by amylase. Secondly, the modified starch is processed. The modified starch has the characteristics of good rehydration, high viscosity, and stable viscosity. It can be used as a binder, thickener, and sizing agent in food, aquatic feed, paper, textile, and other industries. Characteristically, the development of specialty drinks and food: The Qinghai–Tibet Plateau has a high climate and a cold climate, making people demand more wine. Green barley contains 10.1% protein, 1.8% fat, and 70% carbohydrates and is rich in minerals, amino acids, and vitamins the body needs. This is a good brewing material. It is rich in nutrients and has higher nutritional value than rice, corn, and common wheat. It has a large calorific value and is full of hunger and cold. It is easy to carry and suitable for the lives of herders. In addition, it is recommended to develop green barley food, including barley biscuits, barley cakes, barley noodles, and other products, in view of high barley protein content, low gluten content, high amylopectin content, high material viscosity, reasonable processing technology and formula design, optimized operating conditions, high‐quality breakfast foods, snack foods, and nutritional health care for people with high blood lipids.However, further studies are required to assess the desirable characteristics and commercial application of each starch through the analysis of their structural, physical, and functional properties. Studies investigating the impact of modifications on the structural and physiochemical properties of HB starch sampled from the different cultivars are also warranted.The authors declare that they do not have any conflict of interest.This study does not involve any human or animal testing.Written informed consent was obtained from all study participants.


Rice mutants with altered starch components and properties are important genetic resources in rice breeding programmes. In this study, 44 mutants with altered starch components were screened from 135 rice mutants with opaque kernels using a starch–iodine absorption spectrum method, and nine mutants from them were further selected for investigating their starch properties and kernel appearance quality. The results showed that the iodine absorption spectrum parameters, OD620, OD620/550, and λmax, could reflect the changes of starch components in rice mutants, and had significantly positive relationships with amylose content and negative relationships with the proportion of short branch-chains of amylopectin. The endosperm starches from nine mutants all showed A-type crystalline structure and similar short-range ordered structure, but had different relative crystallinities. The changes of starch components in mutants not only resulted in the different gelatinization properties of starch but also changed the appearance quality of brown rice kernels. This study provided abundant genetic plants for studying the molecular mechanism of starch synthesis and the quality regulation of rice kernels. Rice is one of the most important crops, and provides the staple food for over half the world’s population. Starch is the main component of rice endosperm, and its properties greatly influence the eating and cooking qualities of rice and its processing method in industrial use ,2,3. TheIn recent years, some novel rice materials with different qualities have been cultivated through traditional cross breeding, chemical or physical mutagenesis breeding, and advanced biotechnological approaches, including transfer DNA or transposon insertion and RNA interference ,16,17,18In this study, we screened mutants with altered starch components from rice mutants with opaque kernels. Their starch properties and kernel appearance qualities were investigated. This study should provide valuable genetic materials to study the molecular mechanism of starch synthesis, and provide important information for rice quality breeding.Starch contains two main components: amylose and amylopectin. Their contents can be simply detected through measuring their iodine absorption spectra . The iodExcept the starch-iodine colorimetry, the methods for measuring starch components are concanavalin A precipitation, gel permeation chromatograph (GPC), size-exclusion chromatography, near-infrared analysis, and differential scanning calorimetry (DSC) analysis ,19,20. Tgranule-bound starch synthase I gene [The AC in isolated starch was measured using Megazyme amylose/amylopectin assay kit . Accordie I gene ,22.In order to further confirm the component changes of starch, the isoamylase-debranched starch was analysed using GPC . The thrp < 0.01). Similar results have been reported in the previous paper [In order to reveal whether the changes of iodine absorption spectrum parameters can reflect the changes of starch components, their relationships were analysed . The OD6us paper . The prePrincipal component analysis (PCA) is a widely used method to determine the relationships of variables that contribute to differences between physicochemical properties or samples. The iodine absorption spectrum parameters and starch components of ten rice materials were subjected to PCA, and the results are present in granule-bound starch synthase I or soluble starch synthase does not change to the crystalline structure of starch, though starch components change significantly [The X-ray diffraction (XRD) patterns of starches are shown in ficantly . The relficantly ,27,30. H−1 have been linked with order/crystallinity and amorphous regions in starch, respectively. The ratio of absorbance 1045 to 1022 cm−1 (1045/1022 cm−1) is usually used to assess the ordered degree of starch [The branch-chains of amylopectin form two types of helices in starch granules. The helices that are packed in short-range order are defined as the double helical order, and the helices that are packed in long-range order are related to the packing of double helices forming crystallinity . The shof starch . The FTIf starch .Wx alleles and ACs, the starch with lower AC has a higher gelatinization temperature. This is due to the fact that the lower amylose contributes to the higher relative crystallinity of starch, needing a higher temperature to disrupt it [The thermal properties of starch are important functional properties and determine the applications of starch. The thermal properties of starches are usually determined by DSC, which measures the change of heat involved in starch gelatinization. The gelatinization results in the disruption of double helical and crystalline structure of amylopectin in starch granules. The DSC thermal curves of starches from rice mutants are shown in srupt it . In factsrupt it . The chasrupt it ,37. Amonsrupt it . In the The unhulled mature grains are shown in Compared with floury endosperm mutants with slightly changed starch components due to the mutation of storage protein synthesis or aleurone cell development ,13,14, tjaponica rice (Oryza sativa L.) variety (WT), Kitaake, through 60Co gamma irradiation [The rice mutants with opaque kernels were derived from a adiation . These mStarch was isolated from brown rice kernels following the method of Lin et al.  with som2/KI. The sample was scanned from 400 to 900 nm using a spectrophotometer .The iodine absorption spectrum of starch was measured exactly following the method of Lin et al. . BrieflyThe AC in isolated starch was measured using an amylose/amylopectin assay kit .The molecular weight distribution of starch was analysed using GPC following the method of Lin et al. . BrieflyThe crystalline structure of starch was measured using X-ray diffractometer  following the method of Lin et al. . The tes−1, resolution of 4 cm−1, and scan times of 64. The original spectrum was corrected by a baseline from 1200 to 800 cm−1, and deconvoluted using a Resolutions Pro with resolution enhancement factor of 1.9 and peak half-width of 1.9 cm−1.The short-range ordered structure of starch was measured using an attenuated total reflectance–Fourier transform infrared (ATR–FTIR) spectrometer  following the method of Lin et al. . The tesFive mg of starch and 15 μL deionized water were mixed and sealed in an aluminium pan. The sample was equilibrated at room temperature for 2 h, and then analysed using a differential scanning calorimeter . The test conditions included a heating rate of 10 °C/min and a temperature range from 20 to 130 °C.The hulled grains and dehulled brown rice kernels were photographed with a digital camera . The cross section of brown rice kernel at the middle region was photographed using a Stereo Microscope . The lengths, widths, and thicknesses of 30 brown rice kernels were measured using an electronic digital calliper . The 100 brown rice kernels were weighed and repeated three times.The data were analysed using the SPSS 16.0 Statistical Software Program, and the one-way analysis of variance was evaluated using Tukey’s test. The principle component analysis was carried out using Minitab Version 16.0 Software .The starch-iodine absorption spectrum parameters: OD620, OD620/550, and λmax, could reflect the component changes of starch. Forty-four rice mutants with ACs from 2.2% to 14.3% were rapidly screened from 135 rice mutants with opaque kernels according to the changes of starch-iodine absorption spectrum parameters. The screened mutants had significantly different starch molecular weight distributions but exhibited the same A-type crystallinity and similar short-range ordered structures. The mutants with different starch components had different thermal properties of starch, and exhibited significant differences in length, width, thickness, and weight of their brown rice kernels. The newly selected non-waxy floury mutants are valuable genetic materials for further investing the molecular mechanism of starch synthesis and the quality regulation of rice kernels.


SnRK1 gene has been found to increase starch accumulation in several plant species. However, its roles in improving starch quality have not been reported to date. In this study, we found that the IbSnRK1 gene was highly expressed in the storage roots of sweet potato and strongly induced by exogenous sucrose. Its expression followed the circandian rhythm. Its overexpression not only increased starch content, but also decreased proportion of amylose, enlarged granule size and improved degree of crystallinity and gelatinization in transgenic sweet potato, which revealed, for the first time, the important roles of SnRK1 in improving starch quality of plants. The genes involved in starch biosynthesis pathway were systematically up‐regulated, and the content of ADP‐glucose as an important precursor for starch biosynthesis and the activities of key enzymes were significantly increased in transgenic sweet potato. These findings indicate that IbSnRK1 improves starch content and quality through systematical up‐regulation of the genes and the increase in key enzyme activities involved in starch biosynthesis pathway in transgenic sweet potato. This gene has the potential to improve starch content and quality in sweet potato and other plants.Sucrose non‐fermenting‐1‐related protein kinase‐1 (SnRK1) is an essential energy‐sensing regulator and plays a key role in the global control of carbohydrate metabolism. The  Its potential as food and carbohydrate source has been widely recognized  analysis showed that the expression levels of Agrobacterium tumefaciens  and empty vector control (VC)  Figure i. The tr) Figure j–m.IbSnRK1 were significantly higher in the transgenic sweet potato plants than in WT and VC  of amylopectin exhibited a similar pattern in the four transgenic plants and WT/VC, and four peaks were present at approximately degree of polymerization (DP) 7, DP 12, DP 17 and DP 46 in L13, L14, WT and VC, but only three peaks  were found in L17 and L18  to 82.4 °C  with an enthalpy (ΔH) of 11.1 J/g. However, To, Tc and ΔH of starch from the transgenic plants were significantly decreased, indicating their starch was earlier to gelatinize, compared with WT  encoding acid invertase, neutral invertase, sucrose synthase, hexokinase, UDP‐glucose pyrophosphorylase, phosphoglucomutase, AGPase, GBSSI, SSI, SSII, SSIII and SBEII, respectively, were systematically up‐regulated in the storage roots of transgenic sweet potato plants compared with WT and VC  and glucose‐1‐phosphate (Glc‐1‐P) were significantly decreased, but those of uridine diphosphate glucose (UDP‐glc), adenosine triphosphate (ATP), adenosine diphosphate glucose (ADP‐glc) and ADP were significantly increased in transgenic sweet potato plants compared with WT Figure . No diffSnRK1 gene has been shown to increase starch accumulation in several plant species such as barley , work together to catalyse the biosynthesis of amylopectin . qRT‐PCR was conducted to determine the transcript levels of IbSnRK1 using SYBR Premix Ex Taq  with a 7500 Real‐Time PCR system . Specific primers of IbSnRK1 and the sweet potato Actin (AY905538) gene as internal control were listed in Table CT method  of Luhsu 3 using Trozol Kit . The first‐strand cDNA was synthesized using PrimeScriptIbSnRK1 to exogenous sucrose was examined according to the method of Wang et al. (m sucrose or water (control) in the dark at 28 °C, and sampled at 0, 3, 6, 12, 24 and 48 h after treatment to analyse the expression of IbSnRK1.The response of g et al. . Leaf peIbSnRK1 to the circadian rhythm was investigated as described by Leterrier et al.  and bar genes driven by a CaMV 35S promoter, respectively. Embryogenic suspension cultures of Lizixiang were prepared as described by Liu et al.  as described by Nishi et al.  to determine X‐ray diffraction. The crystal type and degree of crystallinity were calculated with Jade 5.0 software .To, peak temperature (Tp), Tc and ΔH were measured with Universal Analysis 2000 .Starch samples and distilled water were mixed , and the suspension was sealed in an aluminium pan with an empty aluminium pan as control for 24 h under room temperature. The samples were heated over a temperature range of 30–95 °C (raising the temperature at 10 °C/min) with a differential scanning calorimeter , and the Expression of starch biosynthesis‐related genes in the freshly harvested storage roots was analysed by qRT‐PCR as described above. The specific primers were listed in Table et al.  and P < 0.01 , respectively.All experiments were independently performed three times, and the data were presented as the mean ± SE. Results were analysed by Student's The authors declare no conflict of interest.Figure S1 Production of transgenic sweet potato plants overexpressing the IbSnRK1 gene.Table S1 X‐ray diffraction patterns of starches from the storage roots of the transgenic sweet potato plants, WT and VC.Table S2 The thermal characteristics of starches from the transgenic sweet potato plants, WT and VC.Table S3 Primers used in this study.Click here for additional data file.


To investigate predictive factors related to graft failure of IgA nephropathy(IgAN) in renal allografts following living donor transplantation.We identified a series of 102 biopsies diagnosed as IgAN in renal allografts following living donor transplantation from July 2004 to January 2017 at our center, and assess the predict value of the Lee’s classification and the 2009 Oxford classification in IgAN in renal allografts, clinical, ultrasonic and pathological characteristics at biopsy and the outcomes were retrospectively analyzed.The 5-year graft cumulative survival rate after transplantation was 91.4%. The 4-year graft cumulative survival rate after biopsy diagnosis of IgAN in renal allografts was 59.6%. The mean time ± SD to disease was 4.7 ± 3.5 years. The color doppler ultrasound and blood flow imagine showed the echo enhancement, the reduced blood flow distribution, the reduced peak systolic velocity of main renal artery, and the increased resistance index of arcuate renal artery were valuable in evaluating the graft dysfunction. The Cox multivariate analysis revealed that the 24-h urinary protein level , estimated glomerular filtration rate (eGFR) , and mesangial C1q deposition  at biopsy were independent predictive factors of graft failure of IgAN in renal allografts.IgAN in renal allografts occurred frequently within 5 years after transplantation. The risk of graft failure should be taken seriously in patients who exhibit heavy proteinuria and/or a declined eGFR as the initial symptoms; a high lesion grade (grade IV-V of Lee’s classification) and/or mesangial C1q deposition may also indicated a poor outcome. The widespread use of effective immunosuppression has markedly reduced the risk of graft failure due to rejection in recent year s. ConversIgA nephropathy (IgAN) is the most common recurrent nephropathy after transplantation, especially living donor transplantatio n, 6. SomeIn a total of 637 living donor transplantations in the First Affiliated Hospital of Sun Yat-sen University  from July 2004 to January 2017, we identified a series of 102 patients with a diagnosis of IgAN in renal allografts proven by biopsy, including 93 indication biopsies and 9 protocol biopsies, repeated biopsies with a same diagnosis were excluded. The series consisted of 72 males and 30 females with a mean age ± SD of 35.5 ± 9.1 years. Sixty-seven patients have been proven primary IgAN by biopsy for native renal in this series.A follow-up was performed in the First Affiliated Hospital of Sun Yat-sen University  and Guangzhou Women and Children’s Medical Center . Routine blood and urine examinations, liver and renal function tests, and therapeutic drug monitoring were conducted prior to biopsy for all patients. The standard maintenance therapy included a calcineurin inhibitor (CNI)  in combination with mycophenolate mofetil (MMF) (Mycophenolate Sodium was initiated at 360 mg q12h) and corticosteroids. Oral prednisone was initiated at 30 mg (qd) and gradually reduced to 5–10 mg (qd) over a 4-week period. Table The Regional Ethics Committee of our center approved this study, and all patients signed informed consent forms.The following indications for biopsy were combination of doctors’ experience and the 2009 KDIGO Clinical Practice Guideline for the Care of Kidney Transplant Recipient s were useNinety-three patients with indications were suggested accept biopsy in 1 month, most of them (80.6%) underwent a timely biopsy, the mean ± SD waiting time for biopsy was 0.7 ± 1.2 months.Nine patients with high risk factors for recurrence, such as a family history or primary IgAN diagnosed by native renal biopsy, underwent protocol biopsies at 6 months and at 1, 2, 5 and 10 years.An ultrasonography-guided needle biopsy was performed using an 18-gauge needle (Bard). Each sample included at least 6 glomeruli visible by light microscopy. Immunofluorescence analyses were performed for all biopsies, and the IgA, IgG, IgM, C3, C1q and C4D levels were graded by two senior pathologists in an independent and blinded fashion. A diagnosis of IgAN in renal allografts based on IgA-positivity due to immunofluorescence staining deposition in the mesangial area  and RI in each renal artery.According to 2002 K/DOQI clinical practice guidelines for chronic kidney disease, we definWe analyzed the time to disease recurrence, the 5-year graft cumulative survival rate after transplantation and the 4-year graft cumulative survival rate after biopsy diagnosis. Because the aim of this study was to investigate the predictive factors related to graft failure of IgAN in renal allografts rather than recurrence itself, we defined the initiation of the follow-up time from the day of biopsy diagnosis instead of the day of transplantation.p values lower than 0.05 were considered significant. Correlation analyses were performed among abovementioned characteristics, Pearson’s and Spearman’s correlation coefficient were used for continuous and categorical variables respectively.The urinary red blood cell (U-RBC) count, 24-h urinary protein level, serum albumin (ALB), eGFR, degree of histopathological injury, ultrasonic and other clinical characteristics at the time of the biopsy diagnosis were recorded. Based on Lee’s classifications, ultrasonic and blood flow index such as cortical thickness, echo enhancement, Vs and RI were compared in different degree of histopathological injury. Variables were compared using the Mann-Whitney U test or Chi-square test, and Univariate Cox proportional hazards models were used to investigate the significance of predictive factors related to graft failure, and significant factors were tested in the multivariate analysis. Kaplan-Meier estimates were used to create graft survival curves, and comparisons were performed using the log-rank test.The data were analyzed using the IBM SPSS Statistics software version 22.0.Ninety-three patients who were diagnosed with IgAN in renal allografts underwent a biopsy for initial symptoms. Proteinuria occurred in 72 patients (70.6%), hematuria occurred in 65 patients (63.7%), hypoproteinemia occurred in 18 patients (17.6%), and increased sCr (> 30% of the baseline) occurred in 28 patients (27.4%). Nine patients (8.8%) did not have abnormalities and were diagnosed using the protocol biopsy.Graft failure occurred in 26 patients. The 5-year graft cumulative survival rate was 91.4% after transplantation , and the 4-year graft cumulative survival rate after diagnosis of IgAN by biopsy was 59.6% . The mean time ± SD to occurrence was 4.7 ± 3.5 years. Sixty-four cases (62.7%) occurred within 5 years after transplantation, and 38 cases (37.2%) occurred after more than 5 years , the reduced blood flow distribution (p = 0.02), the reduced Vs of main renal artery (MRA) (p = 0.04), and the increased RI of arcuate renal artery (ARA) (p = 0.04) were valuable in diagnosing and evaluating the graft dysfunction (Table p <  0.01), the reduced blood flow distribution , the reduced Vs of MRA , and the increased RI of ARA .The color doppler ultrasound and blood flow imagine showed the echo enhancement , time to disease occurrence (p = 0.05), the 24-h urinary protein level (p <  0.01), eGFR (p <  0.01), hypoproteinemia (ALB < 35.0 g/L) (p <  0.01), Lee’s classification (p = 0.02), mesangial C1q deposition (p <  0.01) at the time of biopsy were predictive of graft failure. We found significant correlations between the abovementioned characteristics. For instance, the time to disease occurrence was directly correlated with age , Lee’s classification , and mesangial C1q deposition . The 24-h urinary protein level was directly correlated with hypoproteinemia . The multivariate analysis demonstrated that the 24-h urinary protein level, eGFR and mesangial C1q deposition at the time of the biopsy were independent predictive factors related to graft failure. The hazard ratios (HRs) were 1.6 for 1-g increased 24-h urinary protein (95%CI 1.2–2.0), 1.0 for 1-mL/min/1.73 m^2 declined eGFR (95%CI 1.0–1.1), 3.0 for mesangial C1q deposition (95%CI 1.2–7.4)  (Table p <  0.01) and other factors.Although some characteristics were not independent predictive factors of graft failure in our study, the log-rank test revealed a significant difference between the Kaplan-Meier curves of Lee’s classification (1) Table . Figure Due to differences in diagnostic criteria, discrepancies have been reported in recurrence rates, with a range from 9 to 61 %. Family More and more recent studies confirmed that IgAN in renal allografts had impact on long-term graft survival,, 10, 16 We found that IgAN occurred frequently within 5 years postoperatively and then declined sharply in subsequent years, which was consistent with a previous repor t. Few patAn increased sCr level or declined eGFR as the initial symptoms was generally associated with a high risk of graft failure. This result was confirmed in a previous study on native IgA N. AnotherThe color doppler ultrasound is helpful to evaluate the progress of renal graft diseases, the change in cortical thickness, echo enhancement and the reduced blood flow distribution are typical features of graft dysfunction, like echo enhancement denotes glomerulosclerosis and interstitial fibrosis, which showed in our results. The results indicate that patients with severe histopathological injury (grade IV-V of Lee’s classification) may show lower Vs and higher RI in blood flow than mild or moderate (grade I-III), it coincide partly with early research, which isLee’s classification and the Oxford classification are useful prognostic indicators of disease progression and clinical outcomes of native IgAN, but their utility in IgAN in after transplantation remains unknown. Lee’s classification includes criteria for histopathological grading, such as glomerular sclerosis, crescent formation, interstitial inflammation and tubular atrophy, which were established based on and considered suitable for East Asian patients. Patients with mild or moderate (grade I-III) lesions have been reported to have a benign outcome, whereas patients with grade IV or V lesions develop end-stage renal failure, which isSixteen cases with mesangial C1q deposition in this series. Traditionally, the presence of C1q staining is viewed as a typical pathological lesion of LN rather than IgA N. HoweverMaintenance therapy with medium/low prednisone is used to relieve IgAN progression in combination with angiotensin-converting enzyme inhibitors (ACEIs) to reduce proteinuria. Early steroid withdrawal is a safe intervention in living donor transplantation,, 35 but Our study analyzed the 5-year graft cumulative survival rates after transplantation, but longer-term follow-up from biopsy is needed to observe more outcome events. In most cases, the primary diseases of the recipients were unknown, only 67 cases have been proven primary IgAN by biopsy for native renal in this series, the differences in clinical and pathological characteristics between de novo and recurrent IgAN couldn’t be found in this study. Although major confounding variables were adjusted and our results were consistent with other studies in the literature, bias was unavoidable in the univariate Cox proportional hazards models due to the influence of the lack of information for the intraoperative and donor factors.IgAN is the most common de novo or recurrent nephropathy, especially in living donor transplantation, and occurs frequently within 5 years after transplantation. The color doppler ultrasound and blood flow imagine were valuable in diagnosing and evaluating the graft dysfunction. The risk of graft failure should be taken seriously in patients who exhibit heavy proteinuria (24-h urinary protein level > 2 g) and/or a declined eGFR as the initial symptoms, especially with hypoproteinemia, a high lesion grade (grade IV-V of Lee’s classification) and/or mesangial C1q deposition.


The concentration of copper catalyst (CuCl2) loading was as low as 1/64 equivalent to chlorine atom in the presence of Me6-Tren under UV irradiation. The light-responsive nature of graft polymerization was confirmed by “off-on” impulsive irradiation experiments. Temporal control of the polymerization process and varied graft contents were achieved via this photoinduced Cu(II)-mediated RDRP.Photoinduced Cu(II)-mediated reversible deactivation radical polymerization (RDRP) was employed to synthesize poly(vinylidene fluoride- Atom transfer radical polymerization (ATRP) ,2,3,4 ha2 and excess tris[2-(dimethylamino)ethyl]amine (Me6-Tren) exhibited outstanding efficiency in acrylate polymerizations :[Cu]:[Me6-Tren]:[AN] = 1:(1/32):(6/32):30) was put into the ultraviolet light (λmax ~ 365 nm) while cooling with a hair dryer. Samples were taken at regular time intervals followed by precipitation in H2O/CH3OH (v:v = 1:1) mixture, washed three times with ethyl alcohol, and dried overnight under reduced pressure. The resultant graft copolymer was obtained for characterization.Polymerizations were conducted by using a Schlenk technique . First, d6 containing tetramethylsilane (TMS) as internal standard. Fourier transform infrared (FTIR) spectroscopy of polymer films was performed on a Nicolet iS5 . Differential scanning calorimetric (DSC) analysis was conducted on a Discovery DSC 250 . After rapid heating and cooling cycles (at a rate of 20 °C/min) to remove the thermal history, the sample was heated at a rate of 10 °C/min under nitrogen atmosphere. Thermogravimetric analysis (TGA)  results were recorded on Netzsch STA 449 F3 Jupiter  in nitrogen atmosphere at a heating rate of 10 °C/min.Nuclear magnetic resonance (NMR) spectra were recorded on a Bruker  (Advance III) 400 MHz instrument for solutions in DMSO-co-CTFE)-g-PAN copolymers could be synthesized via Cu(0)-mediated reversible deactivation radical polymerization (RDRP) in batch and flow reactors [2/Me6-Tren was employed to promote the polymerizations of AN with P(VDF-co-CTFE) as macroinitiator. The polymerization results are listed in co-CTFE)-g-PAN (graft content = 20.8 mol %) was obtained with a low copper concentration ([Cl]:[Cu]:[L]:[AN] = 1:(1/32):(6/32):30) (Poly(acrylonitrile) (PAN) and its copolymers have been widely used as precursors for novel carbon materials with outstanding properties and performances . P:30) , run 5 a/32):30) , run 7. co-CTFE) with no PAN graft chains was obtained by the removal of UV irradiation (co-CTFE) afforded no product -g-PAN with varied graft contents (co-CTFE)-g-PAN (graft content = 20.8 mol %) was used as macroinitiator to initiate polymerization of AN ([Cl]:[Cu]:[L]:[AN] = 1:(1/32):(6/32):(30), UV (λmax = 365 nm)). The graft content was increased into 30.2 mol % upon another 6 h exposure.The influence of [Cu]:[L] on graft polymerization was investigated. It was supposed that 1:6 would be better to yield a higher graft content . Under tco-CTFE)-g-PAN was characterized by FTIR, 1H NMR, and 19F NMR. In −1 (–CN) indicated the presence of PAN segments in the copolymer. The 1H NMR spectrum (2CH2CFClCF2–). A new peak appearing around 1.9–2.1ppm was attributed to the proton on the methylene group of the AN unit (–CH2-CHCN–). Another new peak appeared at 3.0–3.3 ppm and overlapped with the proton signals of head-to-tail connections of VDF in P(VDF-co-CTFE), which was assigned to the methine proton (–CH2CHCN–). The signals of –CH2CHFCF2– from hydrogenation of CTFE unites and –CH=CFlCF2– from elimination of HCl from main chains were not observed. This indicated that the typical side reactions did not happen in this photoinduced Cu(II)-mediated RDRP process. The 19F NMR spectrum provided more information about the structure of graft copolymers. In The chemical structure of P(VDF-spectrum  displayeco-CTFE)-g-PAN was explored by using DSC and TGA. In co-CTFE) and the crystal domain size were influenced by the introduction of PAN graft segments. The TGA (co-CTFE) began to decompose at about 400 °C, and about 40 wt % remained at 500 °C. Meanwhile, two polymer degradation stages were observed for the P(VDF-co-CTFE)-g-PAN copolymers. The first stage started from about 300 °C, which corresponded to the decomposition of PAN segment. The second stage began from about 450 °C, which was attributed to the degradation of P(VDF-co-CTFE) backbone chain. It was noteworthy that the weight remaining over 500 °C was enhanced with the increase of PAN content. It was supposed that the PAN was carbonized to carbon material with higher thermal stability.The thermal properties of P(VDF- The TGA  showed tco-chlorotrifluoroethylene)-graft-polyacrylonitrile (P(VDF-co-CTFE)-g-PAN) with varied graft contents were prepared via photoinduced Cu(II)-mediated reversible deactivation radical polymerization (RDRP). Upon UV irradiation, CuCl2/Me6-Tren enabled low catalyst loading and temporal control of the polymerization process. This protocol might have potential application in the area of novel dielectric materials. In summary, poly(vinylidene fluoride-


World Health Organization (WHO) defines tropical diseases as all diseases that occur solely, or principally, in the tropics’ [Historically, tropical medicine emerged from a multidisciplinary background as a result of progress in the areas of public health and hygiene, travel and exploration, biology and evolution, and the germ theory of disease . The Wortropics’ . WHO’s fTropical Medicine and Infectious Disease. It has been endorsed as the Official Journal of The Australasian College of Tropical Medicine (ACTM) and its joint Faculties of Travel Medicine and Expedition and Wilderness Medicine. Both MDPI and the ACTM are celebrating anniversaries this year, being 20 and 25 years old respectively, and are both proud of their respective traditions in the world of publishing. The Australasian College of Tropical Medicine has partnered with long term electronic publisher, MDPI, to develop an international, scientific, open access journal of tropical medicine and infectious disease to be published quarterly online. We have named this new journal, MDPI publishes 150 peer reviewed, scientific, open access, electronic journals in a broad range of fields. It has offices in four cities, being headquartered in Basel, Switzerland and having additional offices in China and Spain. MDPI is a member of all the major professional publisher associations, including OASPA, STM, or COPE and has a strong track record in building the indexing and abstracting support for each of its journals . Part ofAnnals of the ACTM since 1995, initially as conference proceedings and then as a peer-reviewed journal from 2002. All of these are available online [Dictionary of Tropical Medicine [The ACTM is particularly excited about this project being launched in its 25th Anniversary Year. The ACTM itself has been publishing the e online . The ColTropical Medicine and Infectious Disease is to publish authoritative and original articles, critical and systematic reviews, editorials, perspectives, short communications, commentaries, book reviews, letters to the editor and special issues on all aspects of tropical medicine and tropical infectious disease. All articles will be submitted through MDPI’s excellent online journal submission platform and are subject to editorial and peer-review. The MPDI Editorial Process is extremely rigorous [The aim of rigorous , but it Tropical Medicine and Infectious Disease will publish on all tropical diseases of global significance, as well as neglected tropical diseases, as defined from time-to-time by the WHO. The scope of the journal includes, but is not limited to clinical tropical medicine, tropical infectious diseases, parasitology and entomology, bacteriology, mycology and virology, epidemiological and social science studies, chemotherapy and pharmacology, immunology, disease prevention, control and elimination, emerging and re-emerging infectious diseases, emerging public health threats, and global health and One Health.We are very proud to welcome our foundation international Editorial Board, an extremely talented group of more than 20 senior academics and clinicians from six continents with global outreach. We also warmly welcome our Assistant Editor, Anna Liu, who has championed the development of the journal and will be a key contact for both contributors and Editorial Board members. The team will be led by our Editor-in-Chief, John Frean from South Africa, and our Deputy Editor-in-Chief, Peter A. Leggat, AM, from Australia. We will be reviewing and building our Editorial Board and reviewing teams in the coming years and the Editor-in-Chief would welcome nominations from interested leaders in the fields of tropical medicine and infectious disease. We are also looking forward to receiving your research, reviews and other scholarly contributions and we welcome suggestions or ideas that can enhance this new journal.Tropical Medicine and Infectious Disease. On behalf of the Editorial Board, we invite you to submit your manuscripts and suggestions for Special Issues and guest editorials to


Dyslipidemia, an increase in pro-atherogenic low-density lipoproteins (LDL) and its oxidative modifications, oxLDL, is well known to be a crucial factor in endothelial damage, a key early step and a predictor of the development of atherosclerosis. It is also well-known that atherosclerotic plaques develop in regions of low and non-unidirectional disturbed blood flow as is found by branching points, bifurcations, and in the inner curvature of the aortic arch. Our studies demonstrate a novel paradigm for dyslipidemia-induced endothelial damage, an increase in endothelial stiffness induced by the uptake of oxidized lipids.via activation of the contractile RhoA/ROCK cascade, whereas enriching the cells with cholesterol has a rescue effect [Our previous studies showed that exposure to oxLDL in vitro or to plasma dyslipidemia in vivo dramatically increases endothelial stiffness –3. Furthe effect , known te effect . Most ree effect . Specifie effect .stiffness of the vascular wall, dominated by extracellular matrix and layers of smooth muscle cells, and stiffness of the endothelial monolayer. Previous studies established that dyslipidemia, aging and cardiovascular disease are associated with stiffening of the arterial wall of major arteries, attributed to remodeling of the extracellular matrix and vascular smooth muscles (VSMCs). However, while vascular stiffening is well recognized as an important factor in aging and development of the cardiovascular disease (CVD) [In terms of the functional significance, there is a critical distinction between the se (CVD) , the rolvia a different mechanism. Specifically, we found that endothelial stiffening but not the stiffening of the vascular wall could be abrogated by the genetic deletion of CD36 or blocking of oxLDL uptake indicating the leading mechanism of endothelial stiffening in aged vessels is accumulating uptake of oxidized lipids. These findings provide a model to discriminate between the roles of endothelial and sub-endothelial stiffness in age-related vascular dysfunction.Finally, we found that endothelial stiffness increases with age independently from the stiffening of the vascular wall . As was In summary, the novel concept of these studies is in discovering the impact of oxidized lipids/plasma dyslipidemia on endothelial stiffness and a new paradigm for synergism between pro-atherogenic flow and dyslipidemia in inducing EC dysfunction: the stiffening of aortic endothelium, mediated by increased expression of CD36 and enhanced oxLDL uptake. Dyslipedimia/aging-induced stiffening of aortic endothelial cells is proposed to constitute an important confounding factor in endothelial dysfunction independent of vascular stiffening.


The degree of substitution (DS) was calculated using elemental analysis data and showed higher DS values in modified dextrin than modified starch. The onsets of thermal degradation and temperatures at maximum mass losses were improved after modification of both starch and dextrin polymers compared to unmodified ones. Glass transition temperatures (Tg) of modified starch and dextrin were lower than unmodified control ones, and this was more pronounced in modified dextrin at a high molar ratio. Dynamic water vapor sorption of starch and dextrin polymers indicated a slight reduction in moisture sorption of modified starch, but considerably lower moisture sorption in modified dextrin as compared to that of unmodified ones.Pea starch and dextrin polymers were modified through the unequal reactivity of isocyanate groups in isophorone diisocyanate (IPDI) monomer. The presence of both urethane and isocyanate functionalities in starch and dextrin after modification were confirmed by Fourier transform infrared spectroscopy (FTIR) and  Amylopectin is larger than amylose with α(1–4) linked chains and up to about 5% of the glucose residues in α(1–6) branch units [Starch, as the main reserve source and energy storage of plants, is widely available in agricultural products. This biopolymer has excellent applications in many industrial sectors such as food, textile, pharmaceutical, paper, and biofuel. Most of these applications rely on the colloidal properties of two structurally distinct α-lopectin . Amylosech units . Starch ch units ,4.Over the last decades, starch has been modified chemically, physically, genetically, and enzymatically to enhance its functionalities and offer new applications. Chemical modification of starch has been carried out through cross-linking, acid hydrolysis, oxidation, etherification, esterification, cationization, and grafting of polymers ,10,11,12Tg) in unmodified starch but two defined Tg signals at −60 and 35 °C were obtained, which were respectively attributed to the relaxation of the grafted oligopropylene oxide chains and possible glass transition of the starch macromolecules in the network. However, modification of starch polymers with isocyanate functional groups has received very little attention. Barikani and Mohammadi  grafted Isophorone diisocyante (IPDI) is an aliphatic isocyanate that contains primary and secondary isocyanate groups with unequal reactivity. Modification of cellulose nanocrystals with IPDI monomer facilitated the covalent bonding between the modified cellulose and polymer matrix, and brought a greater mechanical strength than that of unmodified cellulose nanocrystals . Gómez-F13C nuclear magnetic resonance (13C NMR) in solid state, thermogravimetric analysis (TGA), differential scanning calorimetry (DSC), and scanning electron microscopy (SEM). The effect of functional modification on water vapor sorption behavior of modified samples was assessed by using a dynamic vapor sorption (DVS) apparatus.Enhancement of the unequal reactivity of IPDI isocyanate groups by using DBTDL catalyst should make possible a reaction with the functional groups of natural polymers, like the hydroxyl groups of starch, through their higher reactive sites . A pendant isocyanate group, which is sterically hindered by the neighboring methyl group, should then be available for reaction with additional monomers of interest. Therefore, this study aims at confirming the hypothesis by studying the highly functionalized pea starch and dextrin polymers with IPDI monomer and DBTDL reaction catalyst. Characterization of the modified starch and dextrin involved elemental analysis, Fourier transform infrared spectroscopy (FTIR), ®, kindly provided by Covestro, Leverkusen, Germany), with a NCO content of 37.8%. HPLC grade anhydrous dimethyl sulfoxide (DMSO)  was used as reaction medium and dibutyltin dilaurate DBTDL  catalyzed the reaction of hydroxyl groups from glucoside unities of starch and dextrin with the secondary NCO groups of IPDI. HPLC grade toluene  and tetrahydrofuran (THF)  were used to wash the polymers and remove the unreacted IPDI after modification.Pea starch and pea dextrin were kindly provided by Emsland-Stärke GmbH . The functionalization of starch and dextrin polymers with isocyanate groups was carried out using isophorone diisocyanate, IPDI  was then added to the IPDI and allowed to stir for 5 min. The dissolved starch was added dropwise to the stirring IPDI/DBTDL mixture with a separatory funnel over 45 min while maintaining the vigorous stirring in the flask. The reaction proceeded at 60 °C for 24 h in a flask with NThe carbon, hydrogen, oxygen and nitrogen contents of unmodified and modified starch and dextrin were determined by elemental analysis using an EuroVector EA 3000 atomic absorption spectrometer  heated at 980 °C with a constant flow of helium.−1 at room temperature, averaging 64 scans with resolution of 4 cm−1. The samples were dried at 50 °C for 24 h prior to the analysis.The chemical structure of the unmodified and modified starch and dextrin as well as of the IPDI monomer was analyzed with FTIR  provided with a MKII Golden Gate accessory  with a diamond crystal at a nominal incidence angle of 45° and ZnSe lens. Spectra were recorded in attenuated reflection (ATR) mode and the evaluation was performed between 4000 and 400 cm13C NMR using a Bruker Avance III 400 MHz equipment . The spectra were recorded using a decoupled sequence at 13C frequency of 100.6338 MHz at 25 °C. Samples were measured at a spinning rate of 10,000 Hz averaging 4096 scans with a recycling delay of 10 s. A time domain of 2 K was employed with a spectral width of 30 KHz.The insertion of isocyanate functional groups in starch and dextrin polymers were verified with solid-state −1, under a flowing nitrogen atmosphere.Thermal stability of the modified starch and dextrin was analyzed using a Q500 TA equipment . Around 5 mg of dried sample (24 h at 50 °C) were heated from 50 to 650 °C at a rate of 10 °C·minTg) of starch and dextrin associated with functional modification with IPDI were performed using a DSC analyzer , from −50 to 200 °C at a heating rate of 30 °C min−1 under a nitrogen flow of 10 mL·min−1. Approximately 5 mg of oven-dried sample (at 50 °C for 24 h) was used for each analysis.The changes in glass transition temperature  as reported previously ,21,22. AThe morphology of the starch and dextrin granules before and after modification was characterized by SEM using a JEOL JSM-7000F equipment  operating at 20 kV and a surrounding beam current between 0.01 and 0.1 nA, taking secondary electron images. Dried samples were placed over a double-sided carbon-based conductive tape and covered with a 20 nm Au in order to make the surface conductive.The results of elemental analysis and DS of the unmodified and modified starch and dextrin are given in 6 OH is more reactive and readily accessible than the secondary ones on C2 and C3. The C2 OH is however more reactive than the C3, mainly because the former is closer to the hemi-acetal and more acidic than the latter [Although the molar ratio of IPDI in reaction medium of MS 6.1 was two times more than MS 3.1, the DS value of MS 3.1 (0.55) was slightly lower than that (0.77) of MS 6.1. This might be explained by the different reactivity of the three –OH groups in a glucose unit of the starch macromolecule. The primary Ce latter . The hyd−1, which is assigned to the stretching vibration of –OH group, and at 2960 cm−1, which can be attributed to –CH bond stretching vibration [−1 was characteristic of the –CO stretching of the polysaccharide skeleton [−1, which indicated that many of these functional groups were consumed. This together with the appearance of the isocyanate band at 2266 cm−1, which is related to the pendant –NCO groups, indicates the success in functionalization starch and dextrin with IPDI [−1, and bending vibration of –NH and stretching vibration of C–N at 1530 cm−1 confirmed the presence of urethane bonds between IPDI and starch and dextrin [FTIR spectroscopy analysis was used to monitor changes in the structures of starch and dextrin polymers upon functionalization with the IPDI monomer. ibration ,25. The ith IPDI ,15,19. A dextrin ,15,26.6 [2, C3, and C5 [1 and C4 profiles are most sensitive to chain conformations [4 and C1, respectively [The structural changes of starch and dextrin polymers after modification were analyzed by means of solid NMR. 6 ,28. The , and C5 . Glycosirmations , and theectively . Modificectively . The appectively ,15. The Tonset) than the unmodified ones. The Tonset of S-control was 297.2 °C and it was increased to 313.2 and 324.4 °C in MS 3.1 and MS 6.1, respectively. D-control showed slightly lower Tonset (292.1 °C) as compared with S-control. Modification of dextrin also increased the respective Tonset of MD 3.1 and MD 6.1 to 318.9 and 319.8 °C (Tmax1) of the polymers occurred. These temperatures were 316.6 and 319.7 °C for starch and dextrin, respectively, and were directly associated with their thermal decomposition [Tmax1 to slightly higher temperatures at ~330–338 °C, which can be attributed to the decomposition of urethane groups into the isocyanate and alcohol groups [Tmax2) at ~413–434 °C for modified starch and dextrin. These results are in accordance with Girouard and coworkers, who found more than one Tmax in cellulose nanocrystals after functionalization with IPDI [Thermal degradation behavior of unmodified starch and dextrin as well as of the modified ones were examined by thermogravimetric (TG) and derivative thermogravimetric (DTG) analyses of samples that were dried previously in a vacuum oven at 50 °C for 24 h. 319.8 °C . The DTG319.8 °C b,d, and position . As indil groups ,34, and ith IPDI . ValodkaTg) of unmodified and modified starch and dextrin after removing the first history of samples are shown in Tg of unmodified starch and dextrin were found to be 113.2 and 89.3 °C, respectively. Previous studies quoted that the moisture content has a considerable effect on shifting the Tg of starch [Tg than former studies. The respective Tg of MS 3.1 and MS 6.1, however, shifted to 112.4 and 105.7 °C. Modification of dextrin caused further decrease in Tg, as MD 3.1 presented a Tg of 63.1 °C and MD 6.1 showed a Tg of 44.1 °C. The obvious decrease in Tg of modified polymers, particularly modified dextrin, might be attributed to the insertion of IPDI in the polymers’ backbones and creation of urethane linkages, which can increase the mobility of polymers due to lower amounts of hydroxyl groups, and thus reduce the hydrogen bonds [Tg. The glass transition temperature  are presented in 13C NMR confirmed both isocyanate and urethane functionalities in starch and dextrin macromolecules after modification. This study demonstrated remarkable improvement in thermal stability of IPDI-modified starch and dextrin. In addition, the moisture sorption of starch and dextrin was greatly reduced by IPDI modification. Functionalization of starch and dextrin polymers with IPDI make them suitable for a wide range of applications. These sustainable materials can be employed as matrices for composites or as reinforcement elements in other matrices without any further functionalization. For example, such polysaccharides would constitute promising substitutes for petroleum-based counterparts, e.g., in polyurethane foams and wood adhesive applications.Pea starch and dextrin were successfully modified with isophorone diisocyanate (IPDI) in order to reduce their hydrophilicity and improve their thermal stability. Assessment of degree of substitution (DS) from elemental analysis data indicated that most of the hydroxyl groups in starch and dextrin polymers reacted with IPDI. FTIR and


Tendon stiffness was measured during ramp and ballistic contractions. After fatigue task, joint stiffness significantly decreased by 20.7 %, whereas mEMG of measured muscles during drop jump did not. After fatigue task, active muscle stiffness with and without stretch reflex significantly decreased by 15.7 % and 21.5 %, and tendon stiffness measured during ramp and ballistic contractions did not change. In addition, the relative change in joint stiffness was significantly correlated with that in active muscle stiffness with stretch reflex , but not with those in the other measured variables. These results suggested that the decline in joint stiffness after repeated hopping exercises would be caused by changes in active muscle stiffness, but not those in tendon properties or neuromuscular activities.The purpose of this study was to elucidate the mechanisms of decline in joint stiffness after repeated stretch‐shortening cycle exercises according to changes in both muscle‐tendon properties and neuromuscular activities. Eleven males performed fatigue task (5 sets of 50 hopping). Ankle joint stiffness and electromyographic activities (mEMG) of plantar flexor and tibial anterior muscles during drop jump were measured before and after fatigue task. Active muscle stiffness with (100 deg·sec In orde1) Kubo, . Therefo1) Kubo, , we havee.g., Kawakami et al., i.e., drop jump) in order to directly clarify the mechanisms of a decline in joint stiffness after repeated SSC exercises.Since 2000, ultrasonography has been used to clarify the significant role of the muscle‐tendon interaction during SSC exersises throughout investigating the relationship between the muscle‐tendon properties and the SSC performence and the behavior of muscle fibers (fascicles) and tendons  would change after repeated SSC exercises, since these jump exercises consist of multi‐joint movements. Indeed, Kuitunen et al.  after repeated SSC exercises using a single joint. The purpose of this study was to elucidate the mechanisms of the decline in joint stiffness after repeated SSC exercises according to changes in both muscle‐tendon properties and neuromuscular activities. We hypothesized that a fatigue‐induced decline in joint stiffness would be associated with decreases in active muscle stiffness with the stretch reflex and muscle activities (including the stretch reflex) during jumping.In the previous studies as mentioned earlier, the contribution of each joint during the measurement of joint stiffness  of 0.8, it was shown that as least nine subjects were necessary for this study. Eleven healthy males  voluntarily participated in the present study. They were asked to avoid intense exercises for 3 days before the experiments. They were fully informed of the procedures to be utilized as well as the purpose of this study. Written informed consent was obtained from all subjects. This study was approved by the office of the Department of Sports Sciences, The University of Tokyo, and complied with their requirements for human experimentation.The sample size was estimated using the data from previous studies  and the mechanical properties of muscle‐tendon complex were measured before and after the fatigue task (see below).Subjects were initially familiarized with the testing procedures (see below) 1 week before the experiments. They performed two tests on 2 separate days, with at least 1 week between sessions and no longer than 2 weeks separating the two sessions. The order of the two experimental conditions  was randomized for each subject. The neuromuscular activity and behavior of fascicles during the drop jump . The load used was 50% of the body mass for each subject. They initially maintained a maximal plantar flexed position. They then exerted plantar flexion torque to maximal dorsiflexion, and rebounded to start plantar flexion until the toe finally lifted away from the footplate of this apparatus. These movements were repeated without a break. The fatigue task consisted of 5 sets of 50 hopping exercises with a rest interval of 1 min. The subjects were requested to jump as high as possible.Subjects performed a unilateral drop jump using only the ankle joint on a sledge apparatus, as described in our previous studies  to become accustomed to the test procedure before testing. The test was repeated five times before the fatigue task and three times after the fatigue task per subject, with at least 1 min between trials. The sliding table of this apparatus was moved to a height of 20 cm from the surface of the force plate to the sole of their foot with the assistance of an experimenter. Subjects were dropped from a height of 20 cm. After landing on the edge of the force plate, they arrested downward motion by eccentric plantar flexion. They then started concentric plantar flexion and took off. We excluded trials with slight knee joint flexion (>5 deg) based on images taken by the high‐speed video camera. The measured values were the means of two trials with the highest and second highest jump heights according to the flight time.The ankle joint angle was measured using open‐source image analysis software . The range of motion of the ankle from touchdown to the lowest position (ROM) was calculated. The ankle joint torque during the drop jump was estimated from the following equation  was used to continuously record longitudinal ultrasonic images of the medial gastrocnemius muscle (MG) during the exercises. At 30% of the lower leg length , the scanning probe  of the apparatus was secured with adhesive tape on the skin. Ultrasonic images were recorded on a video tape at 60 Hz, synchronized with recordings of a clock timer for subsequent analysis. The fascicle length was defined as the distance between the insertions of the fascicle into the superficial and deep aponeurosis. We analyzed the fascicle length of MG from pre‐landing (defined as 100 msec preceding landing) to when the toe lifted away from the force plate.In addition, we recorded electromyographic activities (EMG) from the lateral gastrocnemius muscle (LG), soleus muscle (SOL), and tibial anterior muscle (TA) with bipolar surface electrodes (5 mm in diameter) placed on the belly of each muscle with a 25‐mm interelectrode distance. The electrodes were connected to a preamplifier and differential amplifier with a bandwidth of 5 to 500 Hz . EMG signals were transmitted to a computer at a sampling rate of 1 kHz. EMG was full‐wave rectified and averaged for the duration of the contraction (mEMG). During jumping tests, mEMG values from LG, SOL, and TA were calculated from the pre‐landing, eccentric, and concentric phases, respectively, according to the ankle joint angle. In addition, the mean mEMG in LG and SOL was defined as the mEMG of the plantar flexor muscles (PF).‐1. In order to minimize thixotropic effects as preconditioning  was used to obtain a longitudinal ultrasonic image of MG at the level of 30% of the lower leg length during slow stretching. Ultrasonic images were recorded on a video tape at 30 Hz and synchronized with recordings of a clock timer for subsequent analyses. The fascicle length was defined as the distance between the insertion of the fascicle into the superficial and deep aponeuroses.The passive torque, joint angle, and fascicle length were continuously recorded during slow stretching. The slope of the portion of passive muscle force—fascicle length curve from 90 to 80 deg, was defined as passive muscle stiffness  at a 100‐deg ankle angle with a 2‐min rest period between each trial. The peak torque value within all trials was recorded as the MVC value.−1). For the 250‐deg·sec−1 condition, a 60‐ms period after stretch onset was analyzed in order to avoid any neural effects . During this period (110 msec), the range of motion was about 8 deg, and the angular velocity reached about 100 deg·sec−1.After a 5‐min rest period, measurements of active muscle stiffness were performed using a previously described procedure  at 50% of MVC with the visual aid of exerted torque on an oscilloscope. The measured values were the means of two trials.An additional measurement was performed two times at 0% MVC before the short‐range stretch experiment for data‐correction purposes. The averaged torque during the relaxed condition (caused by inertia and passive elasticity) was subtracted from the measured torque during each of the active stretch trials . At the level of 30% of the lower leg length, the scanning probe of the apparatus was fastened using adhesive tape on the skin. Ultrasonic images were stored at 98 Hz in the computer memory of the apparatus , with the right foot tightly secured to the footplate of the dynamometer by two straps. The ankle joint was set in a neutral anatomical position (90 deg) with the knee joint at full extension. After a warm‐up and submaximal contractions to become accustomed to the tests, subjects were asked to exert isometric plantar flexion torque at two different speeds of contraction, that is, ramp and ballistic contractions. For ramp contractions, they generated increasing torque from zero (relaxation) to MVC within 5 sec . Ultrasonic images were recorded as video signals at 60 Hz with a videotape recorder and synchronized with the torque and joint angle data by superimposing a clock timer for subsequent analyses. To assess the elongation of tendon structures (including outer tendon and aponeurosis), the movement of the intersection made by one fascicle and aponeurosis was measured. Even during isometric contraction, angular joint rotation occurred in the direction of ankle plantar flexion, resulting in the tendon displacement .Descriptive data are reported as means ± standard deviation. A two‐way ANOVA with repeated measures {2 (conditions) x 2 (test times)} and a post hoc analysis were used to analyze data. The F ratios for main effects and interactions were considered to be significant. Significant differences among means were detected using a Bonferroni post hoc test. In ANOVA, Mauchly's sphericity test was performed to assess the homogeneity of variance. Greenhouse‐Geisser correction was applied where assumption of sphericity was violated. Pearson's product‐moment correlation was computed to assess the relationships among the measured variables. The level of significance was set at P = 0.060), ROM significantly increased (P = 0.004), and the joint stiffness significantly decreased by 20.7% (P = 0.003) after the fatigue task. The relative change in joint stiffness was correlated with that in ROM  but not peak torque .Table P < 0.001) (Table P = 0.333) , but not the 250‐deg·sec−1 condition (P = 0.160) (Table −1 condition (P < 0.001), but not 100‐deg·sec−1 condition (P = 0.090) (Table −1 condition (P = 0.001) and 15.7% under the 100‐deg·sec−1 condition (P = 0.024) , the elongation of fascicle significantly decreased by 11.2% (P = 0.011), and the passive muscle stiffness significantly increased by 32.5% (P = 0.003) after the fatigue task , but not with that in the other measured variables  in leg and joint stiffness after submaximal and maximal SSC exercises . Therefore, decline in active muscle stiffness would lead to an increase in ROM during the measurement of joint stiffness. The results on active muscle stiffness under the 250‐deg·sec−1 condition suggest that intrinsic muscle properties without the effect of the stretch reflex changed after repeated SSC exercises. Dr. Komi's colleagues demonstrated that exercise‐induced muscle damage that resulted in changes in the neural effect through modulation of the stretch reflex would be associated with the regulation of joint stiffness , but not to those in the neural drive and neuromuscular activities. Indeed, Trappe et al.  decreased to the same degree as for the 250‐deg·sec−1 condition  after repeated SSC exercises. Furthermore, the relative change in joint stiffness was highly correlated with that in active muscle stiffness measured under the 100‐deg·sec−1 condition, but not that under the 250‐deg·sec−1 condition  was close to that under the 100‐deg·sec−1 condition. Therefore, we cannot deny that the change in active muscle stiffness was influenced by the angular velocity, and not the stretch reflex. At present, we are investigating the effect of the angular velocity on active muscle stiffness in our laboratory.In addition, the active muscle stiffness under the 100‐deg·secThe mechanical properties of tendon structures measured during ramp and ballistic contractions did not change after repeated SSC exercises. Previous studies demonstrated that the elongation and stiffness of tendons measured during ramp contractions did not change after repeated drop jumping and endurance running (Kubo et al., Before the fatigue task, the fascicles of MG shortened during the eccentric phase of the drop jump, whereas the whole muscle‐tendon complex lengthened Fig. . PreviouAnother interesting result of the present study was that passive muscle stiffness significantly increased after repeated SSC exercises. According to previous findings (Jones et al., In this study, we must draw attention to limitations and assumptions associated with the methodology followed. First, we used a previously reported moment arm length in order to calculate the Achilles tendon force. We cannot rule out the possibility that the moment arm length may change due to an acute increase in muscle thickness after the fatigue task. According to the finding of Sugisaki et al. , the momIn conclusion, a decline in joint stiffness after repeated SSC exercises would be caused by changes in active muscle stiffness, but not those in tendon properties or neuromuscular activities. In a future study, we must establish a new endurance training protocol for active muscle stiffness and joint stiffness in order to enhance performance in athletic events such as long‐distance races.The authors declare that they have no conflict of interest.