Provided there is a suitable positioning of dyes, target-bound FIT2 probes emit brighter than mono dye probes, while dye–dye contact in the single stranded state provides opportunities for decreasing background fluorescence. The probes were used to explore the single nucleotide-specific detection of a C → U edited RNA of the glycine receptor (GlyR). We observed strong self-quenching upon single base mismatched hybridization of FIT2 probes, which helped in distinguishing edited from unedited RNA target in cell lysates.Efficient fluorogenic hybridization probes combine high brightness and specificity of fluorescence signaling with large turn-on of fluorescence. Herein, we present an approach to enhance signaling by combining two identical fluorescence base surrogates in FIT Efficient fluorogenic hybridization probes combine high brightness and specificity of fluorescence signaling with large turn-on of fluorescence. An ideal probe would excel in all three performance characteristics. However, in practice, usually only one or two of these criteria can be optimized with a given probe technology. For example, with molecular beacon-type probes4 relying on the distance-dependent interactions between two dyes it is relatively easy to optimize the turn on by spectral matching of fluorescence donors and acceptors, often at the cost of brightness, or by optimizing aggregation of dyes.5–11 Sequence specificity of fluorescence signalling, on the other hand, typically is limited by the fidelity of Watson–Crick base pairing. Considering that oligonucleotide probes must have a sufficient length to provide uniqueness and sufficient affinity, it can be difficult to discriminate targets that differ, for example, by a single nucleotide. This issue is most pressing when single nucleotide-specific measurements ought to be performed at the comparatively low temperatures required for measurements in cells or cell lysates.Fluorogenic oligonucleotide hybridization probes are extremely versatile and enable the direct detection and localization of specific DNA/RNA molecules within complex mixtures such as in cells, cell lysates or PCR-type applications.12–17 In these PNA-, DNA- or RNA-type oligomers, a fluorophore belonging to the thiazole orange family of cyanine dyes replaces a canonical nucleobase, distinguishing them from Light-Up probes18 and Echo probes,6 which tether thiazole orange via flexible tethers at the periphery. The fluorescent base surrogate senses viscosity changes in the immediate environment. In a low viscosity environment TO is virtually non-fluorescent because rotations around the central methine bridge deplete the TO excited state.19 Formation of the probe-target complex places the “TO base” in the double helical base stack. This high viscosity environment renders the TO fluorescent. Adjacent single base mismatches decrease the local viscosity, causing decreases of TO emission.14,20 As a result, FIT probes provide for single nucleotide specific fluorescent signalling regardless of hybridization fidelity. This property has facilitated the real-time detection of single nucleotide alterations in in vitro assays21,22 and fluorescence microscopic imaging.23–30 The FIT probe concept has also been applied for detection of double strands via triplex formation,31–33 and in signalling aptamers.34 To improve turn-on and brightness of fluorescence signalling by FIT-probes, we employed the artificial “thiazole orange base” as energy donor in combination with a second dye serving as energy acceptor.35 However, this affects the single nucleotide specificity of fluorescence signalling, because the “TO base” will transfer energy to the acceptor dye regardless of its sequence environment (matched or mismatched). To enable detection of C → U edited RNA, the approach was applied to a binary probe system, in which target specificity was determined by hybridization fidelity.36Focussing on high sequence-specificity of fluorescence signalling, we have introduced the Forced Intercalation (FIT) probes .12–17 In2 probe such decreases could occur whenLooking for a way to improve hybridization-induced fluorescence enhancements and brightness of fluorescence signalling by FIT probes while maintaining or even increasing target specificity, we considered the combined use of two identical fluorescent base surrogates . The int(1) None of the TO-like dyes is embedded in the high viscosity environment provided by matches of adjacent base pairs.(2) The two dyes form weakly fluorescing H-aggregates.via homo FRET or(3) One TO-like dye in a low viscosity environment quenches the other dye (4) Both dyes engage in excitonic interactions.2 probes could potentially show lower fluorescence than expected for the sum of the fluorescent components. Hybridization will prevent options (1) and (2) making it likely that fluorescence should increase strongly upon binding of target. Highest fluorescence enhancements should be obtainable when options (3) and (4) remain unutilized in the probe-target duplex. This should be the case when the distance between the dyes is high and/or orientation of the dyes does not permit coupling of transition dipole moments. For example, according to molecular ruler work from Asanuma37 and our work,35 a 8 nt distance minimizes energy transfer between dyes embedded in the base stack of a DNA–DNA duplex. Given an optimal positioning, FIT2 probes could therefore provide higher fluorescence intensity and higher turn-on upon hybridization than single dye FIT probes. We furthermore hypothesized that a single base mismatch in the vicinity of one dye will weaken the stacking interactions and allow the dye to adopt conformations that permit energy transfer. As a result, fluorescence signalling by FIT2 probes may discriminate single base mismatches under non-stringent hybridization conditions.Options (2)–(4) cannot exist in single dye FIT probes explaining why single stranded FIT2 probes that enable the single nucleotide specific call for a nucleotide corresponding to the C → U editing site in RNA from a transcript encoding a glycine receptor (GlyR). Measurements in highly viscous cell lysates demonstrate that two identical TO-type base surrogates within a FIT2 probe can indeed discriminate better against single base mismatches than mono FIT probes.Herein we describe FIT2 probes to enable homogeneous detection of GlyR mRNA  as QB next neighbour, as previously described.25We designed FIT probes and FITlyR mRNA .38 Probe2 FIT probes targeting the unedited state of GlyR mRNA  increase the fluorescence turn-on by introducing additional quench options for single or molten strands, (ii) enhance the intensity of fluorescence in the double strand while (iii) providing single mismatch discrimination.2 probes with 2′-OMe modifications, which improve resistance to nucleases and, therefore, facilitate measurements in cells and cell lysates. According to previously defined design criteria, the 2′-OMe RNA probes included a locked nucleic acid (LNA) building block as a 3′ neighbour of each Ser(QB) or Ser(TO) nucleotide and 2′-deoxynucleotides on each side of the serinol-LNA dinucleotides. This design confers high brightness to probe-target duplexes while keeping fluorescence of single strands low. Both QB and TO dyes were introduced at positions used for UE-QB2-3. In addition, we evaluated probes specific for the edited state of GlyR mRNA.Encouraged by the findings, we constructed FIT2 probes  were compared with the corresponding mono labelled FIT probes (UE-Dye-OMe-C1 and ED-Dye-OMe-C1). Fluorescence measurements performed at 25 °C  than the mono probe UE-QB-OMe-C1 (F/F0 = 23 and 85). In addition, the single nucleotide specificity was higher  and the dual dye probe fluoresced with higher intensity.The dual labelled FIT5 °C see  and 37 °2-OMe-1) was higher than for the mono dye probe (UE-TO-OMe-C1). A similar picture emerged for probes targeting the edited GlyR mRNA state. In this sequence context, introduction of the second QB can, again, improve the responsiveness, specificity and brightness of signalling, whereas the positive effects of the second TO were limited to brightness.The TO dye was placed in the same context and again single nucleotide specificity and brightness of fluorescence provided by dual TO probe  and two additional probes (UE-QB2-OMe-4 and UE-TO2-OMe-2) used for detection of target in cell lysate (vide infra). Single nucleotide mismatched duplexes were characterized by efficient self-quenching occurring with 53–85% efficiency meaning that emission intensities of the FIT2 probes corresponded to only 15–47% of the intensity expected for the sum of intensities from the two mono dye probes. FIT2 probes engaged in matched hybridization experienced only 3–53% self-quenching.To expose the hybridization state dependency of dye–dye interactions in FIThed form . This in Table S1. In this45 Regardless of the mechanism involved, self-quenching was, in each case, more efficient in mismatched duplexes than in the corresponding matched duplexes. The single-stranded probes showed a less uniform behaviour with self-quenching ranging between 67% for UE-QB2-OMe-4 to −40% for ED-TO2-OMe-1. Single strands can adopt conformations that enable dye–dye contact. In cases such as in ED-TO2-OMe-1 such a contact may decrease the degrees of freedom for rotations around the dye's methine bridge explaining why the single-strand can fluoresce with higher intensity (leading to “negative” self-quenching) than expected for the sum of the component intensities. While in most other cases turn-on with FIT2 probes is higher than with mono dye probes due to additional quench options in the single strand, the data indicates sequence dependencies.Self-quenching is, amongst other effects, the result of excitation energy homotransfer. This transfer depends on the orientation of transition dipole moments. In single nucleotide mismatched duplexes one of the dyes gains flexibility which facilitates arrangements allowing for efficient transfer of excitation energy. According to an alternative interpretation, the high quenching efficiencies may be a consequence of delocalized excited states emerging from dye–dye interactions. As reported for multifluorophore-DNA such states may be particularly vulnerable to quenching.2 probes in a challenging matrix; 100% lysate of HEK293 cells which contains the multitude of a cell's molecules including RNA. Competing probes, one for detection of the edited state of GlyR mRNA via QB signalling and the other for detection of the unedited state via TO signalling, were equipped either with QB or with TO nucleotides. Out of four tested ED-QB2-OMe probes (Table S52-OMe-1 as the brightest probe in this series. The selection of UE-TO2-OMe-2 (out of 4 tested probes (Table S4TM matching (TM of mismatched TO probe duplexes should be lower than the TM of matched QB probe duplexes) and single nucleotide specificity. The ED-QB2-OMe-1/UE-TO2-OMe-2 probe pair was compared with the corresponding mono dye probes, ED-QB-OMe-C1/UE-TO-OMe-C1s. Both probe sets were pipetted to lysates and RNA target was added. In addition, measurements were performed in PBS buffer without lysate. In buffer, the FIT2 probe system provided a 34-fold enhancement of QB signalling upon addition of edited target compared to a 13-fold by the mono dye pair  as a codye pair , left. Lited RNA , right. 2-OMe-1 has the QB nucleotides in positions used also in ED-QB2-OMe-1. TM matching and high sequence specificity recommended the use of ED-TO2-OMe-4 . This and self-quenching in the single stranded state probably contributes to the high specificity observed when differently coloured FIT2 probes were used as pairs for detection of RNA in a complex matrix such as cell lysate. Our experiments showed that responsiveness and single nucleotide specificity are affected when FIT probes are used in cell lysate. In this regard, the FIT2 approach is an easy-to-implement method to improve specificity as well as brightness and responsiveness of fluorescence.In our pursuit of approaches providing RNA FIT hybridization probes with enhanced brightness, responsiveness and single nucleotide specificity of fluorescence signalling, we have explored FITThe datasets supporting this article have been uploaded as part of the ESI.†S. S. and A. H. synthesized oligonucleotide probes. S. S. and A. K. evaluated properties of oligonucleotide probes. S. S. and O. S. analyzed the data. S. S. wrote the first draft of the manuscript. O. S. conceived the research and wrote the final version of the manuscript.There are no conflicts to declare.SC-014-D3SC00363A-s001


Dear Editor,10) or over 20 min (FC10), with the purpose of assess whether the fluid responsiveness [defined as an increase in the stroke volume index (SVI) ≥ 10%] was affected by the time of infusion . The grey zones for all the statistically significant ROC curves were also calculated considering the low cut-off value including 90% of negative FC responses and a high cut-off value predicting positive FC in 90% of cases . On the contrary, the ROC curve of the changes in SAP after ½ FC20 was not statistically significant (p = 0.11). Our results show a positive moderate linear correlation (r2 = 0.50) between for ΔSAP and ΔSVI FC10. The associated ROC curve constructed showed that ΔSAP > 10% is highly suggestive of FC response , whereas ΔSAP = 0 is highly suggestive of no response . On the contrary, the linear correlation of FC20 was weak (r2 = 0.28), and the ROC curve was insignificant.There was a significant positive correlation between ΔSAP and ΔSVI from baseline to ½ FC administration, during both FC10, still maintains clinical utility, suggesting that no increase after ½ FC is associated with no fluid responsiveness, whereas an increase of at least 10% is associated to fluid responsiveness. In contexts of low resources or when a SVI monitoring is not available, our results may provide practical cut-offs to guide fluid optimization in elective surgical patients. This finding, however, could be partially dependent on the intrinsic mathematical coupling between pressure and flow variables changes after the fluid infusion, since the MostCare® system is based on the high sample rate analysis of the arterial waveform.The interplay between SAP and SVI is based on a complex balance between cardiac factors, arterial load, and resistance , 4. In fRespectfully Yours.


This retrospective study aimed to investigate factors associated with inhibition of early aneurysm obliteration after flow diverter (FD) treatment. We also created the early obliteration inhibition (EOI) score for pre-operative evaluation.We examined 110 cerebral aneurysms in 104 patients who underwent FD treatment. The following parameters were investigated: age, sex, symptoms, aneurysm location and type, maximum aneurysm diameter, parent vessel diameter, neck diameter, and dome–neck ratio. We also noted aneurysm location relative to the curvature of the parent artery and any branches arising from the aneurysm dome. Procedural factors such as FD diameter and length, number of FDs placed, type of FD, and use of adjunctive coiling were also investigated. Aneurysm obliteration was evaluated using digital subtraction angiography 3 months after the procedure. Adequate obliteration was defined as grade C or D on the O’Kelly–Marotta scale.The following factors inhibited early obliteration: 1) extradural location, 2) saccular aneurysm, 3) aneurysm neck located at the outer convexity of the parent artery, and 4) arterial branch arising from the aneurysm dome. Odds ratios were used to create an EOI score. Receiver operating characteristic curve analysis showed that the optimal cut-off EOI score for adequate obliteration was 1.5 .The EOI score, which is based on factors that inhibit early obliteration, may predict early treatment outcomes of FD placement. Briefly, the FD is placed to cover the aneurysm neck, which promotes thrombosis within the aneurysm and provides a scaffold for endothelial cell growth along the device.2 The Pipeline embolization device  was the first FD approved for use. The Pipeline Flex (Medtronic) is an improved version that was introduced in Japan in 2015 for use in wide-necked proximal internal carotid artery aneurysms with a major diameter of at least 10 mm.3 In 2020, the Pipeline Shield (Medtronic), which has improved operability and antithrombotic properties, was introduced for use in internal carotid and vertebral artery aneurysms with a major diameter of at least 5 mm.6 Another new FD, the Flow Redirection Endoluminal Device , is now indicated for use in aneurysms of the proximal anterior and middle cerebral arteries and basilar artery.9 Therefore, a considerable number of cerebral aneurysms can now be treated using a FD.In recent years, remarkable progress has been made in endovascular treatment of cerebral aneurysms. More than half of unruptured cerebral aneurysms treated in Japan are treated using an endovascular approach. Although conventional endovascular treatment involves endosaccular embolization using detachable coils, the introduction of flow diverter (FD) placement for side-wall type unruptured aneurysms has caused a paradigm shift in treatment.11 Early obliteration prevents aneurysmal rupture and enlargement and enables dose reduction and termination of antiplatelet therapy.12 This study aimed to elucidate factors that inhibit early obliteration at the three-month stage after FD treatment and establish an early obliteration inhibition (EOI) score to identify patients and aneurysms less suitable for FD treatment.The reported rates of complete aneurysm obliteration 1 year after FD treatment range from 70% to 80%.Patients with unruptured cerebral aneurysms who successfully underwent FD placement and subsequent imaging follow-up three months after treatment for five years since October 2016, which was when FD was introduced at our hospital, were eligible for study inclusion. We excluded patients under age 18 years and those who had undergone previous coil embolization or FD placement. Patients who required additional treatment within 3 months of FD placement were also excluded. Clinical and radiological data were obtained from a retrospective review of the medical records and neuroimaging studies. All patients provided written informed consent for treatment. The study received institutional review board approval. The requirement for informed consent for study participation was waived owing to the retrospective nature of the study and because all data were de-identified.12 In patients with >550 aspirin reaction units (ARUs), aspirin dosage was increased to 200 mg/day. In patients with ≥210 P2Y12 reaction units (PRUs), clopidogrel was stopped and a loading dose of prasugrel 20 mg was administered followed by 3.75 mg/day. In patients with 60–210 PRUs, clopidogrel 75 mg/day or prasugrel 3.75 mg/day was administered. In patients with <60 PRUs, the prasugrel dosage was reduced to 1.88 mg/day. Repeat platelet function testing was performed again before hospital discharge; dosage changes were made accordingly.12Dual antiplatelet therapy (aspirin 100 mg/day and clopidogrel 75 mg/day) was initiated in all patients 2 weeks before treatment. Platelet function was evaluated using VerifyNow  2 days before treatment.FDs were placed with the patient under general anesthesia. An 8Fr sheath introducer was placed into the superficial femoral artery, followed by administration of 70–100 U/kg of unfractionated heparin. Heparin was titrated to maintain activated clotting time (ACT) >250 seconds. ACT was measured hourly.8 The parent artery diameter was measured at the distal, neck, and proximal positions of the aneurysm. FD size was determined based on the maximum diameter of the parent artery and the length of the aneurysm neck. The FD was guided across the neck of the aneurysm and deployed. Percutaneous transluminal angioplasty was performed if high-resolution cone-beam computed tomography depicted insufficient adaptation of the FD against the parent arterial wall. Endovascular coiling was performed in intradural aneurysms with major diameter ≥10 mm if a bleb or jet blood flow into the aneurysm was observed. Brain magnetic resonance imaging was performed the day after treatment. Patients were discharged home approximately 7 days after the procedure.An 8Fr guiding catheter  was advanced into the parent vessel of the target aneurysm. A 5Fr Navien 115 cm (Medtronic) or 5Fr Sofia Select (MicroVention-Terumo) catheter was used as an intermediate catheter. The Marksman (Medtronic) or Phenom 27 (Medtronic) microcatheters were used for placement of the Pipeline Flex or Pipeline Shield. The Headway Plus 27 (MicroVention-Terumo) microcatheter was used to place the FRED. FDs were guided to the aneurysm through the intermediate catheter. FDs were selected at the operator’s discretion according to the course of the parent artery.Digital subtraction angiography (DSA) was performed 3, 6, and 12 months after the procedure. Dual antiplatelet therapy was continued for at least 6 months. and the DSA results were used to reduce the number of drugs to one at the 6–12-month mark and discontinue therapy at the 18–24-month mark. Since the purpose of this study was to evaluate patients after 3 months of treatment, all included patients were on DSA and on dual antiplatelet drugs at the time of evaluation.13: A, at least 95% of the aneurysm remains; B, 5%–95% remains; C, <5% remains; and D, complete obliteration. Adequate obliteration was defined as OKM grade C or D.The following patient and aneurysm characteristics were recorded: age, sex, symptoms, aneurysm location and type, maximum aneurysm diameter, parent vessel diameter, neck diameter, and dome–neck ratio. We also noted aneurysm location relative to the curvature of the parent artery and any branches arising from the aneurysm dome. Procedural factors such as FD diameter and length, number of FDs used, type of FD, and use of adjunctive coiling were also recorded. Aneurysm obliteration was evaluated using the O’Kelly–Marotta (OKM) grading scalea priori based on previous studies16 because they are likely to be associated with aneurysm obliteration:Patient factor: ageAneurysm factors: intra/extradural location, anterior/posterior circulation location, maximum aneurysm diameter, parent artery diameter, dome–neck ratio, saccular/dissecting aneurysm, aneurysm location relative to the curvature of the parent artery (yes/no) and arterial branch arising from the aneurysm dome (yes/no)Procedural factors: number of FDs, type of FD, and adjunctive coilingContinuous data are presented as medians with interquartile range (IQR). Categorical data are presented as numbers with percentage. A multivariable ordinal logistic regression model was generated to explore factors associated with adequate obliteration 3 months after the procedure. The following factors were selected Ordinal logistic regression analysis was performed to calculate odds ratios (ORs) with 95% confidence intervals (CIs) and determine factors associated with early adequate obliteration. Penalized maximum likelihood estimation was used to allow shrinkage of the effect of factors to correct for possible overfitting. Factors associated with inhibition of early obliteration were used to create a binary logistic regression predictive model. A simple score was developed to predict the EOI score based on the regression coefficients of the binary logistic regression model. Receiver operating characteristic (ROC) curves and optimal cut-off points were calculated. Two-sided P<0.05 was considered significant. Statistical analyses were performed using R software version 4.1.1 .This study included 110 aneurysms in 104 patients. Among the 134 patients treated during the study period, 30 were excluded based on criteria: one patient in whom FD placement was not successful, two under age 18 years, 23 who underwent FD after a previous procedure, three who were lost to follow-up, and one who underwent additional treatment within 3 months of the FD procedure. Eleven patients (10.5%) experienced an adverse event, including cerebral infarction with residual sequelae (n=3) and carotid–cavernous sinus fistula without sequelae (n=4). All four carotid–cavernous fistulae required additional treatment. In one patient, FRED placement was not successful and a Pipeline Shield had to be placed instead. No deaths occurred.Patient and aneurysm characteristics are shown in In ordinal logistic regression analysis, the following factors were significantly associated with inhibition of early obliteration: extradural location , saccular aneurysm , aneurysm neck located at the outer convexity of the parent artery , and arterial branch arising from aneurysm dome . Binary logistic regression performed using these four factors showed that extradural location , saccular aneurysm , and aneurysm neck located at the outer convexity of the parent artery  were significantly associated with inhibition of early obliteration. Although presence of an arterial branch arising from the aneurysm dome failed to reach statistical significance by a small margin , we still included it in the EOI score because of strong sign of association in ordinal logistic regression analysis .The EOI score was created using the regression coefficients from the binary logistic regression model. Scores were weighted by rounding coefficient values: 2 for extradural location and aneurysm neck located at the outer convexity of the parent artery, and 1 for saccular aneurysm and arterial branch arising from the aneurysm dome . Figure A 64-year-old woman presented with a right oculomotor palsy caused by a right paraclinoid internal carotid artery aneurysm. Maximum aneurysm diameter was 18.5 mm and the neck measured 5.5 mm. A Pipeline Shield  was placed as described above. Antiplatelet therapy was administered as described above. DSA 3 months after the procedure showed complete aneurysm obliteration. In this case, the preoperative EOI score was 1 . We therefore expected early obliteration with FD treatment, which was demonstrated on follow-up DSA .A 60-year-old man presented with an incidental asymptomatic right vertebral artery aneurysm that exhibited arterial dissection and partial thrombosis of the dome. A FRED  was placed without complication as described above. Antiplatelet therapy was administered as described above. DSA 3 months after the procedure showed residual aneurysm (OKM grade B). In this case, the preoperative EOI score was 3  and early obliteration was not expected .3 FDs currently available for use include the Pipeline Flex, Pipeline Shield, and FRED. Many cerebral aneurysms are amenable to FD treatment.Since endovascular coiling of cerebral aneurysms was first introduced in 1997, use of endovascular treatment techniques has increased. FDs, which were initially developed to treat sidewall-type unruptured aneurysms occurring on the lateral wall of the parent artery, have been found safe and effective.11 The 1- and 5-year rates of complete obliteration in a large single-center study of the Pipeline embolization device in Argentina were 75.8% and 96.4%, respectively.1 In the Prospective Study on Embolization of Intracranial Aneurysms with the Pipeline device (PREMIER study), the complete obliteration rate at 1 year was 84%.6 Two other studies have reported 1-year obliteration rates of 77.2% and 81.8%, respectively.5 Regarding the FRED, reported 1-year complete obliteration rates range from 62% to 73%.9Numerous large-scale studies have investigated outcomes of FD treatment. The Pipeline for Uncoilable or Failed Aneurysms trial  evaluated the Pipeline embolization device and reported 6- and 12-month obliteration rates of 74% and 87%, respectively.12 Therefore, the ability to predict early obliteration can contribute to the pharmacotherapeutic strategy. In our study, adequate obliteration (OKM grade C or D) was achieved 3 months after treatment in 71.8% of aneurysms, which is comparable to previously reported 1-year rates. We believe our study is the first to report early obliteration status at the 3-month timepoint.These results show that the aneurysm obliteration rate increases over time; nonetheless, not all aneurysms progress to complete obliteration. In these aneurysms, retreatment is warranted. The ability to predict the need for retreatment at an early stage would be clinically beneficial. Furthermore, dual antiplatelet therapy is required after FD placement; however, early discontinuation may be required in certain patients because of comorbidities.The following four factors inhibited early aneurysm obliteration in our study: 1) extradural location, 2) saccular aneurysm (versus dissecting aneurysm), 3) location of the aneurysm neck on the outer convexity of the parent artery, and 4) presence of an arterial branch arising from the aneurysm dome. Subsequently, ORs were used to create an EOI score. In ROC analysis, the optimal EOI score cut-off value for adequate obliteration was 1.5. In other words, an EOI score <1 indicates that early obliteration is likely while a score ≥2 suggests it is not. Our findings may enable clinicians to predict treatment course and perioperative management. Furthermore, they should help determine whether FD treatment is even indicated.Our study has several limitations. It was retrospective in design and was conducted in a single center. Therefore, the sample size was small and selection bias may have been introduced. In addition, we excluded patients younger than age 18 years, those with previously treated aneurysms, and patients who required retreatment after FD placement.We clarified factors that inhibit early obliteration of cerebral aneurysms after FD treatment. The EOI score, which is determined based on these factors, has the potential to predict early obliteration. Future prospective studies are warranted to verify our findings.


Volume resuscitation is a cornerstone of modern anesthesia care. Finding the right balance to avoid inadequate or excess volume administration is often difficult to clinically discern and can lead to negative consequences. Pulse pressure variation is often intraoperatively used to guide volume resuscitation; however, this requires an invasive arterial line and is generally only applicable to patients who are mechanically ventilated. Unfortunately, without a pulmonary artery catheter or another costly noninvasive device, performing serial measurements of cardiac output is challenging, time-consuming, and often impractical. Furthermore, noninvasive measures such as LVOT VTI require significant technical expertise as well as access to the chest, which may not be practical during and after surgery. Other noninvasive techniques such as bioreactance and esophageal Doppler require the use of costly single-use sensors. Here, we present a case report on the use of corrected carotid flow time (ccFT) from a portable, handheld ultrasound device as a practical, noninvasive, and technically straightforward method to assess fluid responsiveness in the perioperative period, as well as the inpatient and outpatient settings. Volume resuscitation is a cornerstone of modern anesthesia care. Finding the right balance to avoid inadequate or excess volume administration is often difficult to clinically discern and can lead to negative consequences. These consequences range from acute kidney injuries, inadequate cellular perfusion, to pulmonary edema or even frank anasarca leading to poor wound healing and increased hospital length of stay ,2. ThereccFT has been validated as a technique for assessing cardiac output , where cOur patient was a 14-year-old girl who was otherwise healthy presenting to the operating room for open reduction and internal fixation of an ankle fracture. She had been NPO for 18 h prior to the start of the surgery. She was 160 cm tall and weighed 65 kg. Her initial heart rate was 114 beats/minute, and her initial blood pressure was 114/65 mmHg. General anesthesia was induced with 200 mg of propofol and 50 mcg of fentanyl, a laryngeal mask airway was inserted, and she was maintained on a concentration of 3.2% inspired sevoflurane throughout the procedure. Carotid flow time was measured 45 min following the induction of anesthesia by placing a handheld ultrasound probe, Butterfly IQ , on the patient’s neck  and locaThis is the first presentation of the use of a portable, handheld ultrasound device to measure changes in ccFT as a practical, noninvasive, and technically straightforward method to assess fluid responsiveness. Although we demonstrate the calculation of ccFT before and after a fluid bolus, the ideal test for fluid responsiveness would employ maneuvers such as a passive leg raise or, in patients receiving controlled mechanical ventilation, an end-expiratory occlusion test, which do not require fluid administration before determining its need. Portable, handheld ultrasounds may be useful for assessing fluid responsiveness in patients such as those undergoing surgery where it may be impractical to have invasive monitoring  and other noninvasive measurements (LVOT VTI) due to lack of access to the chest. Furthermore, it is cost-effective as the portable, handheld ultrasound devices can be easily cleaned and immediately reused on another patient without requiring costly, single-use sensors . We believe that many of our patients would benefit from portable, handheld ultrasound assessment of ccFT to guide intraoperative fluid administration given its ease of use and the increasing availability and affordability of portable handheld ultrasound devices. Further studies across different patient populations are necessary to validate this technique prior to widespread use.


Correction to: Journal of Cancer Research and Clinical Oncology 10.1007/s00432-022-04418-yIn Fig. 5 of this article, there were two errors in Fig. 5c, right panel. Firstly, the title of the graph (in the right panel) that had read “Target tumour” should have read “Off-target tumour”. In addition, the graph had presented the raw tumour volume data, and it should have presented the % tumour volume increase data. Asterisks that depict the level of significance had also been missing;The Fig.


Head and Neck cancers (HNC) are a heterogeneous group of upper aero-digestive tract cancer and account for 931,922 new cases and 467,125 deaths worldwide. About 90% of these cancers are of squamous cell origin (HNSCC). HNSCC is associated with excessive tobacco and alcohol consumption and infection with oncogenic viruses. Genotyping tumour tissue to guide clinical decision-making is becoming common practice in modern oncology, but in the management of patients with HNSCC, cytopathology or histopathology of tumour tissue remains the mainstream for diagnosis and treatment planning. Due to tumour heterogeneity and the lack of access to tumour due to its anatomical location, alternative methods to evaluate tumour activities are urgently needed. Liquid biopsy approaches can overcome issues such as tumour heterogeneity, which is associated with the analysis of small tissue biopsy. In addition, liquid biopsy offers repeat biopsy sampling, even for patients with tumours with access limitations. Liquid biopsy refers to biomarkers found in body fluids, traditionally blood, that can be sampled to provide clinically valuable information on both the patient and their underlying malignancy. To date, the majority of liquid biopsy research has focused on blood-based biomarkers, such as circulating tumour DNA (ctDNA), circulating tumour cells (CTCs), and circulating microRNA. In this review, we will focus on ctDNA as a biomarker in HNSCC because of its robustness, its presence in many body fluids, adaptability to existing clinical laboratory-based technology platforms, and ease of collection and transportation. We will discuss mechanisms of ctDNA release into circulation, technological advances in the analysis of ctDNA, ctDNA as a biomarker in HNSCC management, and some of the challenges associated with translating ctDNA into clinical and future perspectives. ctDNA provides a minimally invasive method for HNSCC prognosis and disease surveillance and will pave the way in the future for personalized medicine, thereby significantly improving outcomes and reducing healthcare costs. Head and neck cancers (HNCs) are the 7th most common cancer in the world, with 931,922 new cases and 467,125 deaths in 2020 . The Org®) are at risk of developing HPV associated oropharyngeal squamous cell carcinoma (OPSCC) [The International Agency for Research on Cancer  has identified diverse risk factors that contribute to the development of HNSCC. Excessive consumption of alcohol and tobacco use are the two major risk factors for the development of HNSCC. It is estimated that at least 75% of HNSCCs are caused by tobacco smoking and alcohol consumption , 11, 56. (OPSCC) .Current diagnostic methods for HNSCC include physical examination, endoscopy, imaging studies, biopsy and tumour biomarker testing . Tissue ®) and Nivolumab (Opdivo®), they are recommended in the National Comprehensive Cancer Network (NCCN) guideline.Currently available treatments for patients with HNSCC include surgery, radiotherapy, chemotherapy, targeted therapy and immunotherapy . TreatmeIn recent decades, new cases of OPSCC are increasing globally due to increasing rates of HPV infections. OPSCC has now surpassed cervical cancer to become the number one cause of HPV-related cancer in the United States and in Australia , 155. ThBefore biomarkers can be integrated into a clinical workflow, they will have to undergo five phases of development  to provPI3KCA mutations are commonly found in HNSCCs [PI3KCA mutations would not benefit PI3K pathway inhibitor. Given the drawback of using tumour tissue to diagnose and predict treatment response in HNSCC patients, alternative methods are urgently needed to better manage patients with HNSCC [The use of precision, targeted genomic therapies in HNSCC lagged behind many other cancer types, leading to poor survival outcomes. As an example, oncogenic n HNSCCs , and patn HNSCCs . Howeverth HNSCC . Liquid Mandel and Metais in 1948 detected for the first time cell-free DNA (cfDNA), which is now referred to as short fragments of nuclear acids in circulation . cfDNA i, revealed that M2-like tumour-associate macrophages (TAMs) can regulate CTCs metastasis by breaking down the basement membrane, promoting angiogenesis and protecting tumour cells from anti-tumour immunization [Circulating tumour (ctDNA) is derived from tumour and is part of the total cfDNA pool, representing only small fraction of cfDNA. It ranges from 0.01% to 90% , but is nization . TAMs canization . The amonization . Muhannanization .Fig. 1ct, DriverDBv3 (N = 2798) [. Also, Dietlein et al.,’s publication in Nat Genet (N = 425 for HNC) [. Notably, the mutational profiles between HPV-negative HNC and HPV-positive HNC are different [The mutational profiles of HNSCC have been reported by The Cancer Genome Atlas (TCGA) and cBioPortal databases. In general, the majority of HNSCCs present loss of function in tumour suppressor genes , and thi = 2798) . Also, Dfor HNC)  summarizifferent , they arTP53, whose mutations are in 73–100% of HPV-negative HNSCC cases [TP53 is a tumour suppressor gene that functions as a gatekeeper for cell growth and division [TP53 mutation is rarely seen in HPV-positive HNSCC, this may be because of the HPV E6 viral protein initiating the degradation of TP53 [TP53 mutation can be regarded as an early event during tumourigenesis in HNSCC [TP53 mutation usually respond poorly to cisplatin-fluorouracil neoadjuvant chemotherapy [TP63 (tumour protein 63) encodes a member of the p53 family of transcription factors. Recurrent focal amplification for 3q26/28 involving the TP63 locus occurs in 15% of HNSCC [The most common mutation in HPV-negative HNSCC is CC cases . TP53 isdivision . This inin HNSCC . HNSCC potherapy , leadingotherapy . In addiof HNSCC .CCND1) and cyclin dependent kinase inhibitor 2A (CDKN2A) are two genes involved in cell cycle and DNA repair pathways. Amplification of CCND1 and deletion of CDKN2A occur in 94% of oral squamous cell carcinoma (OSCC) [CDKN2A [FAT1  (6%) in HNSCC [FAT1 either by mutation or homozygous deletion can activate Wnt signaling pathway to promote tumorigenesis [Cyclin D1 (a (OSCC)  and stru [CDKN2A . In addiin HNSCC . Moreoveigenesis .(EGFR) is a receptor tyrosine kinase (RTK) that is frequently altered in HNSCC [EGFR or other RTKs can active the PI3K/Akt pathway. More than 10% HNSCC patients have amplifications on EGFR of chromosome 7 [EGFR copy number variations (CNV) is linked to poor prognosis in HNSCC [Epidermal growth factor receptor in HNSCC  and stimmosome 7 , 147. Grin HNSCC .(AJUBA), a gene in WNT/β-catenin signaling pathway, is found to inactivate mutations in HPV-negative HNSCC [Ajuba LIM Protein ve HNSCC . It can ve HNSCC .FAM135B) is a cancer-related gene on chromosome 8q. It has been shown to increase progression of esophageal squamous cell carcinoma (ESCC), and mutation of FAM135B in ESCC corelated to poor prognosis [Family with sequence similarity 135, member  encodes a small GTPase in the Rho family, regulating cell motility and tissue development [RHOA mutations in gastric cancers [Ras homolog family member A elopment . RHOA mu cancers  can indu cancers , its rolNSD1) encodes a protein containing a SET domain. Truncating mutations, novel focal deletions (includes homozygous deletions and inframe deletions), missense point mutations and inactivating mutations are found in NSD1 in HNSCC [NSD1 are seen as a favorable prognostic biomarker [Nuclear receptor-binding SET domain protein 1  acts as an oxidative stress factor regulating antioxidant and stress-responsive genes. It only mutates in HPV-negative HNSCC, and heavily related to smoking [Nuclear Factor Erythroid 2-Related Factor 2 ( smoking .CASP8) is located on chromosome 2 and is involved in cell death through the death receptor pathway. Knockdown of CASP8 makes HNSCCs susceptible to necroptosis [CASP8, inversely contributing to the risk of HNSCC development [CASP8 mutation frequency is associated with lower aggressiveness in HNSCC. In addition, CASP8 mutations are found in 10% OSCC tumours [Caspase-8 . Lui’s et al. discovered PTEN gene copy loss in 4/45 HNSCC cases [In HPV-positive HNSCC, PI3K/Akt signaling pathway is the most mutated signaling pathway  and has mutation . HPV-pos tumours . PI3K isCC cases , and thiCC cases , 124.HRAS gene are seen in low frequencies (5%) in both HPV-positive and HPV-negative HNSCC [HRAS is an oncoprotein, which interacts with the PI3K complex in a GTP-dependent manner to increase the catalytic activity of PI3K kinase [Mutations in ve HNSCC . HRAS isK kinase .NOTCH1 causes loss of proliferation and senescence in HNSCC cell lines [NOTCH1 mutation [Although the NOTCH pathway is oncogenic in some types of cancer, its role in HNSCC seems to be tumour suppressive , 139. Nell lines . Approximutation , 139.FBXW7 is a member of F-box protein family and acts as a tumour suppressor gene that mainly targets NOTCH1 [FBXW7 in HPV-positive HNC [FBXW7 can modulate the NOTCH pathway. Studies have also shown that HNSCC patients with TP53 mutations had significantly higher mutation rates in FBXW7 [s NOTCH1 . Its muts NOTCH1 . Lechnertive HNC , whilst tive HNC . It is hin FBXW7 .EP300) is located on chromosome 22 and acts as a histone acetyltransferase. It regulates transcription by chromatin remodeling [E1A binding protein P300 (CREBBP) acts as a tumour suppressor gene and encodes a protein that participates in chromatin remodeling. It is reported to have loss-of-function mutations in many types of malignancies [EP300. Loss of function of CREBBP/EP300 is documented to increase the proliferation ability of tumour cells [cAMP-response element binding protein-BP  mutation (p.D321N and E322K) correlates to Erlotinib sensitivity in HNC patients [and lung . Frameshutations . Mutatioenhancer  regulatienhancer . Furtherpatients , 164.cfDNA is separated either by using centrifugal columns or magnetic beads  for downTP53 mutation at frequencies down to 0.02%. Details of ctDNA detection technologies are summarized in Table Besides the first generation and the next-generation sequencing technologies, Oxford Nanopore Technology (ONT) is the third-generation sequencing that relies on the detection of electrical changes as nucleic acids passing through a protein nanopore. This technology is predominantly used in sequencing long-length sequences, such as genomic DNA . More rectDNA has been widely applied in the early detection of cancer, predict tumour burden, monitor response to treatment . As illuLevels of cfDNA is associated with the stage of the tumour and can indicate disease progress. Hilke et al. sequenced 20 tumour samples from locally advanced HNSCC patients and followed them longitudinally during and post treatment and found that 85% of patients had detectable cfDNA and that the amount of cfDNA correlated with the gross tumour volume . Lin et Circulating HPV DNA (ctHPVDNA) has widely been used as a biomarker in disease prediction and treatment monitoring in patients with HNSCC. Cao et al. reported that pre-treatment ctHPVDNA copy number was closely associated with the metabolic activity of lymph nodes and tumour volume in a 64 HNC patient cohort. A reduction in ctHPVDNA copy number was seen in 14 patients receiving chemoradiotherapy. Similarly, the ctHPVDNA levels were elevated in 13 HNSCC patients coinciding with the time of metastasis, further providing evidence that ctHPVDNA levels can be used as a prognostic biomarker . DahlstrHowever, it is worth mentioning that even though the above studies all focused on ctHPVDNA, they used different probes of HPV, controls, and analysis methods. The following Table Infection with Epstein-Barr Virus (EBV) contributes to the development of nasopharyngeal carcinoma (NPC) . SimilarctDNA mutation profiles have been evaluated to monitor response to treatment in HNSCC patients , 130. HoKRAS, NRAS or HRAS mutations after cetuximab treatment [TP53 mutations in plasma of six HPV-negative HNSCC patients [PIK3CA E545K mutations were detected in the plasma samples from 9/29 HNSCC patients [Schwaederle et al. analyzed ctDNA from various cancer types (HNC = 25) and concluded that HNC was an independent predictor for a higher number of alterations in ctDNA  . Braig ereatment . van Ginpatients . PIK3CA patients .When compared with other cancer types, there is dearth of data relating to tumour tissue mutations and ctDNA mutation profiling in HNSCC. Table CDKN2A, MGMT, GSTP1, and DAPK1) in primary tumour samples and matched serum samples from HNSCC patients (N = 50). They found similar DNA methylation profiles between tumour tissue and serum DNA (21 and 50 respectively). For those patients with serum positive hypermethylated DNA, 5 out of 21 developed recurrence, while only 1 out of 29 patients who relapsed had negative serum methylation DNA [RASSF1 (17.5%), CDKN2A (22.5%), DLEC1 (25.0%), DAPK1 (51.4%) and UCHL1 (64.9%). When combining four-gene methylation markers  in predicting NPC, it gave the highest sensitivity and specificity [EDNRB, p16 and DCC methylation by analyzing serum DNA from HNSCC patients and revealed that serum EDNRB hypermethylation was highly specific for HNSCC but it was not sensitive [SHOX2 and SEPT9 in serum from HNSCC patients (N = 284) correlated with tumour and nodal category and was associated with higher risk of death [CCNA1, DAPK, CDH8 and TIMP3 between FFPE tumour samples (N = 52) and corresponding plasma samples (N = 15). They detected methylation in 73% of plasma samples, while methylation of CCNA1 was related to recurrence-free survival [PENK, NXPH1, ZIK1, TBXT and CDO1 between pre-treatment and post-treatment ctDNA. Table When regulatory regions of tumour suppressor genes are methylated (tumour suppressor genes), their expression levels are reduced, leading to the development of tumour , 110. Sation DNA . Tian etcificity . Mydlarzensitive . Schröckof death . Jesus esurvival , compareDAPK1 51.% and UCHMicrosatellite sequences are short non-coding repeat sequences that vary in length between individuals. Nawroz-Danish et al. reported that 45% (68/152) of HNSCC patients had microsatellite alterations in the DNA isolated from serum samples and was identical to the alterations observed in corresponding tumour samples. Furthermore, 16 HNSCC patients with distant metastasis, 11 had detectable microsatellite alterations in DNA derived from serum with one or more markers . Nunes eAllelic imbalance (AI) is a condition in which the expression levels of two alleles of the same gene differ in the same cell, either as a result of the epigenetic inactivation of one of the alleles or as a result of genetic changes in the regulatory regions. Hamana et al. analyzed AI in tumour tissue and serum samples from OSCC patients (N = 64) at three time points  and found that 52% patients’ serum samples showed AIs in at least one locus and AIs were frequently detected pre-operatively and post-operatively. Importantly, OSCC patients who had AIs during the post-operative period but turned negative 4 weeks post-operatively were free of disease. In contrast, patients who were AI-positive both post-operatively and 4 weeks post-operatively deceased due to distant metastasis. Therefore, microsatellite status in the serum DNA could be used as a potential biomarker in monitoring disease progression .Ahn et al. investigated HPV16DNA levels in saliva and plasma samples from OPSCC patients (N = 93) pre-treatment and post-treatment. For pre-treatment samples combining saliva and serum, the sensitivity, specificity, negative predictive value, and positive predictive values of HPV16DNA were 76%, 100%, 42%, and 100% respectively . Hanna eNext-generation sequencing (NGS) of tumour tissue DNA is emerging as a promising avenue to comprehensively characterize tumour mutation burden. High nonsynonymous tumor mutational burden (TMB), evaluated by WES has shown to correlate with improved clinical outcomes for patients with other types of cancer (lung cancer). However, the use of tumour biopsies to discern clinically available biomarkers have limitations. These include tumour heterogeneity, access to tumour tissues in anatomically challenging locations, insufficient quantity of tumour DNA and the inability to monitor response to treatment in patients who have undergone surgical resections.Liquid biopsy-based applications are revolutionizing the management of patients with cancer . StudiesThe lack of biomarkers to triage the risk of relapse at diagnosis, disease surveillance and predicting recurrence are considered as the main contributors for poor outcomes in HNSCC. To date, most of the research in liquid biopsies has focused on blood-based biomarkers, predominantly using ctDNA. The analysis of ctDNA has several benefits over traditional tumour biopsy testing. Liquid biopsy enables real-time monitoring of response to treatment, also including those patients with tumours in anatomically challenging locations. However, well-designed multi-center clinical trials using homogeneous HNSCC patient cohorts where the use of ctDNA as a biomarker for disease management should provide meaningful benefits to patients before it is broadly implemented clinically.


For staring infrared sensors in geostationary orbit, the clutter caused by the high-frequency jitter and low-frequency drift of the sensor line-of-sight (LOS) is the impact of background features, sensor parameters, LOS motion characteristics, and background suppression algorithms. In this paper, the spectra of LOS jitter caused by cryocoolers and momentum wheels are analyzed, and the time-related factors such as the jitter spectrum, the detector integration time, the frame period, and the temporal differencing background suppression algorithm are considered comprehensively; they are combined into a background-independent jitter-equivalent angle model. A jitter-caused clutter model in the form of multiplying the background radiation intensity gradient statistics by the jitter-equivalent angle is established. This model has good versatility and high efficiency and is suitable for the quantitative evaluation of clutter and the iterative optimization of sensor design. Based on satellite ground vibration experiments and on-orbit measured image sequences, the jitter-caused clutter and drift-caused clutter models are verified. The relative deviation between the model calculation and the actual measurement results is less than 20%. Staring infrared sensors in geostationary orbit can achieve the high-frame-rate detection of fixed positions and have the advantages of high spatial resolution, high temporal resolution, and high sensitivity. They play an important role in the field of infrared point target detection. Under temporal infrared point target detection, the sensor noise and the clutter caused by the line-of-sight (LOS) motion are important factors that affect the sensor detection performance. Sensor noise is basic and unavoidable, caused by various independent noise sources; due to the influence of factors such as the detector material and manufacturing process, the suppressible level of sensor noise is limited. Due to the inhomogeneity of the spatial distribution of the background radiation, the LOS motion leads to changes in the radiant energy on the detector, resulting in jitter-caused (high-frequency) and drift-caused (low-frequency) clutter. With the continuous improvement of the sensor detection sensitivity and spatial resolution requirements, the impact of LOS motion on sensor detection performance is increasing. Therefore, quantitative modeling of LOS-motion-caused clutter is useful for the prediction of system noise and the adjustment of the sensor parameters.CET) into two categories: the drift-equivalent target and the jitter-equivalent target. The drift-equivalent target refers to the detector output fluctuation caused by the LOS drift; the jitter-equivalent target refers to the detector output fluctuation caused by the random shaking of the LOS. Lawrie [CET and described CET as a function of the background power spectral density, optical system parameters, detection distance, and LOS motion rate. Myers [CET, ground sample distance (GSD), LOS drift rate, and jitter amplitude. Fraedrich [The clutter caused by LOS motion has been studied by many scholars. Cota  proposed. Lawrie ,3 analyz. Lawrie ,5 evalua. Lawrie ,7,8 prope. Myers  simulateraedrich  evaluateraedrich  showed aGSD. Hu [Typically, for instruments performing the precise imaging of celestial objects or remote sensing of the Earth’s surface, jitter is a critical requirement for high-quality imaging ,13. ThisGSD. Hu  considerIn this paper, In this section, the jitter is described as the two-dimensional time-varying angular deviation Therefore, the temporal variation in irradiance at a fixed point The temporal variation in radiant flux on a single detector can be related to the spatial variation at time Therefore,The radiant flux change caused by the jitter displacement in the direction of For the staring infrared sensor in geostationary orbit, the radiance on a small rectangular area Similarly, the dimensions of the resulting image area are given byThe average irradiance of the small area is the collected power divided by the imaging area; then,From Equations (1), (7), and (9), the relationship between the radiant flux on the detector and the background radiance is given byGSD constraints on x and y are given byTherefore, it is necessary to differentiate Equation (10). The The GSD, which can be evaluated from two-dimensional image data. The background is expressed as a two-dimensional array of pixel size The integral values on the right-hand side of the above two equations are completely dependent on the background and The unit of Similarly, the gradient component in the y direction is approximately given byEquations (18) and (19), combined with the definition of the gradient, give all the information required to find The calculation result of Then, the relationship between The imaging features of the sensor are related to the background radiation intensity gradient. The equation related to the radiant flux distribution on the focal plane of the sensor to the background radiation intensity distribution has been derived in this section. The relationship between the radiant flux gradient on the detector and the background radiation intensity gradient has been analyzed.For a single detector, the jitter-caused clutter is related to the jitter spectrum, detector integration time, frame period, and background suppression algorithm. These factors can be combined into the jitter-equivalent angle The number of electrons The statistical characteristics of the function ,7. The aX, Y is defined asStatistically, the correlation function of two temporal random variables τ isFor a stationary random process, the autocorrelation function of the time difference In the field of signal processing, Therefore, from Equation (25), From the radiation flux gradient model and Equation (6) in Therefore, from Equations (26) and (27), the correlation function of the radiation flux change isFor the background suppression algorithm of temporal differencing, background suppression refers to the differencing of image DN values or the detector outputs. If the frame period of the sensor is The electrical power of a signal is proportional to the square of the amplitude. The mean squared amplitude of ns,t is (30)<ns,Since the jitter is a stationary random process, In order to calculate Equation (26), the sampling correlation function Then, the mean square differenced output isFrom Equation (22), ssed see  as(33)RThe jitter statistics can be given by the power spectral density Then, the sampling correlation function is expressed by the power spectral density asFinally, substituting Equations (37) and (38) into Equation (32), the mean square differenced output isS of the temporal differencing algorithm asTherefore, the jitter-equivalent angle 2/Hz, S is dimensionless, and the unit of From Equation (41), since the unit of According to Equations (21) and (39)–(41), for a single detector, the RMS differenced output isFor the entire focal plane, since all the factors in Equation (42) are the same except for JCET) for the entire focal plane isAccording to the relationship between radiance and the electron numbers, the average jitter-caused clutter intensity (It can be found from Equation (44) that the jitter-caused clutter intensity is proportional to the jitter-equivalent angle Equation (44) shows how to decompose the jitter-caused clutter into several independent components. The low-frequency drift-caused clutter is directly related to the LOS drift rate DCET) can be equivalently calculated byThe drift-caused clutter intensity (As can be seen from Equation (46), the drift-caused clutter intensity is proportional to the LOS drift rate Since the jitter-caused clutter and drift-caused clutter will reduce the detection performance, predicting the clutter can contribute to the control of LOS stability. Firstly, in order to verify the clutter model, considering the diversity of influencing factors in orbit and the limitations of measurement instruments, the ground measurement experiment was developed and the LOS jitter spectrum was analyzed. Then, the drift displacement of the on-orbit image sequences was measured, and the LOS drift rate was calculated. According to the clutter model, the jitter-caused clutter and drift-caused clutter intensities of different backgrounds were calculated. Finally, with the on-orbit image sequences, the clutter intensities were counted, and the jitter-caused clutter and drift-caused clutter model were verified. The sensor parameters are shown in The staring infrared sensor in geostationary orbit has a long integration time, and it is more susceptible to vibration than a low-orbit satellite sensor. According to the design of the satellite, the active parts mainly consist of a data transmission antenna, solar array drive mechanism (SADA), momentum wheel, and pulse tube cryocooler. The digital transmission antenna does not move during imaging. The fundamental frequency of the SADA is low, and the vibration energy is mainly concentrated in the low-frequency range of 0.2–20 Hz. According to the disturbance test of the SADA, its disturbance force is smaller than that of the cryocooler and momentum wheel. Therefore, the main vibration sources during the imaging of the staring infrared sensor in geostationary orbit are the cryocooler and the momentum wheel.In the experiment, six angular displacement sensors are used to measure the LOS jitter characteristics. The arrangement of the measuring points is shown in The measurement principle of the angular displacement sensor is based on the Sagnac effect, and the source of error mainly includes zero value drift and random variance. Due to the vibration isolation effect of the free boundary simulation device, the high-frequency components caused by the environmental mechanical noise are attenuated and the low-frequency components are amplified. The typical noise spectrum is shown in Since the disturbance frequency of the cryocooler considered in this experiment is mainly 50 Hz and its multiples, and the disturbance frequency of the momentum wheel is distributed above 20 Hz, it is necessary to reduce the noise of the angular displacement sensor. For the disturbance of the cryocooler, filtering and noise reduction were carried out through the band-selective and band-pass methods. For the disturbance of the momentum wheel, the adaptive noise cancellation method was used for noise reduction.The cryocooler provides low-temperature cooling for the infrared focal plane. When the cryocooler is working, the imbalance of the momentum of the moving parts such as the piston and the fluctuation in the pressure of the high-pressure gas will generate disturbance force.The frequency spectra of the RX and RY direction angle vibration of the primary mirror caused by the cryocooler are shown in As can be seen from The jitter power spectral density in the RX and RY direction is approximated asMomentum wheels contain inertial components that rotate at high speeds to generate control torque. Due to processing errors and other reasons, the center of inertial component mass does not coincide with the actual rotation axis, resulting in unbalanced rotation and vibration. The satellite in the experiment has two momentum wheels, which are 0.5 Nm and 0.1 Nm.The frequency spectra of the RX and RY direction angle vibration of the primary mirror caused by the momentum wheels are shown in As can be seen from The jitter power spectral density in the RX and RY direction is approximated asAccording to the frequency spectrum of the angular vibration, the jitter-equivalent angle is calculated. The jitter-equivalent angles caused by different disturbance sources are shown in As can be seen from Based on the continuous image sequence, the LOS drift rate is approximately calculated from image geometric registration. In order to calculate the average LOS drift rate under the typical working conditions, the deviations of the on-orbit image at 10 consecutive moments are required, as shown in IFOV is 16.7 μrad). Therefore, the LOS drift rate is 1.45 μrad/s.As can be seen from In this section, the clutter intensities of different backgrounds are calculated, and the jitter-caused clutter and drift-caused clutter models are verified. The sensor parameters are shown in According to the clutter model in As can be seen from SET) mainly consists of three parts: sensor temporal noise (NET), jitter-caused clutter (JCET), and drift-caused clutter (DCET). Therefore, SET is given byFor the staring infrared sensor in geostationary orbit, the temporal clutter  and actual measurements. In addition to calculation from the model, the The following can be seen from (1)SET calculated by the model is similar to the actual measurement results, and the relative deviation is less than 20%. Under the same system condition, the larger the background radiation intensity gradient is, the greater the SET is.The (2)Under the actual LOS jitter amplitude and drift rate, for scenes with strong clutter, the background radiation intensity gradient is large. The drift-caused clutter dominates, followed by sensor noise and jitter-caused clutter. However, for scenes with weak clutter, the background radiation intensity gradient is small, and sensor noise dominates, followed by drift-caused clutter and jitter-caused clutter.(3)The jitter-caused clutter and drift-caused clutter are related to the background radiation intensity gradient and the LOS motion. The proportion of low-frequency drift-caused clutter and high-frequency jitter-caused clutter is related to the integration time and frequency characteristics of LOS motion.GSD.According to Equations (40), (41), and (44), the jitter-caused clutter intensity is proportional to the jitter-equivalent angle According to the jitter-caused clutter model, the jitter-caused clutter intensity of different scenes with different integration times is calculated, as shown in As can be seen from kG is defined asAccording to Equation (20), the background can be characterized by image pixel data. The statistic kG/GSD is described in terms of the number of pixels instead of the dimension.In order to compare the relative complexity of different spatial structures, the normalized RMS gradient kG/GSD is calculated, as shown in According to the statistical analysis of the on-orbit images, the kG/GSD. Therefore, from Equation (44), the jitter-caused clutter also increases with GSD. According to the fitting results of the power function, kG/GSD.As can be seen from The statistical and fitting results show that the power exponent depends on the spatial structure of the image. Low values occur when there are large areas with little variation in intensity, such as the sea scene, whereas high values occur for images dominated by small-scale structures, such as cloud scenes.GSD reduced. Further reductions in the JCET can be achieved by increasing the complexity of the background suppression algorithm.In order to minimize the jitter-caused clutter, the allowable jitter must be minimized and the SCR) on the image are broken down into requirements of sensor noise and background clutter intensity.In terms of sensor design, the requirements of the target detection performance and signal-to-clutter ratio (SCR (TCR).The minimum target radiation intensity detectable by the system can be calculated from the threshold of NET is fixed. When the temporal target detection method is adopted, the LOS motion converts the spatial variation in background radiation into temporal clutter. Therefore, the LOS motion-caused clutter (CET) consists of two parts.For staring infrared sensors in geostationary orbit with fixed parameters, NET and CET.When the background, sensor parameters, and characteristics of LOS motion remain unchanged, the target detection performance can be quickly calculated, and the main factors can be judged by comparing the values of NET means better sensor performance. The sensor performance can be improved by reducing the readout noise, increasing the aperture, extending the integration time, and increasing the transmittance and quantum efficiency.NET is an important parameter to evaluate sensor performance. A smaller NET, CET, and SET of different backgrounds with different GSD are calculated, as shown in According to the on-orbit image, the GSD increases, the CET and SET increase, resulting in a decrease in the detection performance. Under different scenes, the main factors affecting the detection performance are different. For the backgrounds of sea and land, the main factor is NET. For the background of clouds, the main factor is different with GSD. When the GSD is less than 525 m, NET is the main factor; when the GSD is greater than 525 m, CET is the main factor. At this time, it is the background that mainly affects the detection performance. Improving the GSD can reduce the SET and effectively enhance the detection performance of the system. At the same time, background suppression and spectrum optimization are also primary considerations in improving the detection performance.As can be seen from In this paper, the jitter-caused clutter of a single detector on the focal plane is analyzed, and the clutter model caused by the two-dimensional LOS jitter is established. The model shows that the jitter-caused clutter intensity of the detector is proportional to its corresponding background radiation intensity gradient, and the average jitter-caused clutter intensity of each detector is proportional to the RMS background radiation intensity gradient. Therefore, the distribution of jitter-caused clutter is related to the gradient distribution of background radiation intensity. Based on the equivalence of the LOS drift angular displacement and jitter-equivalent angle, the LOS drift-caused clutter model is established. Taking the on-orbit measured image as the background, combined with the data of the satellite ground vibration experiment and the on-orbit image displacement, the LOS jitter spectrum and LOS drift rate were analyzed, and the jitter-caused and drift -caused clutter model were verified experimentally.The jitter-caused and drift-caused clutter model proposed in this paper provide a reference for the quantitative prediction of clutter and the control of sensor LOS stability.


Twitter has been adopted by physicians across most medical specialties; it allows for the wide dissemination of information and calls to action, brings new stakeholders into collations, promotes academic engagement, and fosters collaboration between academia and private practice. In this review of the literature, we briefly outline the state of advocacy in health care and summarize current Twitter-based advocacy efforts in the major specialties of health care, identifying both successful strategies as well as gaps in Twitter advocacy research. Relevant articles were obtained via PubMed and Google Scholar searches using the phrases “Twitter advocacy healthcare,” “[specialty name] Twitter” and “[specialty name] Twitter advocacy.” Several overarching themes were found to be widely utilized in specialty-specific discussions of Twitter advocacy efforts: organizing under a specific hashtag, fostering dialogue between stakeholders, and tweeting using personalized, action-oriented language. Fields such as pediatrics, heme/onc, ENT, and ophthalmology have most thoroughly embraced the desire to learn how to most effectively advocate on Twitter. Other fields such as OBGYN, cardiology, and surgery have less academic focus on online advocacy. Outside of advocacy efforts, the research and academic benefits of Twitter are well described in nearly every specialty. In conclusion, while clinicians are encouraged to advocate online, only broad strategies for online engagement are currently offered. Additional research into the details of how to successfully create an online profile and Twitter presence is needed to ensure all physicians are able to maximize their advocacy efforts, with clarification of the goals and objectives of this engagement also required. With the broad definition of “speaking on another’s behalf” and working within existing systems to promote the voices of others , advocacSocial media has been well established as an “important tool for scientific collaboration, education, advocacy, patient engagement, and improved patient care” . In the Twitter has been a growing platform for health research, advocacy, and academic efforts . TwitterIn this review, we examine and contrast the Twitter engagement of several of the main medical specialties with a focus on the state of advocacy efforts. We highlight successful organizing strategies employed by different medical communities that may be generalized to other fields. Examples of patient education, medical education, and dissemination of research findings are included in this review for specialties that are not performing more traditional examples of advocacy, as dissemination of information can be a component of advocacy when it is directed to stakeholders unaware of patient needs .MethodsArticles for this review were obtained via PubMed and Google Scholar, using the following search phrases: “Twitter advocacy healthcare,” “[specialty name] Twitter” and “[specialty name] Twitter advocacy.” Articles that were related to the use of Twitter by the medical profession were reviewed and included depending on whether they had relevance to advocacy. All article types were accepted. For specialties without specific Twitter advocacy-related articles, articles that described network characteristics, types of Twitter engagement, or outcomes of Twitter utilization were included. As Twitter is a worldwide platform, articles were not excluded based on their country of origin but were required to be authored in English.Use of Twitter by specific groups of physicians PediatricsPediatricians have embraced online advocacy efforts by organizing around the hashtag “tweetiatrician” . The hasTwitter has also been used for educational campaigns to spread information related to sudden infant death syndrome (SIDS) and safe sleep practices . In an aPsychiatry In psychiatry, Peters and colleagues identify the potential for psychiatrists to intervene in Twitter’s suicide risk detection mechanisms, as well as substance abuse and mood states, particularly for adolescents and young adults . The autNeurologyMishra and colleagues provide one of the only peer-reviewed papers on the use of Twitter in neurology . NeuroloRadiation oncologyIn radiation oncology, Twitter is primarily used for the dissemination of information between academicians. #radonc is used by professional societies with a hub-and-spoke broadcast network wherein posts from a few accounts are shared widely by many other users . In an aResearchers have also leveraged Twitter to promote recent publications, as a correlative analysis between the number of Tweets and the number of academic citations revealed a significant positive correlation between the two . This suDermatologyDermatologic tweets are dominated by personal and educational content. Analysis of a large sample of acne-related tweets revealed that personal tweets were twice as prevalent as tweets pertaining to celebrities or educational dissemination . The mosWhile many dermatologists have amassed large followings on social media, prominent dermatologic influencers are more likely to be females living in metropolitan areas, focusing on cosmetic dermatology in private practice . SimilarConcerning academic engagement, a 2014 review found that most dermatological journals, professional organizations, and patient-centered organizations were using social networking sites, including Twitter, Facebook, and LinkedIn . With thOtolaryngologyThe landscape of Twitter in the otolaryngology field has evolved significantly over the past five years. With limitations on in-person opportunities for the residency application process, there has been a growing academic presence on Twitter . Of the Advocacy-specific efforts in the ENT field have been centered around the hearing loss community, with the majority of Twitter accounts belonging to for-profit or commercial organizations . In a coOphthalmologyTwitter has been recognized as a valuable tool by the field for over a decade, as an early call to Twitter published by Micieli and Micieli in 2012 highlights the many benefits, for journals, professional organizations, and for advocacy . They exAs of 2015, relatively few ophthalmology journals (18.7%) and members of the International Council of Ophthalmology (12.8%) were present on Twitter; however, patient advocacy groups were more popular than journals or other academic organizations . TwitterOBGYNInterest in the use of social media in the field of OBGYN has recently increased in the literature over the last several years and was a popular topic at the 2020 American College of Obstetricians and Gynecologists (ACOG) meeting. At that conference, Noureddine and Chappelle quantified engagement on Twitter. They found that in September 2019, the majority of tweets published using the keywords “obgyn” and “obstetrics and gynecology” were authored by non-healthcare professionals, of which 85.6% were opinion-based and only 9.2% relayed educational content . In contThe academic benefits of Twitter have been relatively less recognized in the field of OBGYN compared to other fields. While the medical education tool “ObGyn Delivered” has been created and tested on various platforms including Twitter, this tool is targeted at undergraduate medical education . As of 2SurgeryIn a 2021 analysis of the top general surgery influencers on Twitter, 50% of influencers were attending general surgeons, 3% were general surgery fellows, and 31% were general surgery residents . The vasWhile seemingly a specialty that would have less opportunity for remote supplementation of care, Leow and colleagues advocate for the use of Twitter as a means for providing diagnostic assistance to surgeons and patients in low-resource settings . In thisBeyond advocacy, surgery has heavily embraced the use of Twitter for academic purposes. Trauma and orthopedic surgery journals with Twitter accounts have higher impact factors; Twitter follower count is positively associated with impact factors . Twenty-CardiologyThe field of cardiology has embraced the academic utility of Twitter. This includes building awareness and interest in conferences , providiWhile there are no publications on traditional advocacy efforts on Twitter in the field of cardiology, there is growing recognition that Twitter can be utilized to survey perceptions, knowledge, and topics of engagement among users outside of the medical field ,66. SuchHeme/OncHematology and oncology fields have been primarily organized using hashtags . DiseaseWhile less proliferative than the field of oncology, hematology has a growing presence on Twitter . With lePathology and other specialtiesAdditional specialties have active roles on Twitter, using the platform to disseminate information. For example, pathology has paid special attention to the teaching capabilities of Twitter. Similar to the teaching taking place between pathologists using a multi-headed microscope in person, Twitter can be used to increase equity in education, as images may be shared and discussed in threads . AdditioInterdisciplinary effortsWhile most physician advocacy on Twitter has been studied in specialty-specific networks, there are examples of specific causes for which multidisciplinary teams of physicians on Twitter have worked together, namely #thisisourlane and #getusPPE. As previously mentioned, #thisisourlane sought to unite physicians, regardless of specialty, to assert the collective role of healthcare providers working to prevent gun violence. While #getusPPE was created by emergency medicine physician Dr. Esther Choo, it rapidly spread to include all healthcare workers who were treating patients during the COVID-19 pandemic. Tweets created using these healthcare-led hashtags were analyzed by Ojo and colleagues (2021), comparing the language used in tweets containing these healthcare-led hashtags to a control hashtag . The tweAcademic physiciansTwitter is a valuable place for open-access medical education that can be easily updated and shared with set groups . AdditioApplications for residency curriculaTwitter has become an important communication tool for physician advocates. In 1996, the Pediatric Residency Review Committee of the Accreditation Council for Graduate Medical Education (ACGME) added a requirement for pediatric residents to have structured educational experiences to “prepare residents for the role of advocate[s] for the health of children within the community” . While oTwitter advocacy by other organizationsThere exists great potential for effective advocacy through partnerships with a variety of individuals and organizations on Twitter. In 2012, Lovejoy and Saxton utilized an information-community-action classification scheme for tweets from the 100 largest nonprofit organizations with Twitter accounts. The majority of such tweets contained information (58.6%), while 13.2% of tweets gave recognition or thanks, and 8.2% of tweets were replies to other messages . The autThe successful advocacy of environmental scientists in the Flint Water Study provides an interesting and relevant case study for advocacy in health care. These scientists used a wide variety of advocacy techniques throughout their tweets, relying most heavily on public education (34.8% of tweets), expert testimony (21.6% of tweets), and dissemination of science/research (13%) . When loTwitter etiquette As more healthcare professionals become involved in online networking activities, questions have emerged surrounding the “proper” behavior of providers on platforms such as Twitter. Many policies are currently rooted in exercising “common sense” while online; however, as Pershad and colleagues assert, more specific guidelines are necessary . MaintaiThere are five general categories of communicative strategies that have been broadly used by Twitter activists from various disciplines, which include information, symbolic activities, organizing, litigious activities, and civil disobedience activities . When coFurther insight into how tweets can be most effectively composed can be gained from a recent experimental study on the different strategies for the dissemination of COVID guidelines. Tweets that had the highest perceived message effectiveness, as well as perceived attitude effectiveness and the highest likelihood of sharing were tweets authored by physicians that contained a personal message . This waAs the primary downside of Twitter is the ability for anyone, regardless of their credentials, to share health-related information with varying degrees of truthfulness, the presence of licensed healthcare providers to provide accurate, evidence-based information is of increasing importance . BecauseThere are commonalities seen in using Twitter for advocacy across multiple specialties. The use of hashtags used to build communities of physicians within specialties (as done in hematology/oncology and pediatrics) and between specialties  warrants prospective study. Likewise, strategies to expand those coalitions to include non-physician stakeholders should be incorporated into training. There are differences in outcomes demonstrated for different groups of physicians; exploring similarities and differences across specialties may further direct the curriculum to enhance the effectiveness of physician advocacy on Twitter.


Gamete-level sexual selection of externally fertilising species is usually achieved by modifying sperm behaviour with mechanisms that alter the chemical environment in which gametes perform. In fish, this can be accomplished through the ovarian fluid, a substance released with the eggs at spawning. While the biochemical effects of ovarian fluid in relation to sperm energetics have been investigated, the influence of the physical environment in which sperm compete remains poorly explored. Our objective was therefore to gain insights on the physical structure of this fluid and potential impacts on reproduction. Using soft-matter physics approaches of steady-state and oscillatory viscosity measurements, we subjected wild Atlantic salmon ovarian fluids to variable shear stresses and frequencies resembling those exerted by sperm swimming through the fluid near eggs. We show that this fluid, which in its relaxed state is a gel-like substance, displays a non-Newtonian viscoelastic and shear-thinning profile, where the viscosity decreases with increasing shear rates. We concurrently find that this fluid obeys the Cox–Merz rule below 7.6 Hz and infringes it above this level, thus indicating a shear-thickening phase where viscosity increases provided it is probed gently enough. This suggests the presence of a unique frequency-dependent structural network with relevant implications for sperm energetics and fertilisation dynamics.This article has an associated ECR Spotlight interview with Marco Graziano. Summary: The viscoelastic properties of salmon ovarian fluid support a novel mechanism of post-mating sexual selection in external fertilisers with important implications for understanding the evolution of sexual traits. The micro-conditions of fertilisation are poorly understood in the majority of animal species . Followiin vivo research using GFP-tagged sperm revealing high levels of activity and interaction between sperm from different males and different areas of the female tract , where a single egg batch can be sired by up to 16 fathers . However, we do not yet know the exact mechanisms facilitating such choice.Some of our most fundamental knowledge about sperm–egg interactions comes from broadcast-spawning marine invertebrates. The associations between bindin molecules  and betwetration , influenetration . Similaretration . In fishing eggs . Ovarianing eggs  and onceing eggs  and infling eggs  and theiing eggs . In salming eggs . There i fathers . Yeates Sperm swimming propulsion is created by the flagellum, whose function is influenced by chemical  and physTo do this, we collected ovarian fluid from mature females during the reproductive season of Newfoundland wild Atlantic Salmon and analysed its rheological properties. Using a modular compact rheometer (MCR), we applied both steady-state and oscillatory viscosity measurements to determine viscoelastic responses in the non-Newtonian regime, when the ovarian fluids were subjected to increasing shear rates and variable angular frequencies comparable to those exerted by swimming spermatozoa moving in the fluid toward the eggs. By evidencing specific shear-dependent changes in the polymeric structure of the fluid and how these occur, we expected to identify an underlying mechanism that influences partner selection to provide an evolutionary advantage.Salmo salar Linnaeus 1758) were collected in early September from a fish ladder at Grand Falls  during their up-stream spawning migration on the Exploits River . Following previous protocols  were kept in sealed glass jars, enclosed with bubble wrap and placed in a cooler of wet ice for transport to the laboratory. Each egg batch was separated from its ovarian fluid using a fine mesh net  was conducted to assess different fluid preservation methods  nor purely elastic (solid-like) and these rheological properties correlate strongly with their function . Structu−1 in 50 equally spaced steps. The resultant shear stresses of the ovarian fluid were measured to determine the apparent viscosity ηa, which was averaged across three aliquots per female (n=11) and plotted as a function of the shear rate.Samples were tested for their resistance to flow in order to measure their viscosity under a specific rate of deformation. To obtain a flow curve, the shear stress was measured for a range of shear rates , a theoretical positive relationship between shear stress and shear rate should be linear and the fit line should pass through zero. When the profiles of water runs were fitted, a positive intercept  of 0.0133 Pa was concluded to be low shear rate instrumental noise. It was subtracted from all the water and ovarian fluid samples for standardisation [(shear stress–0.0133 Pa)/shear rate], creating a small change in values. A comparison of individual ovarian fluid viscosity profiles with distilled water for each of the instrumental replicates allowed us to assess variability among females.a=0.00151±0.00003 Pa s) was independent of shear rate. The apparent viscosity at −1 was roughly 10 times the viscosity of water, but returned within comparable values under increasing shear rates, starting at around 100 s−1 (see Results). For three females the ovarian fluid samples had apparent viscosities ηa in the order of 0.003 Pa s at 10 s−1, showing no meaningful differences with the rheological behaviour of water at the same shear rate. These samples were probably contaminated with urine and/or water during stripping of gametes and for these reasons were not included in the main results. The remaining 11 flow curves were globally fitted to the form ηa00, which must be overcome before there is flow and a viscous component η∞, which represents the viscosity at very high shear rates. This simple form was arrived at when fitted to a more complicated formula, the Herschel–Bulkley equation n that were very close to unity.The apparent viscosity of ovarian fluid decreased with increasing shear rates, in contrast with water whose apparent viscosity (ηquation   resulted0sint) was imposed on the sample at a fixed frequency (ω) and a maximum amplitude (γ0) , where ω is defined as ω=2 π f, where f is the frequency in Hz. The storage modulus:0 is the oscillatory stress applied and δ is the phase angle, were obtained as a function of frequency (ω). The modulus of the complex viscosity η* was obtained from the relation:G″/G′ represents the ratio between viscous and elastic contributions to the viscoelasticity.To preserve finer polymeric structures and obtain a dynamic profile that informs about the viscous and elastic components, we subjected the ovarian fluid to small-amplitude oscillatory shear. For these measurements, a sinusoidal deformation (γ=γude (γ0) . Measure=a σ/G*(ω)|/ω) at a specific oscillatory frequency (ω), that is:The Cox–Merz rule, an empirical method to rationalise steady shear and oscillatory rheological data , was useeriments .G′ and G″ (dependent variables) across all the sampled females were pair-wise compared through t-tests at specific frequencies (independent variables) of interest within two shear stress ranges, 0.001 to 0.105 and 0.105 to 1 rad s−1, to confirm their uniformity within the plateau region and/or alternatively the prevalence of either the viscous or the elastic component of the ovarian fluid in this dimensional range. Normality of the residuals was ensured by using the D′Agostino–Pearson test followed by Shapiro–Wilk test . In all analyses, the statistical significance threshold used was α=0.05.All ovarian fluid measurements and fish morphological data  were summarised using the descriptive statistics function in GraphPad Prism, v. 8.0.0, . Rheometer reads were first standardised for instrumental error and the model fits were applied as described above. Subsequently, the average values of −1 and CV=17% at 500 s−1 . The resulting shear stress responses from the deformed ovarian fluid were measured to determine the apparent viscosity of the material at each of the measuring points.To measure the viscosity under a linearly increasing rate of deformation, the ovarian fluid samples were tested for their resistance to flow for a range of shear rates (10 to 500 s00=0.09 (± 0.01 Pa) and a mean value of the high shear viscosity η∞=2.3 (± 0.8 mPa s) with the ovarian fluid showing an average 97% decline in viscosity as an increasing shear rate was applied through the rheometer's plate ; P≥0.05, t=2.77, d.f.=4] and describe a pure viscoelastic fluid where the elastic and the viscous components of the fluid are comparable. Both G′ and G″ decreased slightly between 0.01 and 0.07 rad s−1 (note the log10–log10 axes) and thereafter maintained constant plateau values until the shear rate reached 1 rad s−1. Note that this plateau value is numerically proximate, given the errors, to the value obtained for the yield stress in the steady shear measurements. Salmon ovarian fluid is therefore a gel-like structure at low frequencies and becomes more dominantly liquid-like at frequencies higher than 10 Hz. Interestingly, this structural shift occurs in a dimensional range that overlaps with the frequencies exerted by salmon sperm when swimming through the ovarian fluid to reach the egg  the loss modulus G″ (mean 0.081±0.02) was marginally higher  than G′ (0.052±0.01). As observed through the study of their first and second derivatives, G′ and G″ trends start to slowly diverge, more intensely from 10 Pa (at 1.59 Hz) onward revealing a breakpoint in the polymer that exacerbates together with increasing shearing rates (−1 (7.58 and 49.66 Hz), showing that the elastic response of the polymer under these frequencies is null (liquid-like); and viscous forces at their maximum in this frequency range instead prevailed. As a result, the absolute value of the complex viscosity (|η*|) decayed until reaching its a minimum of 0.005 Pa s, at a frequency near 8 Hz (50 rad s−1), , see A. Interea) obtained in the flow curves and absolute values of complex viscosities (|η*|) resulting from the small-amplitude oscillatory sweep experiments, were plotted as a function of shear rate and angular frequency, fitted to the best trend and assessed for deviations between the curve profiles , there is an increase in |η*|.The steady-state properties of the ovarian fluid were compared with the dynamic states by applying the Cox–Merz rule. This rule, applied to polymers, enables the identification of secondary flow behaviours and/or breaking down of the fluid's polymeric network under a certain imposed stress. Apparent viscosities  and angular frequencies (SAOS), that were similar to those exerted by sperm swimming through ovarian fluid to fertilise eggs ,b. This Our results indicate that ovarian fluid, which is a gel at its relaxed state (between solid- and liquid-like behaviour), is a shear thinning viscoelastic liquid at low frequencies and may exhibit a shear thickening phase at high frequencies. This shift from gel to a more passable medium, together with minimum viscosity values observed within the range of average beating frequencies exerted by salmon sperm, points out an interesting overlap that might be linked to ‘bio-mechanical co-evolution’ of female and male gametes. Specifically, Atlantic salmon ovarian fluid has a viscosity at its relaxed state that is on average 60 times that of water, being 0.09 Pa. A hypothetical beating frequency of 1 Hz would yield to the absolute value of the complex viscosity |η*| of 0.017 Pa s (five times lower than at its relaxed state), while a beating frequency of 12 Hz yields 0.006 Pa s. Sperm movement occurs at low shearing rate ; 1966 anOncorhynchus mykiss  and methodology used to detect it . Measures mykiss  and ∼80 awytscha . For thiSalvelinus alpinus) when measured at 0.5 Hz under a plate viscosimeter (O. tshawytscha) ovarian fluid viscosity decreased from 0.0042 to 0.0027 Pa s as shear was increased from 7 to 72 Hz  for Arctic charr  and elastic (solid-like) components of the fluid define its changing complex viscosity and cannot be neglected when analysing sperm energetics and outcome. Bos taurus have higher thrust efficiencies of sperm when swimming in a non-Newtonian fluid rather than in a Newtonian one, which could be due to a better energetic exploitation of the elastic responses of the fluid , viscoelastic reproductive fluids have been found to decrease spermatozoa velocities as viscosity increased. However, this was associated to a concurrent increase in their linearity . Bos tauhe fluid . Similarhe fluid . In our S. salar , it is pS. salar .Sperm traits are under strong selection , with reIt is well accepted that the guidance within reproductive fluids occurs by means of chemical and biochemical cues that can differentially enhance the reproductive outcome from different males as demonstrated in a range of external fertilisers .We propose that more consideration should be given to the physical characteristics of the ovarian fluid that could affect sexual selection processes. Females might be able to facilitate the progression of the high quality and fast beating sperm, within and among ejaculates. Also, in view of the variability observed across females, they might have different capabilities to exert this selective potential and such potential could change with the hydration grade of the ovarian fluid as the reproductive season advances.Gadus chalcogrammus) showed values between 1 and 10 Pa  has been linked to augmenting the chances of fertilising the egg  at higher frequencies was attributed to instrument inertia. We cannot exclude that this could also be the case here. Moreover, although the idea that the ovarian fluid may have a natural selective function at very high shear rates is indeed fascinating, this specific aspect lies outside the scope of this study and was not tested specifically. Future experiments should try to provide insights in this regard by testing the ovarian fluid dispersion capacity from eggs under different shear rates that could better simulate the riverbed waterflow that characterises the reproduction of most salmonids. Also, until the shear thickening behaviour of the ovarian fluid is confirmed, the complete picture of how sperm swim in ovarian fluid is not clear. Currently, all the modelling approaches published, have assumed exclusively a shear-thinning fluid at smaller shear flows such as those experienced by sperm in the genital tract of internally fertilising animal models  .10.1242/jexbio.244712_sup1Supplementary informationClick here for additional data file.