Light processable hydrogels were successfully fabricated by utilizing maize starch as raw material. To render light processability, starch was gelatinized and methacrylated by simple reaction with methacrylic anhydride. The methacrylated starch was then evaluated for its photocuring reactivity and 3D printability by digital light processing (DLP). Hydrogels with good mechanical properties and biocompatibility were obtained by direct curing from aqueous solution containing lithium phenyl-2,4,6-trimethylbenzoylphosphinate (LAP) as photo-initiator. The properties of the hydrogels were tunable by simply changing the concentration of starch in water. Photo-rheology showed that the formulations with 10 or 15 wt% starch started curing immediately and reached G’ plateau after only 60 s, while it took 90 s for the 5 wt% formulation. The properties of the photocured hydrogels were further characterized by rheology, compressive tests, and swelling experiments. Increasing the starch content from 10 to 15 wt% increased the compressive stiffness from 13 to 20 kPa. This covers the stiffness of different body tissues giving promise for the use of the hydrogels in tissue engineering applications. Good cell viability with human fibroblast cells was confirmed for all three starch hydrogel formulations indicating no negative effects from the methacrylation or photo-crosslinking reaction. Finally, the light processability of methacrylated starch by digital light processing (DLP) 3D printing directly from aqueous solution was successfully demonstrated. Altogether the results are promising for future application of the hydrogels in tissue engineering and as cell carriers Hydrogels are three-dimensional polymeric networks with the ability of retaining large amount of water within their structures without dissolving. Their crosslinked networks can be formed by either chemical or physical gelation process. The physical gel process involves weak Van der Waals forces, electrostatic interaction, and hydrogen bonding while the chemical gel formation leads to strong chemically bonded networks. Thanks to their hydrophilicity, hydrogels can be utilized by a broad range of sectors including agro-industry, pharmaceutical, medical, and food applications ,2,3,4,5.Photocuring techniques are green and highly efficient for crosslinking polymer resin with photo-curable functional groups. They have many advantages with respect to the thermocuring processes such as absence of Volatile Organic Compound (VOC) emissions, high curing speed, reduced energy consumption, and no heat requirements ,14,15. RStarch is an excellent precursor for preparation of hydrogels. GMA was utilized to methacrylate starch through either a transesterification reaction or by epoxy ring-opening mechanism ,29. Star3D printing by fused filament fabrication (FFF) has gained significant attention as a tool for designing biomedical scaffolds . More reHigh-amylose Hylon VII maize starch (70% amylose) was obtained from Ingredion, Goole, UK. Methacrylic anhydride (MA), triethylamine (TEA) (>99%), dimethyl sulfoxide (DMSO) (ACS reagent P99.9%), ethanol absolute, and bis(acyl)phosphane oxi lithium phenyl-2,4,6-trimethylbenzoyl phosphinate (LAP) were purchased from Sigma Aldrich .High amylose maize starch and DMSO were placed in a round-bottom flask under stirring condition (6 g of maize starch in 200 mL DMSO) and the temperature was increased to 70 °C. After 30 min the starch had been gelatinized and the solution was cooled down to room temperature. Methacrylic anhydride was then added dropwise with a syringe and triethylamine, as a nucleophilic catalyst, was added slowly (molar ratio AGU:MA:TEA of 1:2:0.04). The final solution was kept at room temperature for 18 h. After 18 h the product was precipitated in ethanol, dissolved in deionized water and again precipitated in ethanol. This procedure was repeated two times in order to purify the product. The final aqueous solution was lyophilized.2. The formed hydrogels were named according to the original starch content, i.e., MA-Starch5, MA-Starch10, and MA-Starch15, for the hydrogel prepared from aqueous solution with 5, 10, and 15 wt% of starch, respectively.To fabricate starch hydrogels, aqueous solutions containing 5, 10, or 15 wt% of functionalized starch were prepared. Testing of higher concentration was not possible due to solubility limit of MA-starch in water. The photo-initiator selected was lithium phenyl-2,4,6-trimethylbenzoylphosphinate (LAP), which was added in a content of 1 wt% with respect to the starch weight. The photocurable formulations were then placed in a silicon mold and cured for 1 min under UV-DIMAX lamp with a light intensity of 115 mW/cm2 with an exposure time of 5 s for a layer thickness of 50 µm.The MA-Starch10 formulation  was 3D printed using a lithography-based ASIGA UV-MAX DLP printer . The DLP printer had a light emission at λ = 385 nm and a XY pixel resolution of 62 μm. Approximately 6 mL of starch–water solution was placed in the printer vat. The UV light intensity was set to 10 mW/cm1H-NMR was conducted at room temperature. Approximately 7 mg of each sample were dissolved in 1 mL of DMSO-d6. The 13C-NMR was conducted at 60 °C with 20 mg/mL as starch concentration in DMSO-d6.Starch and MA-Starch were analyzed by Bruker Advance 400 Fourier Transform NMR spectrometer  operating at 400 MHz. The −1 with 4 cm−1 resolution.The FTIR spectra of starch, MA-starch and the crosslinked hydrogels were recorded by Perkin Elmer Spectrum 2000 FTIR spectrometer  equipped with a single reflection attenuated total reflectance (ATR) accessory (golden gate). The samples were scanned from 4000 to 500 cmThe tests were performed with an Anton PAAR Modular Compact Rheometer  using a parallel plate configuration (φ = 15 mm) with a quartz bottom glass. The gap value was set as 300 μm. All experiments were carried out at T = 25 °C. All rheological measurements were performed in triplicate. Oscillatory rheometer operating in time, stress and frequency sweep modes was used to monitor the viscoelastic properties associated with the crosslinking kinetics and to characterize the mechanical properties of the photo-crosslinked networks. The time sweep experiment was performed in the linear viscoelastic region (LVR) at a constant strain amplitude (γ) of 0.5% and a constant frequency (ω) of 6 rad/s to monitor the in-situ gel formation by following the evolution of elastic storage modulus G’ with time. After 30 s, the UV lamp was switched on.−2. The samples were irradiated from the bottom. An amplitude sweep experiment was also performed on the cured hydrogels γ [0.01–100%] at constant ω = 6 rad/s. After that, a frequency sweep experiment was performed on a freshly prepared sample; ω [0.1–100 rad/s] with a constant γ = 1%. It was possible to determine the crosslinked structure parameters using the storage modulus in the frequency independent plateau region (G’P). In this plateau, the energy applied is stored elastically by the polymer segments between two crosslinks that behave as entropic springs. In fact, the polymer chains are not capable of sliding by one another since the stress velocity is higher than the segment relaxation ability. The network parameters crosslink density (νe) and distance between two entanglement points (ξ) were calculated by Equations (1) and (2), respectively.The UV lamp used in the photo-rheology experiments was a Hamamatsu LC8 lamp  with a light intensity of 28 mW/cmA is the Avogadro’s number [R is the universal gas constant, T is the temperature, and Ns number .TM/10 Elite controller using TestWorks® 4 software  The unconfined uniaxial compression was performed with cell loaded of 10 N and a test speed of 0.5 mm/min. The samples tested had cylindrical geometry: φ = 10 mm X h = 11 mm. The data acquisition rate was set as 20 Hz. The compressive modulus was calculated by TestWorks 4 software. The modulus was estimated as the slope of the linear region of the stress–strain curves.The compression tests were performed with MTS QTestw%), the equilibrium swelling ratio (Seq), and the equilibrium water content (EWC) were calculated with Equations (3)–(5), respectively [s, Wd, and Weq are the weight of the sample in the swollen, dry, and swollen at equilibrium states, respectively.The swelling behavior was studied by means of conventional gravimetric procedure. Air-dried hydrogels were placed in deionized water at 25 °C. The weight increase was monitored during different time-intervals by taking the samples from the aqueous media and weighting them after the removal of free water on the surface with filter paper. The swelling ratio percentage S1 is the weight of the dry gel after the treatment with deionized water and W0 is the weight of dry sample before the treatment.The gel content percentage (G%) was calculated according to Equation (6):−1 in a nitrogen atmosphere (rate 50 µL/min). Thermal behavior of the samples was investigated using the following heating–cooling cycles: the temperature was raised from room temperature to 100 °C, then cooled down to −30 °C, and raised again to 200 °C. The glass transition temperature (Tg) was determined from the second heating curve.Differential scanning calorimetry was performed with a Mettler-Toledo DSC 820 . Samples having masses of approximately 7 mg were inserted in 100 μL aluminum pans with pierced lids. The applied heating rate was 10 °C min® CRL-2522™) were cultured on three different 96-well plates at a cell density of 2 × 104 cells/well for 24 h to reach confluence using DMEM-Dulbecco’s Modified Eagle Medium enriched with 10% fetal bovine serum and 1% penicillin/streptomycin . Simultaneously, three hydrogels prepared with three different methacrylated starch concentrations  were soaked for 24 h in 1 mL of DMEM. In each case, 0.1 g of hydrogel was soaked in 1 mL of the medium. After 24 h, the media were collected and filtered through 0.22 µm filters to guarantee sterility. Then, the culture medium was removed from each well of confluent cells and substituted with the supernatant collected from the hydrogels. Controls (CTRL) were obtained using normal medium. After 24 h of incubation, the supernatant was carefully removed and the cell viability assay was performed using non-fluorescent resazurin, which is converted into a highly red fluorescent dye (resorufin) by cell metabolism. Briefly, a volume of 100 µL of 0.1 mg/mL resazurin solution, obtained by diluting a resazurin working solution  into DMEM, was added in each well and the cultures were incubated for 1 h at 37 °C. Then, the fluorescent signal was monitored at 530 nm excitation wavelength and 590 nm emission wavelength using a plate reader . Cell viability was calculated as a percentage value compared to CTRL. Six samples for each condition were used and the experiments were performed three times. GraphPad Prism® software was used for one or two analysis of variance (ANOVA). Values * p < 0.05, ** p < 0.01, *** p < 0.001 were considered statistically significant.Cell viability assay was performed to evaluate the effect of the degradation products of MA-Starch hydrogels on cell viability. BJ human fibroblast cells  and methyl groups -CH3 (δ 1.9 ppm). These spectral changes are consistent with the structural modification of starch. However, as shown in The methacrylation reaction was evaluated and confirmed by spectrum  recorded1H-NMR peaks using Equation (7):1.9 and I5.11 are the integral intensity of CH3 protons (named b in the spectra) and the proton of the anhydrous glucose unit (AGU) unit in α-carbon (1H position). The formula is divided by three since there are three -OH groups in each AGU. The degree of substitution obtained was 0.08 , which can be translated into one methacrylated alcohol-group in every fourth glucose ring (The degree of substitution (DOS) for the hydroxyl groups was obtained as the ratio of integrals of the ose ring . This de13C-NMR spectra of Starch-reference and MA-Starch are shown in 13C-NMR spectrum of MA-Starch, there are additional signals from the methacryl-groups. These are seen at δ = 18.45 ppm (methyl carbon), δ = 136.61 ppm and δ = 127.75 ppm (double bond carbons) and δ = 170.38 ppm (C=O carbon).The −1 is attribute to the -OH vibration. The peak at 2925 cm−1 is the -CH2- stretching vibration. The band at 1638 cm−1 is the deformation vibration of the hydroxyl group of H2O absorbed in the amorphous regions of starch. The peak at 1148 cm−1 is the C-O-C symmetric stretching of cyclic ether group, while the peak at 1076 cm−1 is the C-H bending vibration. The peaks at 994 and 927 cm−1 are assigned to the stretching and skeletal vibration of the C-O-C α-1-4 glycosidic linkage, respectively. Finally, the peaks at 855 and 762 cm−1 can be assigned to the C(1)-H, CH2 deformation and C-C stretching vibration, respectively [−1 can be attributed to the C=O and C-O stretching vibrations, respectively, while the two peaks at 1640 and 815 cm−1 are assigned for the C=CH2 out of plane bending vibrations [The methacrylation reaction was further confirmed by FTIR. ectively ,43. In tbrations ,44,45.The methacrylated-starch was dispersed in water with LAP photo-initiator and the rheological properties of the starch/water/LAP formulations were recorded during the photo-crosslinking reaction to evaluate the reaction kinetics. Initially, an amplitude sweep experiment was performed on the liquid formulations to determine the linear viscoelastic region (LVR). The reaction is considered as completed when a G’ plateau is reached. c) are reported in The photocured hydrogels were fully characterized for their mechanical, rheological, and swelling properties. The compressive stress–strain curves of MA-Starch10 and MA-Starch15 photocured cast discs are shown in For rheological evaluation, the polymer solutions (300 μL) were crosslinked directly on the parallel plate for instant measurements. To determine the linear viscoelastic region of the hydrogels, an amplitude sweep experiment was initially performed. After that, a frequency sweep experiment was performed on a freshly prepared sample with a strain amplitude of 1%. The results of the frequency sweep experiment, including G’, G’’, and |η*|, are shown in p at ω = 10 rad/s and collected in The complex viscosities of the three hydrogels have approximately the same slope of −0.9, which means that all the networks exhibit a pseudo-plastic behavior . All theg)  see . There wHuman fibroblast cells were cultured on the supernatants from the three hydrogel formulations. Starch has previously been utilized in many biomedical applications for fabrication of scaffolds, hydrogels, and drug delivery systems ,53 showic = 13 kPa) close to the elasticity of muscle tissue (12 kPa) [The fast photo-curing reactivity of the formulations and the obtained mechanical properties indicated that the MA-Starch could be suitable photo-curable resin for the vat 3D printing technology . The 3D (12 kPa) . Simple (12 kPa) . In the 1H-NMR, 13C-NMR, and FTIR spectroscopy. The methacrylated starch was dispersed in water at different concentration in the presence of LAP photo-initiator. Photo-rheology confirmed rapid in-situ gel formation by following the evolution of elastic storage modulus G’ with time. The measured G’ plateau values were 1.5, 39.9, and 67.6 kPa for the hydrogels obtained in the presence of 5, 10, and 15 wt% of MA-starch. These values are comparable with other methacrylated biobased hydrogel systems reported in the literature. Rheological characterization of the crosslinked materials was performed and the network parameters were evaluated on the G’p at ω = 10 rad/s. Increasing the MA-Starch concentration increased the crosslink density, which led to decreased swelling capability. The supernatants of the methacrylated starch-based photo-crosslinked hydrogels did not exhibit cytotoxicity against human BJ fibroblast cells in any of the three tested hydrogel concentrations. Finally, the 3D printability of 10 wt% MA-Starch was successfully demonstrated. These findings are promising for future applications of light processed starch in tissue engineering and as cell carriers.Methacrylated maize starch was successfully light processed to hydrogels either by photo-curing with UV-lamp or by DLP 3D-printing directly from aqueous solution. First, starch was methacrylated and the reaction was confirmed by


Neurons in some sensory areas reflect the content of working memory (WM) in their spiking activity. However, this spiking activity is seldom related to behavioral performance. We studied the responses of inferotemporal (IT) neurons, which exhibit object-selective activity, along with Frontal Eye Field (FEF) neurons, which exhibit spatially selective activity, during the delay period of an object WM task. Unlike the spiking activity and local field potentials (LFPs) within these areas, which were poor predictors of behavioral performance, the phase-locking of IT spikes and LFPs with the beta band of FEF LFPs robustly predicted successful WM maintenance. In addition, IT neurons exhibited greater object-selective persistent activity when their spikes were locked to the phase of FEF LFPs. These results reveal that the coordination between prefrontal and temporal cortex predicts the successful maintenance of visual information during WM. Here, the authors show that beta-band coordination between prefrontal and temporal cortex predicts working memory performance. Moreover, inferior temporal neurons exhibits greater memory activity when coordination between these areas is high, suggesting that this interaction supports object memory maintenance. How these areas interact with sensory areas to maintain task-relevant information is not well understood. Neurons in prefrontal cortex exhibit sustained spiking activity that encodes the content of WM2. However, since feature selectivity in some PFC areas seems insufficient to match the resolution of WM, a sensory recruitment model of WM has been suggested, in which brain areas involved in sensory processing also contribute to memory maintenance4. Such models involve interactions between PFC and sensory areas during memory maintenance. Variations of this idea range from a more modular perspective, in which specific portions of PFC exhibiting memory activity send that signal to sensory areas6, or a split of abstract vs. detailed information7 (perhaps varying based on task demands8), to more distributed versions emphasizing the content-specific communication between areas rather than spiking activity within either9. These conceptual models are not necessarily mutually exclusive, but they all predict that memory-related activity in sensory areas should be related to WM performance, a prediction that is discrepant with much of the existing neural data. Studies have shown that even when the content of WM is present in spiking activity within sensory areas, it is either only weakly correlated11 or not correlated at all13 with behavioral performance, casting doubt on its role in memory maintenance.High-level brain areas such as prefrontal cortex (PFC) and parietal areas are believed to be central in the control and execution of behavioral plansIn order to examine how the PFC interacts with sensory areas in support of WM maintenance, we simultaneously recorded spiking activity and LFPs within the FEF and IT in monkeys performing an object WM task. We sought to determine which aspects of the neural response were most closely linked to successful WM maintenance. We found that neither the spiking activity nor the LFP power spectrum within either of these areas was a strong predictor of the animal’s performance on this task. However, the synchronization between the two areas , particularly in the beta band, was a key predictor of successful object memory maintenance. Even though the object selectivity within IT did not differ between correct and wrong trials, the timing of spikes in IT were coordinated with the phase of the FEF LFP, and when the two areas were more strongly locked, IT spiking activity showed greater selectivity for the object held in memory. Thus, beta band synchrony between IT and FEF predicted both behavioral performance and the strength of object-selective persistent activity in IT.16, we focused on visual stimuli that evoked the strongest visual responses in IT, as they are more likely to be involved in maintaining the memory of those stimuli. There was a correlation between object selectivity during the visual and delay periods  in two monkeys  using two single electrodes . All of the FEF and IT units in the analysis had overlapping RFs, both responding to visual stimuli in the FEF RF. To examine the neural basis of object WM, we trained two monkeys to perform an object delayed-match-to-sample (DMS) task Fig.  in whichods Fig. . Therefo17, the population of FEF units  exhibited persistent activity, measured as an increase in normalized firing rate (NFR) during the delay period compared to their baseline . Moreover, there was no significant difference in the delay period spiking between the correct and wrong trials . However, the magnitude of this selectivity did not vary with performance , alpha , beta , low-gamma , and high-gamma  frequency bands. During the delay period, FEF power in the high-gamma band was greater than baseline, while theta, alpha, beta, and low-gamma power was lower  and IT sites  using a wavelet transform (see Methods section). All effects were tested in the theta . An increase in alpha and beta phase locking between areas also occurred during the sample period compared to baseline ; PPL was calculated across time and frequency, and shuffle corrected (see Methods section). Importantly, for correct trials we observed a significant enhancement in beta band PPL during the delay period compared to baseline . No other frequency band showed similar significant object selectivity of the PPL , we found that the beta band PPL between the FEF and IT carried information about the identity of remembered objects , but not on wrong trials . The variability in phase difference values across sessions was also reduced on correct trials ; this difference was only present during the delay period . In addition, there was an increase in the beta band SPL between FEF LFPs and IT spikes during the Pref, In compared to the NPref, In condition, indicative of the object selectivity of this SPL . In contrast, neither the beta SPL between FEF spikes and IT LFPs, nor within-IT beta SPL, nor within-FEF beta SPL was predictive of performance . For FEF spikes and LFPs, only the high-gamma band SPL was significantly different for correct vs. wrong trials . To calculate SPL, each spike was assigned a phase value based on the simultaneously recorded LFP; SPL was the circular average of all spike phases at a particular frequency (see Methods section). We calculated SPL for both FEF spikes and IT LFPs, and IT spikes with FEF LFPs, but only the latter showed significant correlations with behavior Fig. . Figure nce Fig. . There w15. Beta coupling can also reflect the identity of the object held in WM18. We sought to determine if there was a relationship between beta coupling and the object-selective persistent activity in IT. To test for this relationship, we quantified beta coupling on individual trials (see Methods). We sorted trials by beta band PPL value and designated the third of trials with the highest PPL as “High” and the lowest third as “Low” trials (Methods). Using ROC analysis, we calculated the time-course of the object discriminability of IT spiking activity for High and Low PPL trials . For the Out condition, there was lower object discriminability on High vs. Low PPL trials  or FEF . We next sought to test whether this modulation by beta coupling was dependent on the strength of object discriminability of the IT unit. Across the IT population, for the In condition, there was a correlation between IT units’ object discriminability and the discriminability difference for High vs. Low PPL trials  or FEF . These results support the idea that beta coupling between the FEF and IT, unlike power within either area alone, predicts the maintenance of object identity within IT cortex.Object-selective persistent activity in IT has often been suggested as a basis of WM maintenanceals Fig. . For thePrefrontal areas are known to play an important role in WM. In order to understand how PFC interacts with sensory areas during WM, we simultaneously recorded from a prefrontal area (FEF) where neurons exhibit spatially selective persistent activity and from IT, where neurons exhibit object-selective persistent activity, during an object WM task Fig. . Whereas19, or interference from samples remembered during previous trials20. Effects of recent reward history, on the other hand, presumably influence choice during the target selection stage21. Errors in target selection could arise due to impulsivity or selection biases, either based on recent reward history or more persistent biases in target location or object identity. Despite this heterogenous mix of sources of errors, we were nevertheless able to identify inter-areal signatures that correlated with memory performance during the delay period. Observed differences between correct and wrong trials during the delay period may either reflect failures in memory, or failures to encode the stimulus. Clearly, if a sample is not encoded, it cannot be remembered, and thus error trials caused by failures of encoding should exhibit no memory-related activity. Errors in target selection following successful encoding and memory maintenance should result in mislabeling of trials with sufficient memory activity as ‘wrong’ trials, weakening the strength of behavioral correlations. Nevertheless, we observe significant behavioral correlations for neural signatures during the delay period, suggesting that either the magnitude of the effects we observed would be stronger when limiting analysis only to trials with failures of memory, or that there were comparatively few target selection errors.It is important to note that multiple sources of errors are possible during the object DMS task. Encoding errors could result from lapses in attention during the sample period, or if attention was incorrectly directed to the alternative sample location. During the delay period, there could be failures of maintenance22. The known influence of FEF neurons on the visual responses in visual cortex25, and the evidence that FEF neurons send memory-related spiking activity directly to visual areas26, further support the idea that FEF spiking activity has a direct impact on IT during WM. It is important to note that this is not the first time a spatial signal has been shown to enhance feature selectivity. In the case of covert spatial attention28, spatial cues have been shown to increase feature information in V430 and IT31. A similar gating of feature information by a spatial signal is observed when attention is exerted overtly, when the representations of targets of upcoming eye movements are enhanced35. More specifically, FEF spiking activity, lacking robust feature selectivity, has been shown to contribute to feature selectivity in these areas37. A spatial tag has also been suggested as a basis for binding object features39, including during short term memory41; consistent with this hypothesis, retroactive spatial cues can improve object memory46. In our data, coherence between IT and the FEF could reflect the deployment of spatial attention for feature binding during object memory maintenance48. However, in the absence of a competing stimulus, this FEF contribution might not be apparent49. These results suggest that perhaps, similar to the case of spatial attention , a non-selective signal highlighting relevant information is sufficient to boost object-selective spiking activity in IT.Our results support the idea that the interaction between prefrontal and sensory areas contributes to WM maintenance. Although we cannot rule out the possibility that one or more other areas drive WM selective signals in both the FEF and IT during WM, the findings that FEF leads IT in phase, and that inter-areal coherence predicts IT object selectivity, are consistent with a direct gating of object-selective persistent activity in IT by the FEF. This leading role of the FEF dovetails with previous results showing Granger-based causality of the FEF on IT during memory maintenance, but not the reverse58, leading to the suggestion that maintenance of the current state, whether motor or cognitive, is the unifying principle of beta oscillations59. Indeed, beta band coherence between IT sites is correlated with object WM performance in humans57. Such a content-agnostic role for beta oscillations in maintaining current states and information, whatever those might be, is consistent with the idea of a non-selective gain signal from the FEF gating object information within sensory areas. Note that despite the lack of feature selectivity in the FEF, we nevertheless observed spatial and feature selective coordination between the FEF and IT  demonstrates the robust involvement of this frequency band. Moreover, given their level of significance, the beta band effects remain significant even when subjected to multiple comparison tests  were used in this study. M1’s experiments were performed in the School of Cognitive Science at IPM, and M2’s experiments were performed at Stanford University. All experimental procedures were in accordance with the National Institutes of Health Guide for the Care and Use of Laboratory Animals and the Society for Neuroscience Guidelines and Policies. The protocols for all experimental, surgical, and behavioral procedures were approved by the Institute Fundamental Science committee for M1, and by the Stanford University Institutional Animal Care and Use Committee for M2. Structural magnetic resonance imaging  and CT scan (M1) were performed to locate the arcuate sulcus and prelunate gyrus for the placement of a recording chamber in a subsequent surgery. All surgical procedures were carried out under Isoflurane anesthesia and strict aseptic conditions. Prior to undergoing behavioral training, each animal was implanted with a stainless-steel custom-made chamber, attached to the skull using orthopedic titanium screws and dental acrylic. For M1, within the 30 × 70 mm chamber a craniotomy was performed . For M2, a craniotomy was performed within the a 20 mm diameter circular chamber (centered at 23 mm M/L and extended from 25 mm A/P to 5 mm A/P).60. Visual stimuli were presented on an LED-lit monitor , positioned 65.5 cm in front of the animal’s eyes. A photodiode  was used to record the actual time of stimulus appearance on the monitor. For M2, eye position was monitored with a scleral search coil and digitized at 500 Hz . The spatial resolution of eye position measurements was «0.1 deg. Stimulus presentation, data acquisition and behavioral monitoring were controlled by the CORTEX system. Visual stimuli were presented on a 29 × 39 colorimetrically calibrated CRT monitor (Mitsubishi Diamond Pro 2070SB-BK) with medium short persistence phosphors. A photodiode  was used to record the actual time of stimulus appearance on the monitor, with a continuous signal sampled and stored at 32 KHz.Animals were seated in custom-made primate chairs, with their head restrained and a tube to deliver juice rewards placed in their mouth. For M1, eye position was monitored with an infrared optical eye tracking system , with a resolution of <0.01° RMS; eye position was monitored and stored at 2 KHz. The EyeLink PM-910 Illuminator Module and EyeLink 1000 Plus Camera  were mounted in front of the monkey, and captured eye movements. Stimulus presentation and juice delivery were controlled using custom software, written in MATLAB using the MonkeyLogic toolboxThe fixation point, a ∼0.25 dva white circle, appeared either centrally or displaced by ±10dva in the horizontal or vertical axis, and the monkey maintained fixation within a ± 1.5 dva window for 1.5 s to receive a reward.62. Monkeys fixated within a ± 1.5 dva window around the central fixation point, and 10 objects (from 44 possible) were pseudo-randomly presented for 150 ms each with 100 ms between stimuli. Responses from the recording site in IT cortex were monitored audibly and visually by the experimenter. Stimuli evoking the highest response, an intermediate response, and little or no response were selected for use in the subsequent DMS task; final assignment of objects as Pref/Inter/NPref for analysis was based on responses during the DMS task. Only three objects were used per recording session in order to maximize the number of trials  for each condition.The RSVP task was used to quickly screen 44 sample objects and identify at least one which evoked IT visual responses for subsequent use in the DMS taskWe identified FEF sites by eliciting short-latency, fixed-vector saccadic eye movements with trains (50–100 ms) of biphasic current pulses . RF estimation was conducted by having the monkey fixate within a ±1.5 dva window around the fixation point , while stimulation was applied to evoke a saccade. The average endpoint of evoked saccades (landing points) was used to estimate the center of the FEF RF.63, and typically include the low eccentricity (<13 dva) contralateral visual hemifield in which all our FEF RFs were centered. Both FEF and IT neurons responded to stimuli placed in the estimated FEF RF .The DMS task is a widely used WM task; the version we use requires object WM. Monkeys fixated within a ± 1.5 dva window around the central fixation point. After 500 ms of fixation, a 3 dva sample object was presented either inside the FEF RF, or 180° away in the opposite hemifield, and remained onscreen for 300 ms. The animal then remembered the sample identity while maintaining fixation for 1 s (memory period) before the central fixation point was removed and two targets appeared (the sample object and a distractor). Target positions were rotated 90 degrees relative to the sample location, so no target appeared at the sample location, nor did the sample location predict the correct target location. The monkey had to shift its gaze to a ± 4 dva window around the sample object to receive a juice reward. If the monkey completed the trial but selected the distractor object, it was labeled a wrong trial. Note that in this version of the task, the position of the sample is irrelevant for task performance. In designing the task, we intentionally chose a long sample duration (300 ms) in hopes of minimizing encoding errors (estimated 50 ms/sample needed for encoding each item65. This was done by training a binary support vector machine (SVM) classifier using 70% of sorted spikes with an equal number of samples for each pair of clusters from a given recording site. The average classifier performance for the remaining 30% of samples across cluster pairs was taken as the isolation quality of sorted neurons in each session. The clusters of spikes with high isolation quality (>90%) were selected as well-isolated units66 . For M1 and FEF of M2, varnish-coated tungsten microelectrodes , shank diameter 200–250 μm, impedance 0.2–1 MΩ (measured at 1 kHz), were advanced into the brain for the extracellular recording of neuronal activity. For IT recordings in M2, a 32 gauge (235 mm outer diameter) guide tube was advanced ~15 mm through the brain (at a rate of ~0.75 mm/minute) by a custom-modified electronic motor-driven microdrive, stopping at or just above the upper bank of the Superior Temporal Sulcus. An electrode (75–100 μm diameter) was then advanced another 5–12 mm using a hydraulic microdrive (Narashige); high compressive strength was necessary for the electrode to exit the guide tube without bending. Activity was recorded extracellularly with varnish-coated tungsten microelectrodes (FHC) of 0.2–1.0 MΩ impedance (measured at 1 kHz). For M1, single-electrode recordings used a pre-amplifier and amplifier , filtering from 300 Hz–5 KHz for spikes and 0.1 Hz–9 KHz for LFPs. Spike waveforms and continuous data were digitized and stored at 40 kHz for offline spike sorting and data analysis. Extracellular waveforms were digitized and classified as single neurons using manual offline sorting . We also used cross-validation classifier performance to define a criterion for the accuracy of the well-isolated clusterss66 Fig. . Area IT67 during the DMS task. For IT LFP data, object selectivity in multiunit responses at each recording site was used to determine the suitability of the LFP data . As a result, LFP data from 8 IT sites  were excluded due to lack of object-selective multiunit activity, and LFP data from 15 IT sites  were discarded due to insufficient SNR. When comparing correct and wrong trials, some LFP data and units were excluded due to low numbers of wrong trials in a given condition ; number of sites and units included in each analysis are reported in the text with the statistics.Data are reported from 92 simultaneous FEF and IT recordings . For IT spiking data, Pref/Inter/NPref objects were determined based on the peak response across the visual and delay periodsAll analyses were carried out with custom code written in Matlab (MathWorks). The raw LFP data was low-pass filtered (1–300 Hz) and resampled at 1 kHz. The LFP for each site was normalized by subtracting the mean and dividing by the s.d. . Analysis focused on the 600 ms window in the late delay (300–900 ms after stimulus offset), in order to exclude both the stimulus evoked response and presaccadic activity (~100 ms before target onset); this is the time window used for analysis unless otherwise stated. In the trial-matching procedure, the number of correct and wrong trials was matched by subsampling the larger distribution 1001 times, and taking the mean of that distribution.For behavioral analysis we calculated performance based on the percentage of completed trials in which the monkey selected the correct target . Reaction times were calculated based on the time between target appearance and the eyes leaving the fixation window, and the Fig. For firing rate plots Fig. , individt) with the complex Morlet wavelet w to obtain the complex output signal y, also denoted as the analytic signal, where f0 denotes the desired center frequency of the wavelet function. We used a value of c = 7 wavelet oscillations. The center frequencies f0 were 1–130 Hz.To calculate LFP power and phase Fig. , we extrf effect we calculated Z-transformed values for each trial by subtracting the mean of the baseline and dividing by the standard deviation of the baseline across sites for each condition. The time-frequency map was smoothed with a window of 2 Hz in frequency and 100 ms in time. Then, Z-transformed values were averaged across sites.Time-frequency composition of the power spectrum was calculated by the amplitude of the analytic signal for each area. We averaged power values for each condition. In order to remove the 1/f.t) the Kendall correlation between FEF power and IT power across trials was calculated for each condition. In order to compare the conditions, we calculated Z-transformed correlation values by subtracting the value during the baseline and dividing by the standard deviation across pairs for each condition.The power-power correlation was calculated over time and frequency. For each point .The phase difference distributions were calculated similar to PPL, Eq. , except 55.SPL measurements in Fig. 68. N was the number of time points in the window. We subtracted the PPL value of the baseline from the delay period PPL to remove the 1/f effect. Trials were ranked based on single-trial PPL values within each condition. We selected trials with PPL <33th percentile as Low and trials with PPL >66th percentile as High trials. Object discriminability was quantified using the receiver operating characteristic (ROC) analysis69. ROC analysis was performed on the distributions of neuronal firing rate. The areas under ROC curves (AUC) when comparing responses to different sample objects were used as an index of object discriminability, and were calculated as in previous studies36. Sometimes there were not enough trials of a given condition with High or Low PPL to compute an ROC . For the timecourse of object discriminability of IT units, ROC was calculated for a jumping 300 ms window (10 ms steps). For Figs. n = 1001; p < 0.05). Figures High − AUCLow)/(AUCHigh + AUCLow). In Figs. P-value was calculated by permutation test (n = 1001). In Fig. In Fig. p-values and effect sizes are reported up to 3 digits and values below 0.001 are reported as p < 10−3. Effect size  or rank sum (for unpaired comparisons), unless otherwise specified. Statistical comparisons were performed across the population. For example, in comparing PPL for correct vs. wrong trials: for each LFP-LFP pair, PPL in each frequency band was calculated for correct trials, and for wrong trials. The Wilcoxon sign-rank test was then used to compare PPL values for correct trials vs. wrong trials across the population. Statistics for individual monkeys are reported in Table ze Table  was calcFurther information on research design is available in the Supplementary InformationPeer Review FileReporting Summary


There is a growing emphasis on diversity and its impact on health care. Our research has focused on migrants who are Mandarin speaking Chinese and Puerto Rican. Through a series of focus groups conducted in their native languages we discovered that barriers to service access differ across the two groups. In addition, many of these services are administered by individual states rather than the federal government. The diversity across migrant groups and state policies creates a challenge in using findings to establish national or international standards for best practices with older migrants. How then can we apply lessons about diversity and the impact of programs on a national and international scale? We propose the use of Social Interaction Modeling as a method for understanding patterns of behavior at both national and international levels, while preserving the unique character of each migrant group and the context within which they live.


All samples exhibited shear thinning non-Newtonian behavior. Butter proved its ability to maintain stiff network formation with starch molecules forming lipid-starch complexes characterized by high retention properties and increased stability due to high thixotropic and yield stress values. The pseudo-plasticity combined with high self-supporting ability  comprised by MP samples allows them to better behave during advanced technological processes such as extrusion 3D food printing.The effects of agar, alginate, butter, olive oil, and carrots on the mechanical and rheological properties of potato puree prepared by two different cooking methods (microwave heating (MP) and boiling (BP)) were investigated and interpreted in terms of starch microstructural changes. Microscopic observations revealed more aggregated and densely concentrated starch granules in MP samples. This consequently led to a significant increase ( Potatoes are considered the third most consumed food crop world-wide, and today due to the shift towards more convenient nutritious food in ready-to-eat meal markets, vegetable potato purees would serve as a good alternative for money-rich time-poor countries, among which are the European countries. In fact, the unique functional properties of potato puree highlight it as a potential substrate in many advanced food technological processes, such as extrusion based 3D printing. Potato starch are characterized by large-sized granules comprised of versatile biopolymers; highly branched amylopectin and linear amylose chains, making them easily malleable to external stresses and to further modifications, whether mechanical  or bio-chemical . Moreover, the abundant pectin substances in the middle lamella and the cellular wall of the potato cell facilitate the modulation of the textural properties of the starch under physical heat treatment  and widen its applicability as a thickening or gelling agent in the food technology sector Despite existing knowledge, accepting these perishable materials requires an intensive understanding of their physical properties. For example, studying the mechanical and rheological behavior is greatly necessary for developing an acquired product with the desired textural and quality characteristics. Additionally, this knowledge aids in predicting the analysis of process design and flow conditions in food processing and handling, such as in 3D printing . In fact, 3D printing is an innovative additive manufacturing technology that aims to produce innovative shapes with customized food structures and personalized food nutrition sources for an individual person. All the newly published research in the field of 3D food printing focuses on optimizing and characterizing the properties of the printed substrate for achieving best printability ,3. For iTherefore, in food industrial applications, potato purees are generally mixed with different food substrates to enhance their functionality, stability, and texture as well as improve its performance during processing by modifying its rheological-mechanical properties to recommended values . DetermiSolanum Tuberosum L cv Kennebec) were purchased from a local supermarket. Based on the primary visual inspection, potato tubers composing almost the same size, shape, and rigidity were selected. Tubers were then washed, peeled, and cut into cubes of approximately equal sizes. Afterwards, 100 g of potato cubes were subjected to two different cooking treatments: Microwave heating and boiling. For microwaving, potatoes were placed in a Sivica Porcelain Square  and then cooked at 700 W for 6 min in the absence of water whereas for boiling, potatoes were set in a beaker filled with distilled water at a proportion of 2:1 (w/w) at 98 ± 2 °C for approximately 20 min. Olive oil, butter, alginate, and agar were then added separately at 1% w/w for each type-cooked potato. Carrot puree was incorporated at a proportion of 1/3 of the potatoes. Carrots were boiled or microwaved respective to whether they were added to boiling (BP) or microwave heating (MP) groups. For each cooking treatment, each formula was prepared 4 times with the mentioned ingredients. All samples were set in an incubator (for approximately 25 min) until they maintain a temperature of 20 °C preceding any measurements.Fresh potato tubers . The micrographs were taken at 10× magnification.−1, (recording a maximum force) after which the probe returned to the original position [The mechanical characteristics , consistency (kg s), and cohesiveness (kg)) of the different blends were tested using a textural analyzer  coupled with a back extrusion cell and a 35 mm disc. Samples were placed in a standard-sized cylinder. During the test, the disc penetrated a distance of 30 mm at a speed of 2 mm·sposition . Each saThe rheological measurements were performed in a viscometer  controlled with commercial computer software . Steady rotational tests were performed to study the flow behavior, thixotropy, and yield stress. Yield stress was estimated as the point in which viscosity as a function of the shear stress (η = f(τ)) changed abruptly . The tem−1, infill speed = 40 mm·s−1, printing speed = 40 mm·s−1, flow % = 100, and retraction speed = 40 mm·s−1A RepRap BCN3D+ extruder printer  coupled with a syringe tool of 100 mL volume and 4 mm nozzle diameter was used to print 3D samples of potato purees. The code for the desired 3D object was transferred through an SD card from a CAD program (CURA 15.02.01). Speeds set in the CURA program were as follows: Travel speed = 100 mm·sQuality of 3D food printing was evaluated in terms of smooth printing (continuous extrusion without detachment), self-supporting structure (able to hold up its 3D shape properly post printing), and creamy surface appearance at the end.p < 0.05) using analysis of variance, one-way ANOVA, and Tukey’s HSD comparison test.Statistical analyses of the data were conducted using Minitab 18 . The data concerning textural characteristics were tested for significant differences  versus BP (822.8 g·kg−1) facilitated stronger gel formation, as reflected through the higher compacted microwaved starch granules observed under the microscope [In addition, the water content present in starch governs the internal microstructure and affects its textural and rheological properties . The lowcroscope . This dip < 0.05) from BP samples  which was able to elevate the viscosity approximately two times more than that of BP and MP. This could be attributed to the chain length as well as the degree of unsaturation of fatty acids that had different effects on the formation of starch lipid complexes. These starch lipid complexes are favored more by the presence of lower degree of unsaturation and shorter chain length of fatty acids , which cAdditionally, only agar combined with MP showed a distinct viscosity shape  compared to the other samples (instant decrease in viscosity) a. This cIn fact, BP samples expressed much lower yield stress values which could be attributed to the further scattered starch granules, acquiring a reduced fraction of force for cellular separation . In relaConversely, olive oil reduced the yield stress for both BP and MP. Olive oil has a malleable structure that facilitates its penetration inside the starch molecule, weakening the extent of bonding and producing a labile starch structure. In contrast, carrot addition maintained a similar yield stress regardless of the cooking treatment used. Nevertheless, the use of alginate caused a slight decrease in yield stress for MP while increasing that of the BP samples. This could be attributed to the compacted structure of MP granules stimulating more of the repulsive effect between the phosphorous groups on the potato and the anionic charges on the alginate, creating a less continuous network characterized by lower stability. Likewise, Liu et al.  reportedAll potato samples exhibited a hysteresis loop but with varying areas, indicating that all samples possessed thixotropic behavior. Again, MP samples possessed higher thixotropic areas compared to BP . It is aAgar showed an ability to produce the highest thixotropic areas for both BP and MP. The major effect between both cooking treatments was detected when butter was added. Generally, butter exhibits a strong thixotropic nature due to the action of two types of bonds that govern the fat crystal network and, thus, the lipid starch complexes: Irreversible bonds contributing to network stiffness and reversible bonds via Vander Waals interactions that would be disrupted under the shear rate effect but recover slowly via recrystallization . MoreoveMP + olive oil showed a higher hysteresis loop than MP alone, although olive oil demonstrated its ability to decrease the viscosity and yield stress of MP . This coIn 3D food printing trials of the different puree combinations, it was shown that MP samples exhibited better printability. The relative low viscosity that MP samples hold at high shear rates  in combination with the high cohesiveness and consistency values they possess facilitates their smooth continuous extrusion from a nozzle tip. Moreover, the stronger thixotropic and yield stress values expressed by MP samples enhances its internal material properties to reform rapidly once deposited and to minimize shape deformation under the hydrostatic pressure of consecutive layers ,21,22,23The incorporation of 1/3 carrot, 1% alginate, and 1% butter to the MP produced samples with similar viscosity ranges that provided smooth extrusion but printed end products with different degrees of stability, mainly related to differences at the level of yield stress and mechanical characteristic values. The lower mechanical characteristic values recorded by MP + 1/3 carrot made it more susceptible to deformation upon removal post-printing compared to alginate and butter samples, whereas the high yield stress value displayed by butter enhanced the self-supporting nature of the printed material while providing a creamy surface texture and attractive appearance . SimilarConversely, BP samples generally exhibited softer pastes , resulting in an overflow of sample deposition and hampered printing details. All printed BP samples showed a compressed bottom layer due to their poor ability to resist the gravity of subsequent extruded layers. The effect is an increase in diameter and decrease of height of the targeted printed product d compareMicrowaved and boiled potato samples possessed swollen gelatinized starch granules compared to raw potatoes, with microwaved potatoes revealing more aggregated and densely concentrated starch granules. The lower moisture content in microwaved potatoes provided a stronger inter-cohesive starch network, leading to higher values of mechanical and rheological properties.Agar and butter substrates were able to elevate the mechanical properties  as well as the viscosity, yield stress, and thixotropy of both MP and BP samples, due to the formation of a complex network between starch and additives.The ability of butter to possess relatively low viscosity (pseudo-plastic behavior) and to re-solidify when left to rest at room temperature due to its high thixotropic values as well as its strong internal elasticity (possess high yield stress), widens the applicability of butter as a potential substrate to be used in advanced food technological processes such as extrusion-based 3D food printing.BP samples showed smooth extrusion but with an overall deposition of material, characterized also by low textural properties to withstand the weight of the above printed layers, leading to hampered detailed-printed products.MP samples showed better printability in terms of smooth continuous extrusion and product shape self-retention. Yet, the best printability was attained with MP + 1% butter, providing an elegant printed product with a smooth creamy surface.The developed 3D printed products could be assessed as key formulas for delivering innovative shapes in restaurants or hotels and widen the applicability of more nutritive personalized colorful foods for children and elderly facing mastication-swallowing problems, thus promoting healthier snacks for them.


Dear Readers,Analytical Cellular Pathology (ACP). Please allow me to introduce myself before I outline my vision for our journal. I earned my PhD in Tissue Engineering/Molecular Biology in 2006 at University College London, UK. After completing my PhD, I served as a Postdoctoral Fellow at the University of Texas Southwestern Medical Center and later at the Schepens Eye Research Institute at Harvard Medical School. My research focus was on the development of new therapies to prevent the devastating effects of corneal fibrosis. In 2013, I moved to the Dean McGee Eye Institute, Department of Ophthalmology at the University of Oklahoma Health Sciences Center, where I currently reside. My research is focused on corneal trauma, corneal dystrophies, and the development of noninvasive therapeutic agents. Throughout my career, I have given priority to interdisciplinary, translationally relevant approaches. I strongly believe that science is moving towards embracing these approaches and that they are key for future success.I am delighted and honored to begin my role as Chief Editor for I would like to apply some of those approaches, as we move forward with revamping ACP. A journal is only as good as its editorial team and the quality of its reviewers. Hence, building and maintaining a strong team of editors alongside a supporting team of editorial staff are the minimum requirements for a successful journal. My vision for ACP, as we move into this new chapter, is to attract contributing authors that will provide us global recognition and ensure that our journal is competitive, exciting, and useful for the scientific community. I, therefore, will do what I can in order to raise the journal's profile, maintain high standards in our publications, and increase our reach. Excitingly, our recent impact factor has been released  and showIn terms of ACP's aim and scope, as a multidisciplinary, translational research journal, our focus should be on topics and areas that scientists, clinicians, and industry have prioritized. We have therefore updated those statements and will soon be available on our website.The field of cellular pathology is broad and involves a whole spectrum of clinicians, scientists, and engineers. Our journal faces several challenges, which we will try and navigate through together, in the coming years. Perhaps the most significant is convincing enough young people to enter the field of cellular pathology and contribute to future discoveries and treatments. With translational medicine becoming popular and human studies a must, our journal has to attempt to lead the field to the new era. This approach will be one of the strategies, if not the only one, that will keep us competitive and allow us to grow as a unit.The renewed ACP will focus on cross- and multidisciplinary publications that have the potential to revolutionize treatment of diseases. We will actively create opportunities for all those who are interested to submit and publish their work with us. The team and I are pushing for progress, we are pushing for excellence, and we will need all the help we can get from our Editorial Board members. In this era, communication is critical, and we will do what we can to keep the journal visible in medical and scientific communities. I understand that change is always difficult, but the challenges are too great and we must embrace the future.In closing, I would like to extend my thanks to the team at Hindawi for this wonderful opportunity. I look forward to working with each team member in the future. On a more personal note, I would like to thank friends and colleagues who have always been very supportive of my decision to assume this role. I will continue to look to each of you for guidance and support as I begin this new task.I am very excited about the future of ACP, and I look forward to working with our editorial team, the authors, the reviewers, and our readership as we continue to grow as a journal.Dimitrios Karamichos


Skin grafting is an essential aspect of burn and wound reconstruction. Split-thickness skin grafts (STSGs) harvested from the scalp are used for wound and burn reconstruction. Skin grafts from the scalp bear hair and hair particles. Residual hair fragments and pieces of hair in the graft have been associated with many complications, including foreign body reaction similar to pseudofolliculitis and chronic inflammation that can lead to infections. It is important to remove the hair and the hair particles from the scalp graft before its application to the donor site. Traditionally, surgeons have employed some techniques including saline agitation and mechanical removal of the hair particles with forceps. These techniques are time consuming and can subject the graft to mechanical damage. There is another technique that has been described using an adhesive tape. This technique uses Ioban™ , followed by a saline wash to remove hair from grafts prior to grafting. In this paper, we introduce a novel technique for intraoperative hair depilation prior to graft application to recipient site. We used Dermabond®  to remove residual hair particles from the STSG donor. Our technique has several advantages: it is expeditious, it allows minimal mechanical damage to the graft, and can be used for patients with allergies to Ioban. Intraoperative Dermabond depilation of scalp STSGs is safe, easy, and effective. The scalp is one of the many sites for obtaining split-thickness skin grafts (STSGs) for burn reconstruction and wound reconstruction. The scalp donor site has many advantages, including rapid wound healing, low risk of complications, and excellent cosmesis . HoweverAn STSG is harvested from the scalp using a dermatome. The graft is wetted with saline and placed on a flat surface. The graft is then patted dry using a dry gauze. A thin layer of Dermabond is then applied to the epidermal surface of the graft. The Dermabond is allowed to dry and forms a thick coat on the graft Figure . It takes approximately three to four minutes for the Dermabond to dry. Beginning at an arbitrary area within the graft, the Dermabond is meticulously peeled off the graft using Adson or Bishop-Harmon forceps. Once an edge is developed, the Dermabond peels off easily. The hair fragments remain firmly embedded within the dried Dermabond, revealing an almost complete hair-free graft Figure .Most of the hair was removed with one application of Dermabond. The few remaining hair fragments were easily removed upon rinsing the graft in saline. No graft sustained any punctures or tears indicative of mechanical injury. Each graft was rinsed again to ensure that any residual hair fragments or particles of Dermabond were removed. This resulted in an STSG of the scalp, devoid of any embedded hair and mechanical damage . The depilated grafts were then grafted to the recipient site.The scalp is a hair-bearing area, and STSGs from the scalp contain a large amount of residual hair particles after harvesting. Residual hair within the graft has been associated with foreign body reaction similar to pseudofolliculitis -4. The hMany techniques have been used intraoperatively to remove hair from grafts prior to graft application. These techniques include the use of forceps to remove the hair and saline agitation. The most recent technique described in the English language literature involves the use of Ioban and saline to remove hair from scalp STSGs . This teIn this article, we present a technique for intraoperative hair removal from the scalp donor STSG. This method uses Dermabond and saline washing for graft hair removal. Dermabond is a topical skin adhesive containing 2-octylcyanoacrylate . To the Intraoperative Dermabond depilation of STSGs is safe, easy, effective, and expeditious.


Objective: To report the University of Florence technique for robot-assisted kidney transplantation (RAKT) from living donor (LD) and deceased donor (DD), highlighting the evolution of surgical indications and technical nuances in light of a single surgeon's learning curve.Materials and Methods: A dedicated program for RAKT from LDs was developed at our Institution in 2017 and implemented later with a specific framework for DDs. All RAKTs were performed by a single highly experienced surgeon. Data from patients undergoing RAKT between January 2017 and December 2019 were prospectively collected in a dedicated web-based data platform. In this report we provide a comprehensive step-by-step overview of our technique for RAKT, focusing on the potential differences in peri-operative and mid-term functional outcomes between LDs vs. DDs.Results: Overall, 160 KTs were performed in our center during the study period. Of these, 39 (24%) were performed with a robot-assisted laparoscopic technique, both from LDs (n = 18/39 [46%]) and from DDs (n = 21/39 [54%]). Eleven (11/39 [18%]), 13(13/39 [26%]), and 15 (15/39 [30%]) RAKTs were performed in 2017, 2018, and 2019, respectively, highlighting an increasing adoption of robotics for KT over time at our Institution. Median time for arterial (19 min for LD and 18 min for DD groups), venous  and uretero–vesical (18 min for LD and 15 for DD) anastomosis were comparable between the two groups , as the median rewarming time . The rate of postoperative surgical complications according to Clavien–Dindo classification did not differ between the two study groups, except for Clavien–Dindo grade II complications . Overall, 7/39 (18%) patients  experienced DGF; two of them were on dialysis at last FU.Conclusions: Our experience confirms the feasibility, safety, and favorable mid-term outcomes of RAKT from both LDs and DDs in appropriately selected recipients, highlighting the opportunity to tailor the technique to specific recipient- and/or graft-characteristics. Further research is needed to refine the technique for RAKT and to evaluate the benefits and harms of robotics for kidney transplantation from DDs. While still being considered an experimental procedure , robot-aTo date, most groups performing RAKT replicated the Vattikuti-Medanta technique . NeverthIn this report we describe the University of Florence technique for RAKT from both LD and DD, highlighting the evolution of both indications and technical nuances over a 3-year period, as well as the intraoperative, perioperative and mid-term functional outcomes.A dedicated program for RAKT was developed at our Institution in 2017 .After Institutional Ethical Committee approval and obtained the patients' informed consent, data of patients undergoing RAKT from LD or DD were prospectively collected in our institutional web-based database. All consecutive patients undergoing RAKT between January 2017 and December 2019 were included in the analytic cohort .n > 100) and robotic urological surgery . The first LD-RAKT was performed on January 25th, 2017, while the first DD-RAKT on October 16th, 2017.At our center, the KT team includes 4 surgeons; however, all RAKTs were performed by a single surgeon (G.V.) with high experience in open KT  age <18 years; (b) absolute contraindication for robotic surgery; c) severe atherosclerotic plaques at the level of iliac vessels and (for DD-RAKT), and (d) lack of all key logistical requirements including availability of the operating theater .Regarding DD-RAKT, our program included both donors after brain death (DBD) and donors after circulatory death (DCD). However, since 2019, RAKT has been prioritized for DBD donors in light of a potentially higher risk of adverse peri-operative outcomes in the DCD donor population .Preoperative evaluation of donors, postoperative management, and follow-up after RAKT were performed according to the principle of the latest EAU Guidelines on KT .All recipients underwent CT-angiogram to assess the vascular anatomy, potential atherosclerotic plaques and anomalies of iliac vessels before RAKT .Chronic Kidney Disease Epidemiology Collaboration formula was used to calculate estimated glomerular filtration rate (eGFR) in patients aged <70 years , while tWarm ischemia time was defined as the time from the circulatory arrest of the organ procured to the begging of cold storage, while cold ischemia time as the time of cold storage, until its interruption before graft introduction into the recipient.Surgical postoperative complications were classified according to the modified Clavien–Dindo system .Delayed graft function (DGF) and primary non-function (PNF) were defined as the need of dialysis in the first postoperative week and the need for dialysis after RAKT with ultrasonography confirming adequate perfusion of the graft, respectively.In the context of DDs, the following key phases were crucial to evaluate the technical and logistical feasibility of RAKT:Phase 1 (team): after the Regional Allocation Center for Organs and Tissues provided the alert of a potential donation, the feasibility of RAKT depended on the availability of the dedicated surgical team.- Phase 2 (recipient): in the meantime, the recipient was admitted to the Nephrology Unit and a careful preoperative evaluation, including anesthesiologic work-up and CT scan, was performed to ensure that all patients' selection criteria are respected.- Phase 3 (robotic operating room): the feasibility of RAKT also required that the robotic operating room was available. Of note, our robotic OR is available for RAKT from DDs with no restrictions during the night-time and in the week-ends.- Phase 4 (cold ischemia): RAKT should be started within 16 h from the beginning of cold storage, keeping the overall cold ischemia time >24 h.- Phase 5 : RAKT was performed only after careful evaluation of the graft at bench surgery, provided that no highly complex graft vascular anatomy  was present. In this case, open KT is still preferred as multiple anastomoses might potentially increase the rewarming time during RAKT.- For LD nephrectomy, the da Vinci Si robotic platform  in a three-arm configuration was used, while for DDs we followed the European Association of Urology (EAU) Section of Transplantation (ESTU) guidelines for organ procurement . Organ pDuring preparation of renal vessels at bench surgery, the anterior margin of the vein was reshaped by cutting away a slice of venous tissue to improve visualization of its posterior margin to facilitate the subsequent venous anastomosis.In case of right-sided grafts from DDs, the length of the graft renal vein was increased during bench surgery through an inferior vena cava (IVC) patch.While at the beginning of our experience of RAKT from DDs we always used the Carrel's patch to facilitate the arterial anastomosis, from the 11th case onward (provided that no multiple arteries were present), the patch was always removed . As the During bench surgery, in case of multiple vessels, two techniques were used according to the graft vascular anatomy and surgeon's preference: (I) reconstruction of a single arterial vessel using the pantaloon technique to allow the performance of a single anastomosis in case of two arteries close to each other of approximately equal caliber; (II) modeling of the aortic patch to allow the performance of a single anastomosis in case of two arteries close to each other from DDs . In caseAn ureteral stent (5 F/14 cm) was placed during bench surgery to facilitate uretero–vesical anastomosis during robotic transplantation.The graft was finally wrapped in a gauze jacket filled with ice, with the renal artery fixed to the gauze with a landmark stich, to improve the visibility for the subsequent venous anastomosis.RAKT was performed using either the da Vinci Si or Xi robotic platform  in a four-arm configuration.Currently, we used a 0° lens and set the Trendelenburg tilt at 20°. When RAKT was performed in the left iliac fossa, an additional lateral tilt of the operative table of 5°-15° was used to better expose the left iliac vessels .Our surgical technique follows the principles of the Vattikuti Urology Institute-Medanta technique , 2 with Specifically, port placement followed the principles of robot assisted radical prostatectomy (RARP), with only one 12 mm port for the bed-side assistant and a variable triangulation toward the transplantation site at right or left iliac fossa. Pneumoperitoneum pressure was set at 8–10 mmHg and maintained constant through the use of the Airseal system. The da Vinci robot is docked between the patient's legs (Si platform®) or on the lateral side (Xi platform®), according to surgeon's preference and/or patient-specific characteristics.For the first 17 cases, a 5 cm periumbilical incision was made on the midline to place the GelPOINT™ device , to allow the introduction of the graft into the peritoneal cavity.Thereafter, to reduce unnecessary costs, from the 18th case onward, the Alexis® retractor  was used. Moreover, from the 18th case, a Pfannestiel incision was performed  to place the Alexis® device, allowing (I) better aesthetic results, (II) easier placement of the graft directly into the iliac fossa site, and (III) a direct access for the bed-side assistant to the operative field in case of intraoperative complications .In case of recipients affected by Autosomal Dominant Polycystic Kidney Disease (ADPKD), not requiring previously native kidney nephrectomy, port placement resulted slightly different, to tailor the surgical strategy to the specific patient's anatomy .After port placement, the peritoneum was incised at the level of the external iliac vessels, and both the external iliac artery (EIA) and external iliac vein (EIV) are skeletonized. For LD RAKT, extent of iliac vessels skeletonization was limited to the transplantation site, while for DDs (both DBD and DCD) it was extended until the bifurcation of the iliac vessels. Then, an extraperitoneal pouch was created for subsequent allocation of the graft after completion of vascular anastomoses. The Retzius space was developed, and the bladder prepared for the subsequent uretero–vesical anastomosis.After this step, the robotic instruments are temporarily removed, the robot is undocked, and the Pfannenstiel incision is performed to allow placement of the Alexis® retractor or the GelPOINT device . This suThen, the graft is gently introduced into the abdomen through the Alexis® device, with the hilum oriented toward the site of the iliac fossa, adding 250 mL of ice into the abdominal cavity to achieve regional hypothermia.A pictorial overview of our technique for venous anastomosis is described in a previous publication . In someAfter careful evaluation of the site for venous anastomosis and placement of bull-dog clamps, a venotomy is performed on the EIV. Of note, for the first 8 cases, Potts scissors were used to perform the venotomy. Afterward, the venotomy was performed by using cold scissors, that allowed an easy modeling of the venotomy as well as a reduction in terms of costs for surgical instrumentation.After flushing of the lumen of the external iliac vein with heparinized saline, venous anastomosis is performed in an end-to-side fashion to the external iliac vein using a 6-0 GORE-TEX suture  on a CV-6 TTc-9 needle . TechniqAn overview of the steps for arterial anastomosis is described in a previous publication . After mMoreover, at the beginning of our robotic program for DD RAKT, we performed arterial anastomosis using the Carrel's patch, provided that no significant atherosclerotic plaques were present at the level of the renal artery ostium during bench surgery.Nowadays, the arterial anastomosis during RAKT from DDs is usually performed without the aortic patch. This approach might result in an easier anastomosis due to: (i) the shorter arteriotomy; (ii) the more anatomic anastomosis thanks to the similar caliber and structure of graft renal artery and external iliac artery, and (iii) the reduced risk of significant atherosclerotic plaques at the level of graft renal artery as compared to Carrel' s patch  Figures.The renal artery is anastomosed in an end-to-side fashion to the external iliac artery using a 6-0 (or 5.0 in case of DDs) GORE-TEX suture on a CV-6 TTc-9 needle with two half running sutures using two different threads .The integrity of the venous and arterial anastomoses as well as the graft revascularization are checked through a macroscopic inspection for color and turgor. Moreover, to assess the graft and ureteral reperfusion, intraoperative duplex ultrasound and intraoperative fluorescence vascular imaging with indocyanine green (FireFly™ fluorescence imaging for Xi robotic platform) are employed, as previously reported , 17, 18.After checking of the ureteral reperfusion using fluorescence imaging, the uretero–vesical anastomosis is made according to a modified Lich–Gregoire technique over the pre-placed JJ stent creating an anti-refluxing mechanism, as previously described , 3.By Institution protocol, all recipients after KT are admitted in intensive care unit (ICU) and then in the Nephrology and Dialysis Unit for the post-operative course. A standardized institutional follow-up protocol, including serial duplex-US, was applied in all patients .Descriptive statistics were obtained reporting medians  for continuous variables, and frequencies and proportions for categorical variables, as appropriate.U tests, as appropriate.Differences in baseline donor, graft, and recipient characteristics, as well as differences in peri-operative, and functional outcomes between the LD and DD RAKT cohorts were assessed using the Pearson's Chi-square and Mann-Whitney Due to the limited sample size, no formal comparison of intra- or postoperative outcomes between robotic and open KT from DDs was performed.p < 0.05.Statistical analyses were performed using SPSS v. 24 . All tests were two-sided with a significance set at Overall, 160 KTs were performed in our center during the study period. Of these, 39 (24%) were performed with a robotic technique  . ConsideNotably, in our series RAKT from DDs was never impeded by the unavailability of the robotic OR. Yet, during the early phase of the learning curve, two patients were not deemed eligible for RAKT despite the availability of the surgeon due to a previous KT in the right iliac fossa (open KT in the left iliac fossa was preferred in both cases).Preoperative donor, recipient and graft characteristics are shown in Median (IQR) cold ischemia time (CIT) was 1 h (1–1.5) for LDs while 17.5 h (16–19.5) for DDs. Median (IQR) BMI was 26 (21–27) and 24 (21–25) in the LD and DD groups, respectively.p = 0.3).Overall, in the setting of LD RAKT, the graft was introduced through a periumbilical incision in 33.3% while through a Pfannestiel incision in 66.7%. These proportion were 19 and 81% in the setting of DD RAKTs  and the left iliac fossa in 3 cases (14.3%).p = 0.3). Similarly, median (IQR) rewarming time was not significantly different among the two cohorts .Overall, median (IQR) console time was 190 (160–240) and 180 (160–220) min for LD and DD RAKTs, respectively (p = 0.008) (Median (IQR) overall operative time was 270 min (225–365) and 240 min (200–255) in the two cohorts, respectively (= 0.008) .p > 0.05).Of note, as shown in Moreover, as depicted in In one case from a DD (1/39 [2.5%]), an intraoperative bleeding requiring the positioning of an extra 5 mm trocar was recorded, without need for intraoperative transfusions or conversion to open surgery.p = 0.042). Nevertheless, of Clavien–Dindo grade II complications, only one  in the LD group and two (2/16 [12.5%]) in the DD group were represented by peri-operative blood transfusions.Specifically, the rate of postoperative surgical complications according to Clavien–Dindo classification (CDC) did not differ between the two study groups, except for the rate of Clavien–Dindo grade II complications, which were higher among patients undergoing RAKT from DDs  major surgical complications were recorded. Of these, 4 (2 in the LD and 2 in the DD group) were Clavien–Dindo grade IIIa and 1 IIIb Clavien–Dindo grade complication (in the DD group). In particular, 2 recipients required percutaneous placement of a nephrostomy tube with antegrade placement of a JJ stent; in one recipient a symptomatic lymphocele was treated with percutaneous drainage and in one patient a transplant arterial stenosis (TRAS) required percutaneous transluminal angioplasty (PTA) and stenting. One patient in the RAKT DD group required graft nephrectomy due to an arterial thrombosis  .Regarding functional outcomes, 7/39 (18%) patients  experienced DGF. Two of them were on dialysis at last FU .2 in the LD and DD groups, respectively (p = 0.6).An increasing trend of eGFR was progressively recorded from the first postoperative day to the last FU . In partAlthough RAKT is still being performed only at selected referral Centers worldwide, current evidence confirms is feasibility, safety and reproducibility , 19, 20.Nevertheless, the introduction of RAKT might be challenging from surgical, economical and logistical perspectives; such challenges may indeed hinder its widespread adoption in Transplant Centers. In particular, the availability and organization of a robotic operating room (OR) in condition of “emergency,” coupled with a compelling need to check the donor-, recipient-, and graft-related selection criteria for RAKT in a relatively short timeframe (as well as to keep CIT to a minimum) may represent barriers for implementation of RAKT programs from DDs.As such, there is still lack of evidence on the outcomes of RAKT from DDs, though extending the number of minimally invasive KTs in both Urologic and Transplant Centers is becoming an increasingly relevant need.In this scenario, taking advantage of previous pioneering studies , 2, in 2In this report we reported an updated step-by-step overview of the University of Florence technique for RAKT and its mid-term outcomes.A key finding from our experience is that RAKT was technically and logistically feasible in a wide range of several clinical settings . Of note, all RAKTs in our series were performed by a single surgeon; the analysis of his learning curve allows to transparently evaluate the steps required to translate the knowledge and expertise in robotic urologic surgery to KT. As a proof of concept, no recipients required conversion to open surgery, and RAKT was successfully performed even at challenging times  in case of DDs. While the technique for RAKT did not significantly differ between LDs and DDs, except for specific nuances such as the management of the Carell's patch , from a From a surgical standpoint, the codified University of Florence technique for RAKT represents the evolution of the Vattikuti-Medanta technique with specific technical modifications introduced by our surgical team over time. Such modifications were introduced with the aim to simplify the technique, improving its reproducibility and allowing a larger number of (robotic) transplant surgeons to safely approach the procedure. In brief, the main technical nuances introduced by our group included: (a) the performance of a Pfannestiel incision for graft insertion into the abdominal cavity, to have direct access to the iliac fossa while improving the cosmetic result; (b) the systematic use of two single threads for performance of the arterial anastomosis, to ensure a watertight anastomosis avoiding the risk of reducing its caliber; (c) the removal of the Carrel's patch during bench surgery in cases of DD-RAKTs; (d) the performance of the venous anastomosis without the IVC patch in few cases of right-sided grafts from DDs; (e) the use of intraoperative fluorescence vascular imaging to assess the graft and ureteral reperfusion. Notably, intraoperative fluorescence vascular imaging may allow surgeons to tailor the length of the ureter according to the degree of its vascularization before uretero–vesical anastomosis, aiming to reduce the risk of subsequent stenosis of the uretero–vesical anastomosis. While promising, this technique requires further evaluation .Our experience also suggests that the financial burden of RAKT might be at least partly reduced by refining the cost-effectiveness of surgical instrumentation . Of note, such revision of the surgical instrumentation did not influence the intraoperative or postoperative outcomes, in part being counterbalanced by increasing surgical dexterity and confidence during the learning curve.Our experience confirms that RAKT achieved favorable intraoperative, peri-operative, and functional outcomes at both a short- and mid-term follow-up , as prevDespite being one of the few prospective experiences on RAKT worldwide, our study is not devoid of limitations. First, it is a preliminary experience with a relatively small sample size and only a mid-term follow-up. Second, we carefully selected the recipients for RAKT, especially at the beginning of the learning curve. In addition, while the current exclusion criteria are less stringent, DD-RAKT might have not been performed at our center due to logistical challenges, introducing a selection bias. Third, in light of the study design and of the limited sample size, we could not formally compare the outcomes of RAKT and the current gold standard (open KT) in both the LD and DD setting, nor evaluate the impact of the surgeon's learning curve on peri-operative and functional outcomes. Finally, in light of the demanding logistical challenges and financial costs of RAKT , the generalizability of our findings may be limited to selected referral centers.Acknowledging these limitations, our experience provides additional evidence to further explore the potential added value of robotics for KT in both the LD and DD settings.Further research is needed to: (a) evaluate whether RAKT provides incremental benefits for surgeons in both “standard” and “complex” cases, and whether these benefits translate into measurable improvements of the surgeons' learning curve and of patient outcomes; (b) assess the cost-effectiveness of RAKT from LDs and DDs using appropriate, standardized metrics; (c) to explore the impact of rewarming time on long-term functional outcomes; and (d) to refine the technique for RAKT by means of extended-reality platforms and/or immersive technologies such as 3D printing .riccardo.campi@unifi.it.The data analyzed in this study is subject to the following licenses/restrictions: Requests to access these datasets should be directed to The studies involving human participants were reviewed and approved by Comitato Etico Area Vasta Centro—AOU Careggi. The patients/participants provided their written informed consent to participate in this study.RC, FS, GV, and SS: study design. NM, PB, FC, AG, AB, AP, LG, NF, and MM: data collection. RC: statistical analysis. GV, RC, IG, and FS: manuscript writing. MG, AS, VL, and SG: critical revision of the manuscript. SS: supervision. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


The purpose of this systematic review was to investigate and summarize the evaluation methods of graft maturation on second-look arthroscopy following anterior cruciate ligament (ACL) reconstruction.A literature search was performed on articles before December 2017 to identify the literature that has evaluated graft maturation on second-look arthroscopy following ACL reconstruction. Only studies using human grafts, evaluating graft maturation with two or more gross findings were included. Study design, grafts, surgical techniques, follow-up period, evaluation parameters, and categories were compiled.Twenty-eight studies were included in this study. All studies evaluated graft maturation with two or more of the following three findings: graft integrity, tension, and synovial coverage. Two to four categories were used for evaluating each parameter, but the criteria for classification were slightly different for each study. Several studies reported neo-vascularization of grafts and the total maturation score by summing up the scores assigned to each evaluation parameter. Three studies reported that there was no correlation between second-look findings and patient-reported outcomes.Graft integrity, tension, and synovial coverage were the most frequently evaluated for graft maturation on second-look arthroscopy. However, there is no uniform criterion for evaluation. Therefore, development of a valid, uinform criterion is required.Level IV, systematic review of level I–IV investigations. After anterior cruciate ligament (ACL) reconstruction, the graft tendon undergoes a maturation process called “ligamentization”. Amiel et al.  describeMany studies have evaluated graft maturation using second-look arthroscopy –9. This However, there was no consensus on which parameters should be evaluated and which criteria should be used for each parameter to evaluate graft maturation on second-look arthroscopy. Surgeons have been using their own criteria for evaluating graft maturation. Toritsuka et al.  evaluateThe purpose of this study was to review the previous literature (1) to investigate how to evaluate graft maturation on second-look arthroscopy following ACL reconstruction and (2) to determine if second-look arthroscopy had an objective evaluation value. We hypothesized there would be a lot of different methods to perform second-look arthroscopy following ACL reconstruction and second-look findings would be less correlated to the clinical outcomes.We performed a systematic review of all the literature on second-look arthroscopy following ACL reconstruction, regardless of the level of study, graft type, and surgical technique . Searches were restricted to papers published in English. Only articles with detailed description on how to evaluate graft maturation were included. The mean follow-up period of all included studies was more than one year.Our literature search consisted of searches in PubMed and Embase for the terms “anterior cruciate ligament”, “reconstruction”, and “second-look” from the inception of these search engines till December 2017. After removing duplications, we checked the titles and abstracts of all articles to determine whether they fit the previously established inclusion criteria. The included articles identified by the search were each analyzed by a senior author to ensure their appropriateness.Only human ACL reconstruction studies using human grafts and evaluating graft maturation on second-look arthroscopy with two or more gross findings were included. As it was difficult to comprehensively evaluate graft maturation with only one parameter, we included only those studies that evaluated two or more gross findings of the grafts. In addition, studies with synthetic grafts, case reports, studies for which only abstracts had been published without full text, and systematic review studies were excluded. The results of this literature review are outlined in the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) diagram in Fig. The data from each of the 28 articles meeting the inclusion criteria for our systematic review were compiled. We collected demographic data, such as the graft type, number of second-look arthroscopies performed, and follow-up period from reconstruction to second-look arthroscopy. The reconstruction techniques  were recorded from all studies that reported them. We investigated the parameters used for evaluating second-look arthroscopy and categories that were used for each parameter to evaluate graft maturation in those studies.To assess the methodological quality, the modified Coleman Methodology Scores (mCMS) and subscales were determined for each included study . The finAn overview of the 28 studies showing year and journal of publication, level of evidence, graft type, number of second-look arthroscopies performed, reconstruction technique used, and follow-up period from ACL reconstruction to second-look arthroscopy is provided in Table The literature review described above yielded 28 manuscripts that met all inclusion criteria  evaluated the integrity of the graft on second-look arthroscopy. The most common classification of graft integrity involved three categories according to the severity of graft tear: intact, partial  tear, and complete  tear , 25, 31. evaluateTwenty-two of the total studies evaluated the tension of the graft by probing during second-look arthroscopy. In addition, most studies objectively evaluated the graft tension using physical examination, Telos stress radiography, and KT-arthrometer. The most common classification of graft tension also involved three categories depending on how the graft moved during probing: taut, slightly (mildly) lax, and lax. Nine studies used these categories –29, 34. Most studies 26/28) used synovial coverage to evaluate graft maturation in second-look arthroscopy. The most common classification of synovial coverage involved three categories. However, the criteria for assigning the categories vary by study. Kondo et al.  classifi used synOnly three of the included studies used neo-vascularization for evaluating graft maturation , 11, 18.Seven studies tried to evaluate graft maturation comprehensively by combining various parameters. They calculated the total graft maturation score by summing up the scores assigned to each evaluation parameter. Details of each study are summarized in Table Nine of the included studies compared second-look findings with clinical outcomes Table . Most stOn the other hand, only three studies investigated the correlation between second-look findings and patient-reported outcomes  , 22, 32.This systematic review summarizes how graft maturation was evaluated on second-look arthroscopy following ACL reconstruction. All included studies evaluated graft maturation with two or more of the following three gross findings: graft integrity, tension, and synovial coverage. Two to four categories were used for evaluating each parameter, but the criteria for classification were slightly different for each study. Several studies also used the parameter of neo-vascularization to evaluate the viability of grafts.Follow-up period from ACL reconstruction to second-look arthroscopy varied according to the study, but the average length was about 1–2 years. Second-look arthroscopy to evaluate a graft should be performed only after the maturation of the graft ends. However, previous studies have reported different results on the timing of graft remodeling. Falconiero et al.  reportedGraft integrity is the most basic evaluation method and almost all studies used integrity for evaluating graft maturation after ACL reconstruction. This was expressed as the presence and extent of the graft tear. In the majority of studies, graft integrity was classified into three categories: intact, partial  tear, and complete  tear. However, intact graft with no graft tears were mostly categorized as normal, whereas Ohsawa et al. classified normal integrity as over 80% , 29 and Tension of the graft is one of the most important evaluation parameters because it assesses the stability of the knee joint, which is the main purpose of ACL reconstruction. The tension of grafts was evaluated as taut, slightly lax, and lax by probing during second-look arthroscopy. This classification is problematic in that it depends on the subjective judgment of the examiner. Some studies described the use of rulers at the time of probing or the use of anatomical indicators such as femoral condyle to measure the tension of grafts, but this may also not be objective. In addition, the measurement of stability by arthroscopic probing has the disadvantage that it is difficult to evaluate the stability of rotation. Therefore, a method is needed that can be used to evaluate stability of rotation more objectively.While the integrity and tension of grafts represent mechanical properties, synovial coverage has been used as a measure of biologic maturation. Synovial coverage of the graft was also mostly categorized into three categories, while the criteria for classification varied by study. Some studies regarded the best category as complete coverage , 28, 30,Recently, long term follow-up studies of ACL reconstruction have been published, and long-term survival of grafts is recognized as important. Bottoni et al.  reportedSeveral studies calculated the total maturation score by summing up the scores assigned to each evaluation parameter , 17, 18.Although a few studies reported that graft tension in second-look arthroscopy was significantly correlated with objective stability test results , 22, secThis study has several limitations. First, only five randomized trials, among 28 studies, were included for this systematic review. The quality of included studies was assessed to be relatively low (mean mCMS of 46.4 ± 6.7). Second, this study summarizes previous methods for evaluating graft maturation on second-look arthroscopy following ACL reconstruction, but it does not answer which evaluation method is appropriate. Consensus and validation are needed to determine which method is the most appropriate evaluation method. Third, the follow-up periods of the studies were relatively short and no long-term follow-up studies were included, and most studies had difficulty identifying the relationship between clinical outcomes and the evaluation of graft maturation. Finally, almost all the studies were from northeast Asia, including South Korea, Japan, and China; therefore, most studies are limited to specific races.Graft integrity, tension, and synovial coverage were the most frequently evaluated criteria for graft maturation on second-look arthroscopy following ACL reconstruction. However, there is no uniform criterion for evaluation of each parameter. Therefore, development of a validated evaluation method for second-look arthroscopy is required.


The decision was made by bearing in mind the substantial contribution made by this profession to the population’s health, and that this year marks the bicentennial of the birth of Florence Nightingale, one of the founders of modern nursing. states that, although nurses are half the health workers globally, it is necessary to add nine million of these professionals before 2030. The aforementioned seeks to reach the universal coverage in health, which is an indispensable goal to achieve the planet’s sustainable development objectives.(The 2020 initiative: International Year of Nursing is supported by the WHO, the International Confederation of Midwives, the International Nurses Council, the campaign Nursing Now, and the United Nations Population Fund. Throughout the year and globally activities will be undertaken to celebrate the work of nursing and which show to public opinion the difficult conditions these professionals encounter, situations that must be improved by promoting investment to increase training and employment of this personnel as part of each country’s commitment with health for all.(Investigación y Educación en Enfermería, has exalted the essential role of nurses in caring for individuals and communities, and will continue supporting efforts on the dissemination of knowledge to collaborate in the growth of this discipline, which, although young, presents a wealth of research as never seen before.(For 38 years, our journal, Happy International Year of Nursing