CSF MPO concentration correlated with disease duration in male and female patients (p < 0.01). CSF MPO concentration was significantly higher in men with disease duration ≥12 years versus the remainder of the male subjects (p < 0.01). Changes in CSF MPO in women were not significant. Serum MPO concentration and activity were significantly higher in all PD patients relative to controls (p < 0.0001). CSF MPO was not correlated with serum MPO. Serum AOPP were detected in all patients, but CSF AOPP was undetectable in 53% of patients. AOPP were not quantifiable in controls. Conclusions: CSF MPO is not a good biomarker for PD because mean CSF MPO concentration and activity are not different between the cohort of patients and controls. CSF MPO concentration positively correlated with disease duration in men and women, but CSF MPO is significantly enhanced only in male patients with disease duration longer than 12 years. It can be hypothesized that the MPO-related immune response in early-stage PD might be weak in all patients, but then the MPO-related immune response is progressively enhanced in men, not women. Since the blood-brain barrier is intact, and CSF MPO is not correlated with serum MPO, CSF myeloperoxidase would reflect MPO content in brain cells, not blood-derived cells. Finally, serum AOPP was detected in all patients, but not controls, which is consistent with the occurrence of chlorinative stress in blood serum in PD. The study of CSF AOPP as biomarker could not be assessed because the ELISA assay was hampered by its detection limit in the CSF.Background: Myeloperoxidase (MPO) and advanced oxidation protein products, or AOPP (a type of MPO-derived chlorinated adducts), have been implicated in Parkinson´s disease (PD). Human MPO also show sex-based differences in PD. The objective was to study the relationship of MPO and AOPP in the cerebrospinal fluid (CSF) with motor features of idiopathic PD in male and female patients. Methods: MPO concentration and activity and AOPP content were measured in the CSF and serum in 34 patients and 30 controls. CSF leukocytes and the integrity of the blood-brain barrier were evaluated. Correlations of MPO and AOPP with clinical variables were examined. Results: The blood-brain barrier was intact and CSF leukocyte count was normal in all patients. CSF MPO concentration and activity were similar in the cohort of patients and controls, but CSF MPO content was significantly higher in male patients than in PD women ( The enzyme myeloperoxidase (MPO) belongs to the family of heme-peroxidases, which are essential components of the immune defense system. This defensive function is based on the generation of microbicidal compounds through two biochemical pathways: the halogenation and peroxidation cycles. Antimicrobial molecules are powerful oxidants and pro-inflammatory molecules ,2,3,4,5,substantia nigra pars compacta and contributes to α-synuclein pathology, has been implicated in Parkinson’s disease (PD) [substantia nigra in diseased patients and animal models of PD has been reported [substantia nigra, in the MPTP animal model of PD [MPO is a normal constituent in granules of peripheral neutrophils and monocytes/macrophages as well as in brain glial cells (astrocytes and microglia). MPO biochemical relies on the production of hypochlorite, hypochlorite-modified proteins, chlorinated amines, and other reactive species ,16,17,18ase (PD) ,20,21,22reported ,24,25. Tel of PD ,26. In ael of PD ,28,29, ael of PD ,31. SincAmong hypochlorite-modified proteins, Advanced Oxidation Protein Products or AOPP are suggested to play a role in neuroinflammation and oxidation processes in PD, and they are closely linked to MPO ,34,35,36Brain MPO and hypochlorite-modified derivatives have been proposed to be the target for antiparkinsonian treatment ,37,38,39123I-Ioflupane DAT-SPECT by expert physicians from the Service of Nuclear Medicine [This prospective, observational, and cross-sectional study was carried out at Hospital Macarena, Sevilla, Spain. Thirty-four patients and 30 controls were evaluated after careful selection. Patients had age-at-PD onset of 45–75 years, and they were diagnosed of PD if they presented rigidity, bradykinesia, and resting tremor (classic motor signs of Parkinsonism), as well as a reliable loss of dopamine-transporter signal on dorsal striatum ,43,44,45Demographic information was obtained from patients and controls: Age, gender, body mass index (BMI), and years of education. Clinical data included the modified Hoehn–Yahr staging, the International Parkinson and Movement Disorder Society-Sponsored revision of the Unified Parkinson’s Disease Rating Scale (MDS-UPDRS), the modified Schwab–England activities of daily living scale, age-at-PD onset, disease duration in years, and the antiparkinsonian medication regimen. Antiparkinsonian medication  was expressed as levodopa equivalent daily dose , according to established formula ,46,47,482. All participants were non-smokers, non-alcohol drinkers, and non-coffee drinkers, according to established criteria [Individuals suffering from renal, hematological, liver, and cardiovascular dysfunctions, as well as malabsorption, morbid obesity, dementia, autoimmune diseases, acquired immunodeficiency syndrome, cancer, infectious conditions or taking anti-inflammatory drugs or statins were excluded from the study. MPO and AOPP levels in the CSF can be altered in these conditions ,50. Morbcriteria ,52,53.TM AOPP assay kit, ref. STA-318, Cells Biolab Inc., San Diego, CA, USA), following the manufacturer’s instructions. The sensitivity of the ELISA assay with human biofluids is ~10 pg/mL (MPO) and ~0.1 µM (AOPP). Myeloperoxidase activity was measured with an Enzyme activity kit , following manufacturer’s instructions. This assay kit can be used to detect myeloperoxidase as low as 0.010 mU per mL. The reaction period was 10 min. Each sample was analyzed in duplicate .MPO and AOPP were measured in the CSF. In addition, to further discern whether CSF MPO and AOPP are related to blood molecules, serum MPO and AOPP were also evaluated. CSF was collected by physicians by using lumbar puncture. Two ml of CSF were collected, aliquoted, coded, and rapidly frozen at −80 °C. A 0.5-mL fresh collection was employed to observe the absence of traumatic puncture, and to quantify red cells (CSF with >500 red cells/microL was discarded). Five ml of blood was collected by cephalic vein puncture to obtain serum with a serum separator tube (BD Vacuotainer). Serum was centrifuged at 2500 rpm during 10 min, and then it was immediately aliquoted, and frozen at −80 °C in 0.5 mL aliquots. CSF and serum aliquots were unfrozen and sonicated with homogenizing solution . Myeloperoxidase and AOPP concentration were evaluated with commercially available Enzyme-linked Immunosorbent Assay kits , and these subgroups were compared. Ninety-five percent confidence interval and sample size power were calculated to avoid making Type II error, and the statistical power was set at β = 0.2 or >80% (α = 0.01).Quantitative variables were compared with the Student’s Protocols were approved by the internal ethics and scientific boards of Junta de Andalucia , Hospital Universitario Macarena (CEI #19/05/2010 and CEI #2149-29/10/2013), and University of Seville (CEE-27/05/2010 and Seccion Investigacion/Gd-21/06/2010). The subjects consent was obtained according to the Declaration of Helsinki .Demographic and CSF analytical data were not found to be different between controls and patients with PD, as shown in p < 0.0001)  . Based o 0.0001) .p = 0.0049, p = 0.0045, n = 14) than in the remainder patients  and controls .CSF MPO concentration and activity significantly correlated with each other  relative to patients with moderate-advanced PD . CSF MPO activity was found not to be significantly different in patients with early PD  relative to patients with moderate-advanced PD . Finally, AOPP correlations were not studied because AOPP content in CSF samples was below the detection limit of the ELISA assay in many patients, as explained.No significant correlations between CSF MPO concentration or activity and other parameters such as serum MPO content or disease severity  were observed . Thus, Cp = 0.0064), although no differences with controls were found, as shown in p = 0.0054; MPO activity, t = 5.6278, p < 0.0001). Besides, serum MPO content and activity in women was significantly higher in PD patients than in controls  than in the remainder male patients , and control men .As for effects of disease duration, CSF MPO concentration was observed to be significantly higher in male patients with disease duration ≥12 years  relative to patients with moderate-advanced PD . CSF MPO activity was found not to be significantly different in male and female patients at early-stage PD  relative to patients with moderate-advanced PD . The lack of influence of disease severity on CSF MPO concentration or activity in men and women is illustrated in No significant correlations between CSF MPO concentration or activity and other parameters such as disease severity or serum MPO content were observed . CSF MPOsubstantia nigra pars compacta and contributes to α-synuclein pathology [The enzyme myeloperoxidase, which is expressed by microglia and neurons in the athology ,22,54, hathology ,20. Howeathology .A novel finding of this study is that CSF MPO concentration is correlated with disease duration in male and female patients. MPO concentration is significantly augmented in the CSF in men with disease duration longer than 12 years. However, in female patients, there is a progressive increase in CSF MPO content but differences with the control women are not significant. Therefore, MPO is reliably upregulated in the CSF in male patients with PD, not women, and after long duration of the disease. It can be hypothesized that the MPO-related immune response at early-stage PD might be weak in all patients with PD, but the MPO-related immune response and recruitment of MPO-positive cells would be progressively enhanced in men, not in female patients. In women, the MPO-related immune response seems to be weak throughout the disease.Regarding sex-based differences, it is known that female patients have a higher mortality rate and faster progression of PD , and womsubstantia nigra in PD [Many authors have reported that MPO is over-expressed in postmortem brain tissue ,25,39, ara in PD ,25. On tIn contrast to CSF MPO, serum MPO is significantly enhanced in all patients, confirming previous findings . The finAnother novel finding is that advanced oxidation protein products are detected in blood serum in all PD patients but not in controls. In the CSF, the ELISA assay was hampered by its detection limit, thus challenging its reliability. AOPP are pro-inflammatory chlorinated products that are created through the reaction of hypochlorite and chloramines with proteins ,61,62,63The study has some limitations that should be acknowledged. We cannot infer causation, because it is a cross-sectional design, not a longitudinal study. The cohorts of participants were relatively small, and the detection threshold of the ELISA kit was a major limitation for quantifying AOPP concentration in the CSF. The results must be confirmed by means of larger samples in future studies, and the use of more sensitive methods. The strengths of our study include well-characterized patients with PD, rigorous data collection and selection of participants, the analysis of biofluids that are in close contact with brain tissue, and the measurement of concentration and activity of myeloperoxidase.CSF MPO is not a good biomarker for PD because mean CSF MPO concentration and activity are not different between the cohort of patients and controls. CSF MPO concentration positively correlated with disease duration in men and women, but CSF MPO is significantly enhanced only in male patients with a disease duration longer than 12 years. It can be hypothesized that the MPO-related immune response in early-stage PD might be weak in all patients, but then the MPO-related immune response is progressively enhanced in men, not women. Since the blood-brain barrier is intact, and CSF MPO is not correlated with serum MPO, CSF myeloperoxidase would reflect MPO content in brain cells, not blood-derived cells. Finally, serum AOPP was detected in all patients, but not controls, which is consistent with the occurrence of chlorinative stress in blood serum in PD. The study of CSF AOPP as a biomarker could not be assessed because the ELISA assay was hampered by its detection limit in the CSF.


TM and EuroForMix resulting in a significant probative gain of information, .When a minor DNA component to a binary mixture is present at a weight ratio of approximately 1:50 or less, the presence of this minor donor is undetectable (or barely detectable) by standard mixture deconvolution approaches. In an attempt to retrieve probative minor donor DNA profile information, multiple quintuple cell subsamples were collected from a 1:50 DNA mixture using direct single cell subsampling (DSCS) paired with probabilistic genotyping (PG), the latter validated for use with single or few cells. DSCS employs a simplified micromanipulation technique paired with an enhanced DNA profiling approach, involving direct cell lysis and a sensitive PCR process, to genotype individual cells. Multiple five-cell subsamples were used to interrogate sufficient cells from the mixture such that some of the created 5-cell “mini-mixture” subsamples contained a cell from the minor donor. The latter mini-mixture subsamples, which now comprised weight ratios of 1:4 as opposed to the bulk mixture 1:50, were analyzed with the PG systems STRmix Standard DNA mixture analysis typically targets 0.5 ng to 1 ng DNA for amplification from a bulk DNA extraction process that basically “homogenizes” the mixture components. This can be problematic in instances in which one donor’s DNA is present in such a large proportional concentration that it effectively masks the presence of a minor donor due to PCR reaction kinetics. In many cases, alternative approaches such as physical separation by way of differential extraction ,3. HowevProbabilistic genotyping (PG), until recently, has primarily been applied to bulk samples extracted from DNA stains, and although highly useful, does not always result in probative DNA information for all donors to a mixture due to the presence of allele overlap, artifacts such as stutter, and in the case of this paper, the extreme minor presence of one of the donors ,5,6,7,8.A previous report has shown that isolation of multiple groups of cells from mixtures results in sub-populations with differing constituent weight ratios  that, inThe purpose of this study was to develop a cell subsampling strategy to enable the detection and analysis of a minor donor to a binary mixture that would otherwise be un-detectable or barely detectable using standard mixture interpretation methods. In the present work, the direct single cell subsampling approach, which utilizes the simplified micromanipulation technique paired with direct cell lysis and a sensitive PCR process to genotype individual cells, was used to collect five-cell subsamples from a complex binary mixture (1:50) in which the minor donor was virtually undetectable by standard methods. This resulted in new mini-mixture subsamples (with weight ratios of 1:4 as opposed to 1:50) that could then be analyzed and interpreted by way of probabilistic genotyping, resulting in a gain of significant probative DNA information from the minor donor.Buccal cell samples were collected from volunteers using procedures approved by the University of Central Florida’s Institutional Review Board. Samples were obtained from each volunteer by swabbing the inside of the mouth and cheek with a sterile cotton swab.−4, and the resulting solutions were centrifuged at 300 RCF for 7 min to create an epithelial cell pellet. The supernatants were discarded, and 300 μL of TE−4 was added to each cell pellet to create a cell suspension. The Countess™ II FL  automated cell counter was used according to manufacturer recommended protocols to determine the cell concentration of each cell suspension. The appropriate volumes of each donor cell suspension were mixed to achieve the desired donor mixture ratio . The mixture cell suspension was stored frozen (4 °C) before use.Freshly collected or dried/frozen buccal swabs from individual donors (“SA79” and “S3”) were agitated in 300 μL of TE® Gel-Film®  was affixed to a clean glass microscope slide via its adhesive backing, and the clear protective covering removed. Once the desired mixture was created, 60 μL of the mixture solution was pipetted onto the Gel-Film® microscope slide and spread out using a sterile swab. The slide was then stained with Trypan Blue for 1–2 min and gently rinsed with nuclease-free water. Slides were air-dried overnight.Gel-PakA 3M™  adhesive slide reservoir was prepared prior to cell collection, in which double-sided tape was used to attach the 3M™ adhesive to a clean glass microscope slide. The 3M™ adhesive backing was removed, and the slide was stored in a desiccator until needed ,17.TM  present in sterile 0.2 mL PCR flat-cap tubes by way of a sterile tungsten needle and water-soluble 3M™ adhesive. The tungsten needle was used to acquire a small ball of 3M™ adhesive, which was then utilized to adhere selected visualized cells from the mixture slide. Still under the microscope, the adhesive-tipped tungsten needle containing adhered cells was inserted into an amplification tube (0.2 mL) containing PunchSolutionTM  until the 3M™ adhesive is observed to solubilize [Cells were visualized using a Leica M205C stereomicroscope (190–240× magnification) and collected directly into 5 μL PunchSolutionlubilize ,17. An olubilize .TM  in 0.2 mL PCR flat-cap tubes. Samples were incubated until the PunchSolutionTM  evaporated using a lysis protocol of 70 °C → 30 min. After lysis, GlobalFilerTM Express reaction mix was added to each mixture consisting of 2 μL PCR mix, 2 μL primer mix, 1 μL 5X AmpSolutionTM . Samples were amplified using a modified protocol of 95 °C → 1 min; 32 cycles: 94 °C → 3 sec, 60 °C → 30 sec; 60 °C → 8 min; 4 °C → hold.Cells were collected directly into 5 μL PunchSolutionExpress™ Forensic DNA Extraction System  using the PrepFiler Express™ Forensic DNA Extraction Kits . An extraction blank was included with each extraction set performed.DNA was extracted from reference buccal swabs and 60 μL of the mixture cell suspension using the AutoMate TM  amplification kit according to the manufacturer’s recommended protocols. One nanogram of input DNA was used with an amplification protocol of 95 °C → 1 min; 29 cycles: 94 °C → 10 sec, 59 °C → 90 sec; 60 °C → 10 min; 4 °C → hold. Positive and negative amplification controls were included in each amplification.Amplification of the donor reference samples and the bulk DNA mixtures was conducted using the GlobalFiler® size standard . Samples were injected on an Applied Biosystems’ 3500 Genetic Analyzer using POP-4™ polymer and Module J6  and analyzed using GeneMapper™ ID-X v1.6 software .One microliter of GlobalFiler™ or GlobalFiler™ Express  amplified product was added to a mixture comprised of 9.5 μL Hi-Di™ formamide  and 0.5 μL GeneScan™ 600 LIZTM, the sub-source log (LR) was reported. The FBI Caucasian database was used for all allele frequencies in all mixture experiments [Probabilistic Genotyping Software STRmix™ v2.8 and EuroForMix v3.1.0 (Quantitative LR MLE based) was previously internally validated for use with standard bulk mixtures and reference samples. Each sample was analyzed according to the known number of donors. With STRmixProbabilistic Genotyping Software STRmix™ v2.8 [An infographic of the quintuple cell DSCS method is provided in p = 0.02. The binomial probability is 0.092, meaning that if one performed 75 subsamplings of 5 cells, then approximately 6–7 of them would result in mini mixtures containing the minor donor. A similar 75 subsamplings for a 1:100 mixture ) would result in an expectation of approximately 3–4 of the 100 subsamplings to be comprised of minor donor containing mini-mixtures.To determine at what mixture ratio a minor donor would become undetectable using our standard STR analysis system, two-person DNA mixtures ranging from tenfold ratios of 1:20 to 1:50 were prepared and analyzed. It was determined that a donor ratio of approximately 1:50 was the point at which a minor donor became virtually undetectable using standard methods. Thus, we proceeded to develop and investigate a DSCS strategy that would be applicable to undetectable/unresolvable mixtures with a minor donor ratio of approximately 1/50 but which would also probably be applicable to mixtures with lower weight ratios of down to approximately 1/100. The objective of the subsampling is primarily to interrogate a sufficiently large number of mixture cells such that some from the minor donor are recovered and able to be genotyped. The simplest approach would be to remove single cells as per the standard DSCS approach ,7, whichBased upon the above subsampling considerations, seventy-five five-cell subsamples were collected from the original 1:50 mixture. In accordance with the binomial sampling expectations, 6 of the 75 collected subsamples consisted of mini-mixtures, which could then be analyzed by the previously validated PG methods for single or few cells . ProbatiPG analysis of the standard bulk 1:50 mixture returned uninformative LRs  for The DSCS approach was applied to mixtures in which a minor donor was virtually undetectable by standard approaches resulting in a probative gain of DNA profile information. Collecting multiple five-cell subsamples permitted the genetic interrogation of a large number of cells (75 × 5 = 375 cells) in an attempt to detect the presence of a minor donor. This subsampling procedure deliberately created mini-mixtures containing cells from the minor donor and resulted, after PG replicate analysis, in highly probative inclusionary minor donor log(LR)s (~11). Although as a proof-of-concept study we took 75 × 5-cell subsamplings, laboratories could choose different subsampling schemes. For example, one could take 35–40 subsamples and evaluate the results to see whether a minor donor was detected (as might be expected if there was a 1:50 barely detectable minor donor mixture) then decide whether it would be necessary or useful to perform another subsampling. In our experiment, we detected the minor donor on the twelfth collected subsample , therefore 40 × 5-cell subsamples may be sufficient to obtain three replicates of the minor donor.The approach described could prove useful to not only improve the quality of results obtained from mixtures with an undetectable minor donor when both donors are male or when the minor donor is female, but also could provide additional inclusionary support for true donors in those mixtures that currently need to rely on Y-STR analysis for the detection of the person of interest. Although this proof-of-concept experiment was applied to a binary mixture, similar subsampling schemes to interrogate more complex mixtures with >2 donors (to recover probative DNA profile information from undetectable/unresolvable minor donors) can be envisioned.


The aim of this study is to validate the application of the sellar barrier concept for predicting intraoperative CSF leak in endoscopic endonasal surgery for pituitary adenomas with a machine learning approach.Recently, it was defined that the n = 11/155) and inliers into separate datasets, the latter further separated into training (n = 115/144) and inlier test (n = 29/144) datasets. The machine learning analysis was performed using different novelty detection algorithms , whose performance was assessed separately and as an ensemble on the inlier and outlier test sets.We conducted a prospective cohort study, from June 2019 to September 2020: data from 155 patients with pituitary subdiaphragmatic adenoma operated through endoscopic approach at the Division of Neurosurgery, Università degli Studi di Napoli “Federico II,” were included. Preoperative magnetic resonance images (MRI) and intraoperative findings were analyzed. After processing patient data, the experiment was conducted as a novelty detection problem, splitting outliers , MRI barrier: strong (p = 1.405 × 10−6), MRI barrier: weak (p = 4.487 × 10−8), intraoperative barrier: strong (p = 2.788 × 10−7), and intraoperative barrier: weak (p = 2.191 × 10−10). We recorded 11 cases of intraoperative leakage that occurred in the majority of patients presenting a weak sellar barrier (p = 4.487 × 10−8) at preoperative MRI. Accuracy, sensitivity, and specificity for outlier detection were 0.70, 0.64, and 0.72 for IF; 0.85, 0.45, and 1.00 for LOF; 0.83, 0.64, and 0.90 for oSVM; and 0.83, 0.55, and 0.93 for the ensemble, respectively.According to the type of sellar barrier, patients were classified into two groups, i.e., strong and weak barrier; a third category of mixed barrier was defined when a case was neither weak nor strong. Significant differences between the three datasets were found for Knosp classification score (sellar barrier at MRI and its in vivo features as observed during endoscopic endonasal surgery. The novelty detection models highlighted differences between patients who developed an intraoperative CSF leak and those who did not.There is a true correlation between the type of  Endoscopic endonasal approach, representing the most suitable technique , is indiPituitary macroadenomas present a predominantly vertical growth pattern and, albeit being round and soft, displace and compress pituitary gland tissue: the latter gets to be a part along with arachnoid and sellar diaphragm of the interface between the tumor and the supradiaphragmatic area . Accordivia standard endoscopic corridor more troublesome, leading to an increased risk of CSF leak  or significant artifacts on the images used for the analysis were excluded.Patients with intra-suprasellar subdiaphragmatic pituitary adenoma scheduled for tumor resection All patients underwent radiological preoperative assessment with a specific MRI protocol for the sellar region that included sagittal and coronal slices in T1-weighted volumetric sequences, with and without contrast; with axial and sagittal slices of the sealing region in T2-weighted, fluid-attenuated inversion recovery (FLAIR), and Echo Spin gradient sequences (1.5 and 3.0 T resonator).sellar barrier were made with the Horos for Mac-OSX . The measurements were made as explained in previous publications  approach –24. A Kaapproach .k categories to k − 1 indicator variables. The final feature set comprised the following:1. Age2. Knosp classification score3. Gender4. Status: Growth hormone (GH) secreting5. Status: GH Prl secreting6. Status: Prl secreting7. Status: Nonfunctioning8. Size: Microadenoma9. MRI barrier: Strong10. MRI barrier: Weak11. Intraoperative barrier: Strong12. Intraoperative barrier: Weak13. Presurgical treatmentGiven the distribution of classes within the datasets, the experiment was treated as a novelty detection problem. Therefore, outliers  and inliers were split into separate datasets. Then, the latter was further separated with an 80%/20% proportion into training and inlier test datasets. The only variable with missing values within all datasets was the modified Knosp classification score . An imputer based on the mode of this parameter was fit on the training set (mode = 2) and used to transform all datasets to remove the missing values. Then, continuous variables were normalized using a min–max scaler (range = 0–1), also fit exclusively on the training data and used to transform all datasets. Finally, principal component analysis was used to reduce the dimensionality of the data to two vectors, again by fitting only on the training set features and transforming all datasets.Patient data were processed based on their nature using the pandas, numpy, and scikit-learn Python packages . OrdinalThe machine learning analysis was performed using the scikit-learn Python package . DiffereAll statistical tests were conducted in R . Continuous data are presented as mean and standard deviation. Categorical and ordinal data are presented as value counts and proportions. The Shapiro–Wilk test was used to assess the normality of distribution of continuous data. Analysis of variance and Fisher exact tests were used to assess for differences in variable distribution among the training and test groups. Precision , recall , accuracy , and f-score  were calculated as accuracy metrics.One hundred and fifty-five patients were enrolled in the study . Regarding the pituitary adenomas’ features, 129 (83%) were macroadenomas and 26 (17%) were microadenomas; 81 were nonfunctioning tumors (52%), while 43 (27%) were GH secreting, 15 (9.6%) were adrenocorticotropic hormone (ACTH) producing, 12 (0.6%) were prolactinomas, and finally 3 (0.2%) were GH/prolactin (PRL)-secreting adenomas. According to Micko grading scale, i.e. modified Knosp classification .The distribution according to the sellar barrier subtype on MRI was as follows: 108 (69.7%) adenomas had a strong barrier, 13 a weak barrier (0.8%) –3, and fp = 0.0015), MRI barrier: strong (p = 1.405 × 10−6), MRI barrier: weak (p = 4.487 × 10−8), intraoperative barrier: strong (p = 2.788 × 10−7), and intraoperative barrier: Weak (p = 2.191 × 10−10). Accuracy, sensitivity, and specificity for outlier detection were 0.70, 0.64, and 0.72 for IF, 0.85, 0.45, and 1.00 for LOF; 0.83, 0.64, and 0.90 for oSVM; and 0.83, 0.55, and 0.93 for the ensemble. Confusion matrices and accuracy metrics are presented in The training dataset included 115 (74%) patients without intraoperative fistula, while the inlier and outlier test sets included 29 (19%) and 11 (7%) patients, respectively. Patient clinical and demographic data are presented in The possibility to predict outcomes is critical to ensure the highest standards of surgical care, above all to satisfy patient inquiries with regard to the pros and cons of the procedure they are about to undergo.CSF leak is one of the most threatening complications of transsphenoidal pituitary surgery, and also per its related potential complications, such as meningitis  or tensiYears of peculiar care and efforts, in terms of materials and reconstruction techniques, have been given to this issue , 29–35, It is crucial to highlight that albeit intraoperative CSF leak rates are reported as high as 10.3%–69%, postoperative CSF leak rates lower down to 1.3%–8% , 40–54.In the present case series, we found a rate of intraoperative CSF leak of 7.1% and postoperative CSF leak of 0%, which are similar to those reported in the literature for this kind of surgery. These findings suggest that the rate of patients with intraoperative CSF leak who finally developed a postoperative CSF leak is negligible; this is mostly because of the improvement of reconstruction techniques over the years and the refinement of surgical skills in skull base surgeons .However, there are only few reports concerning the possible risk factors that can be detected preoperatively to predict an intraoperative CSF leakage, but univocal consensus has not achieved yet: different groups claimed an increased risk of postoperative fistula upon the opening of the third ventricle, or in cases of patients with higher BMI –11.sellar barrier concept and its role in predicting the risk of intraoperative CSF leakage was recently introduced , MRI barrier: weak (p = 4.487 × 10−8), intraoperative barrier: strong (p = 2.788 × 10−7), and intraoperative barrier: weak (p = 2.191 × 10−10).Our ML model identified with outstanding accuracy that there is a cogent correlation between the weak barrier type and the intraoperative CSF leakage MRI barrier: strong (locus minoris resistentiae,” whose careless manipulation during tumor removal can expose the increased risk of intraoperative CSF leakage.These findings are relatively new and might provide further issue to be considered when defining the surgical planning. Hence, also per intraoperative observation, a weak barrier represents a “In the near future, this computer-aided decision-making tool might improve surgical quality by regularly identifying those patients at higher risk of developing intraoperative CSF leakage and related complications; validated machine learning tools might change routine surgical practice if properly setup: the creation of a computerized predictive algorithm can be a crucial step to further refine modern neurosurgery.Our teams are working on a risk classification of intraoperative CSF leak to refine the most appropriate surgical strategy and adequately inform the patient about its postoperative course.Claude Bernard used to say, “who doesn't know what he's looking for, doesn't understand what he finds.” The clinical application of the sellar barrier concept will allow the skull base surgery team to predict the scenario they will encounter. Thanks to this, you will be able to carefully select the method to be used in the reconstructive phase.During preoperative consultation, the use of imaging software allows the surgeon to show the sellar barrier to the patient and explain with a graphic support about their risk of CSF leakage. Thanks to this, the surgeon can speak clearly and precisely with the patient. It allows the surgeon to inform the patient about his/her possible postoperative evolution: postoperative nasal symptoms, surgery time, and the risk of postoperative CSF fistulae, among others. This type of information would have legal implications in the postoperative period –14.The concept of the sellar barrier cannot be considered as a totally independent predictor factor of CSF leakage.This is a prospective cohort study with a small series of patients. A multicenter study with more extensive patient series is required to validate this concept and its clinical applicability.Finally, it is worth reminding that the downside of the ML model can be its troublesome application to a clinical context: the interpretation of this model conceives the human–machine interaction as its highest moment.The sellar barrier is a new parameter to be considered in the risk assessment of intraoperative CSF leak. The present study demonstrates the efficacy of the sellar barrier concept in patients operated for endoscopic endonasal pituitary removal and strengthens its clinical applicability.sellar barrier at MRI and its in vivo features as observed during endoscopic endonasal surgery. The novelty detection models highlighted differences between patients who developed an intraoperative CSF leak and those who did not.There is a true correlation between the type of


PO) intended to estimate the capacitance and equivalent serial resistance of a capacitor (ESR). The implemented observer consists of a dynamic second-order discrete-time system. The input signal of the observer is the voltage at the terminals of the capacitor measured during its discharge across a variable resistance in two steps. The implemented observer can be used in quasi-online or offline mode. The theoretical and experimental supporting materials provide a comprehensive picture of the implementation and conditions of use of the PO. The experimental verification was carried out with a microcontroller with Cortex®-M7 core architecture. The sampling time of the PO was 20 μs, and the estimation of the parameters was obtained before the end of the discharge of the capacitor. In the cases described in the paper, this means approximately 25 ms. Due to the PO’s capabilities —proved by the experiments carried out on three electrolytic capacitors of 100 μF, 220 μF and 440 μF—the implementation is of interest for several applications, primarily in the field of power electronic applications.This paper describes the implementation of a parameter observer ( Electric capacitors are passive electrical devices that are indispensable for electrical circuits in electronic applications, hereinafter referred to as processes. In designing the circuits, depending on the operating regimes to which the capacitors are subjected in the processes, we use several topologies of electrical models, from the simple capacity, or capacity in series with a resistance, to models with more capacities, resistances and inductances. In this context, the parameters of the models are called equivalent parameters of a capacitor.In many processes, capacitors play key roles in filtering and storing energy, being indispensable elements. As an example, we mention the aluminum electrolytic capacitors used in power electronic converters and in the circuits of photovoltaic panels (solar PV). In general, we must refer to their applications in power electronics ,2,3. TheeC and seR of an R-C serial model of a capacitor. This does not estimate the state variables, but rather the time constants of several first-order linear circuits that appear during the two-stage discharge of the capacitor. From a dynamic point of view, the discharge corresponds to the free regime of a first-order linear system, the signal having the dynamic characteristics of the system’s impulse response. The parameter observer is a second-order dynamic system in discrete time that has a single input signal, namely, the one for which it calculates the equivalent time constant. The use of a single input signal makes it more reliable and less vulnerable to disturbances.Various principles are used for monitoring the equivalent parameters of capacitors. One of these is based on the use of observers . UsuallyZ(f), C(f) and seR(f), respectively [C(f) and seR(f) are not constant shows that the parameters of the capacitor change during dynamic operating regimes, whereby the voltage across the capacitor terminals varies in time. In this context, the “equivalent” attribute takes on an additional connotation, referring not only to the model associated with the capacitor and to its operating regime.Often, we investigate the behavior of capacitors using the frequency characteristics of impedance, capacitance and ESR, i.e., ectively ,15,16. TFrom the perspective of the evaluation of capacitors in relation to the processes in which they are involved, evaluation methods of capacitor parameters are considered to be of three types: online, off-line and quasi-online . In prinPO proposed in [PO implementation structure and to illustrate its capabilities in real cases. The novelties introduced in this paper compared to [PO on a microcontroller and its validation on electrolytic capacitors.The implementation of a eC and seR of the capacitor during the discharge process, which is about 20 ms.The real-time estimation of the values eC and seR.An improvement of the estimation method for the time-equivalent constants used to calculate This paper presents a real-time implementation of the posed in . The imppared to  are:The PO is presented synthetically, and the hardware and software aspects related to the implementation on a microcontroller are analyzed in detail. PO in the case of three different capacitors and summarizes the experimental results. PO implementation in applications. PO’s expediency.The remainder of the article is organized as follows. In 1.T(t) of the non-autonomous, first-order, unforced dynamical system (1) with properties (2) can be determined using the discrete-time parameter observer (3).The time-varying parameter In this section, we summarize under points 1 and 2 the results from , which rS and PO-T blocks in PO-T.PO will always contain a transient interval to be omitted. For this reason, only the discrete time signal 2.The main application considered in  is to deSince As shown in , the propK = 2 × 104 s−1, iK = 108 s−2 and pK and iK. The values of Thus, we obtain the variations in Under the conditions of the experiments performed in , we deteTo measure the equivalent parameters of capacitors by the method of discharging capacitors in two stages over external resistors, as presented in ,18, the measuring the value of monitoring the values of As the accuracy of calculating the values of Under the assumption that the deviations PO-T and are not used for calculating the parameters. The intervals III and V are used to calculate The application of the time diagram is carried out according to the simplified flowchart in stage is initialized with the value 1, which is maintained during the first download stage. Later, when new variables are initialized in the second stage, it takes the value 2. The parameter stage = 1 and the value stage = 2, respectively. Likewise, the parameter stage = 1 and the value stage = 2, respectively. ®-M7 of NXP core architecture [The schematic used to measure the equivalent capacitance and ESR of a capacitor is represented in itecture , which aPO-T algorithm, processes it. To reduce the influence of the input impedance of the microcontroller’s sampling circuit over the discharge process, we inserted the operational amplifier IC1 as a voltage repeater with high input impedance and low output impedance. The IC1 is powered from PO-T. For example, for calculating a logarithm of a real simple precision number (float), the execution time is about The input signal Before the beginning of the discharge, the signal P1 ensures the saturation of the bipolar transistor T1 that keeps the P-MOS transistor T2 open, and thus, the capacitor C is charged through the resistor R3 until time At the moment At the moment Note that, from the viewpoint of Equation (4), For the first stage, we chose the discharging time interval To establish the value of the sampling period, we performed, using the MATLAB environment, a Fourier analysis on the signal r 8 bits . The maxThe timing of these operations is illustrated in As a result, we developed an application program running with an oversampling period of To perform the necessary sampling and processing operations, we used the ADC.h library included in the Teensyduino add-in . CallingThe circuit in PO, experiments were carried out with three capacitors as follows:1C— 2C— 3C— To investigate the possibilities of experimentally determining the values of the parameters The study was conducted on the basis of the following scenario: for each of the three capacitors, 5 series of 20 experiments were carried out with the schematic in PO for PO for the capacitors 1C, 2C and 3C. The columns written on a white background contain four average values The main goal pursued in each individual series was the precision of the values obtained with the PO, and the values The last four columns, written on a light-blue background, refer to the last experiment in each series of 20 experiments. The values The charging/discharging characteristics To qualitatively validate the accuracy of the results in The main observations on the data in PO proposed in [PO, being a quasi-online method, can be implemented on the same processor that already serves the process. We consider that our objective has been achieved, and we acknowledge this through the explanations below.The aim of our paper was to illustrate that the posed in  can be iposed in . ConsidePO proposed in [In posed in , and we PO is described in ®-M7 architecture. The necessary sampling and processing operations were performed using the ADC.h software component included in the Teensyduino add-in library. We must mention that in this paper, we present the main elements necessary for the reproduction of the application. Note that the cost of hardware support is low. In this context for the PO, we used a The implementation of the pated in , and thePO is illustrated in PO.The actual behavior of the implemented PO for providing precise results, i.e., values of The low dispersion highlights the potential of the PO in real applications involves ensuring the quasi-online estimation framework used in this paper [The experiments described in this paper were performed on independent capacitors. This approach may also be found in other research, for instance, the work in . The useis paper . On the It should also be noted that the experiments reported in this article correspond to some discharges of capacitors during which the voltage spectrum on the capacitor is predominantly at relatively low frequencies. The frequency ranges in For reasons of safety in operation and maintenance, numerous systems require the monitoring of capacitors placed in key positions in their electrical diagrams. The parameter observer whose implementation is the subject of this paper can serve the mentioned requirement in real time. It requires ensuring the possibility of discharging the monitored capacitor over a well-known variable resistor in each of the two stages.PO-estimated values of C and ESR are consistent with the frequency characteristics measured with an LCR bridge.The implementation carried out on a common microcontroller illustrates that the method has good precision and fairly good accuracy in terms of determining the parameters of some electrolytic capacitors subjected to signals with not very high frequency spectra. The sampling time with which the voltage signal was acquired from the capacitor terminals and with which the parameter observer worked was


Is biology and knowing biological ancestral information essential to the construction of identity? Bioethicist James David Velleman believes this is the case and argues that donor gamete conception is immoral because a portion of genetic heritage will be unknown. Velleman is critical of sperm donation and the absence of a biological father in donor-assisted families. His bioethical work, specifically the 2005 article ‘Family History’, is oft-cited in articles debating the ethics surrounding gamete donations and diverse family formations. However, I wonder to what extent Velleman’s ethical stance is exhibited in contemporary culture? Velleman suggests that innate knowledge of bio-superiority helps readers and audiences appreciate the importance of biological family structures in literature and film; he says, ‘When people deny the importance of biological ties, I wonder how they can read world literature with any comprehension’ . Velleman understands the stories of Oedipus, Moses, Telemachus and Luke Skywalker as demonstrating a universal cultural comprehension that genetics are essential to identity construction. I adopt Velleman’s list of stories and ask if they really can support an antidonation sentiment and suggest that most of the stories actually support diverse family structures. By exploring the significance of story-telling in cultural understandings of family and identity, it is possible to identify the ways in which story-telling can impact how society negotiates complex issues such as assisted reproduction, donor conception and donor industry regulation. This thought was expressed in his 2005 article ‘Family History’, in which he argues that both genetic parents should raise their offspring. For Velleman, biological ties and ancestral roots are essential for identity construction; consequently, he argues that donor conception is immoral because a portion of genetic heritage will be unknown, resulting in a ‘disadvantaged’ individual.The publication of ‘Family History’ fits in with a range of work from other philosophers who also argued for the nuclear family as an ideal and relied on ancient stories to support their position see . The surThe Odyssey, 700 BCE) and Luke Skywalker —examples he calls ‘world literature’—Velleman presents us with the opportunity to explore the significance of story-telling in cultural understandings of family and identity. While Velleman’s antidonor conception stance is unconditional, his reference to literature allows us to further consider how society negotiates complex issues in ART, such as, for example, industry regulation. However, while Velleman uses these narrative examples to argue for the ethical importance of biological ties, I suggest that these texts show that diverse family formations have always existed and that nuclear family models are not necessarily enshrined as idealistic.By considering the stories of Sophocles’ Oedipus (circa 429 BCE), Moses , Telemachus (attributed to Homer’s Star Wars example as the saga—often described as a contemporary myth—stretches from 1977 to 2019 and straddles important developments within the donor conception industry including the birth of the first in vitro fertilisation (IVF) child in 1978, concerns over cloning in the 1990s and donor industry regulatory shifts in the 2000s.I will use Velleman’s examples to challenge three arguments he poses: that identity is best shaped by biology, that donor conception results in disadvantaged children and that donor-assisted families fall short of an ‘adequacy’ standard. I will mainly focus on sperm donation and the importance Velleman places on the biological father in donor-assisted families and within mythical story-telling. To do this, I will concentrate on Velleman’s ‘A long time ago in a galaxy far, far away’ .For Velleman, offspring have an ‘inborn nature’ that reflects the ‘natures’ of their biological parents. Consequently, a heterosexual couple rearing their biological progeny will be ‘naturally prepared’ for child-rearing. Furthermore, biological parents will be able to care for their child ‘with sympathetic understanding’, and they will be able to show the child ‘how to recognise and reconcile some of the qualities within itself’ 371. TheIn supporting the idealism of the nuclear family model, Velleman remarks: ‘First comes love, then comes marriage, and then the proverbial baby carriage. Well, it’s not such a ridiculous way of doing things, is it?’ . These nostalgic words from a playground nursery rhyme colloquially called ‘The Kissing Song’ are not ridiculous, but this song does present a universality of heteronormativity to the exclusion of LGBTQ+ and solo-parent families. Velleman also universalises the ‘audience’. It is notable that Velleman does not analyse these stories, but rather focuses on how the audience presumably receives and interprets the themes of family and identity. Velleman asks, if not for the importance of biological heritage:How do they (the audience) make any sense of Telemachus, who goes in search of a father he cannot remember? What do they think is the dramatic engine of the Oedipus story? When the adoptive grandson of Pharaoh says, “I have been a stranger in a strange land,” do they take him to be speaking merely as an Egyptian in the land of Midian? 369However, the stories of Oedipus, Moses and Telemachus are open to interpretation and while Velleman presents these works as quintessential examples of the importance of fathering, they can also be read as articulating the significance of non-biological connections to identity construction.On the surface, Sophocles’ Theban plays depict a man, Oedipus, searching for his biological family and the tragedy of unknown lineage resulting in unwitting incest and patricide. This reading is the one Velleman wants us to accept. However, this tale does not represent positive biological bonds and the importance of being raised by both biological parents. The patricide/incest tragedy would have been avoided if his biological parents had not attempted infanticide. The name Oedipus means ‘swollen foot’ and represents how his parents intended him to be murdered: tied to a tree by his wounded ankles and exposed to the elements 140. TheMoses is perhaps a better example. As Velleman notes, in Exodus 2.22 Moses says, ‘I have been a stranger in a strange land’ and, indeed, his heritage as a Hebrew among Egyptians defines his story. But Moses is also adopted. His adoption by the Pharaoh’s daughter sets Moses on a path that leads him to rescue the Israelites. Moses does not reunite with his birth family and his birth mother is not mentioned again. When the Lord speaks to Moses he says, ‘Who gave man a mouth, or who makes (one) dumb or deaf or seeing or blind? Is it not I, the Lord? […] I will be with your mouth, and I will instruct you what you shall speak’ (Exodus 4:11–12). The Lord here is creator and nurturer and plays a more central role than an earthly parent. In Exodus, complex issues surround the political and ancestral links to Israel; but without adoption, Moses would not have become Judaism’s pre-eminent prophet.knows how they feel, in the sense that it experiences them as leading it through a natural emotional sequence’ . However, the audiences for these stories span different times and cultures and can therefore form different inferences as to what the stories convey.The story of Telemachus is more fitting for Velleman because it details a son earnestly waiting for his father to return from a long quest. However, all three ancient texts deal with illegitimacy, adoption and biological parentage in diverse ways as these mythical characters are from opposing philosophical traditions; to suggest that biology provides centrality to identity is a low-resolution way of understanding the historical and sociopolitical contexts of these stories. Mythical form contributes to an idea of what family can mean, and we can use legends and fiction to explore family formations and identity while still appreciating that they do not promise to reveal a universal ‘truth’. In Velleman’s ‘Narrative Explanation’, an article on story-telling, he argues that an audience ‘understands the narrated events, first, because it gnêsioi), conceived illegitimately (nothos) and adopted (poiêtos), with priority given to healthy blood offspring . All these problems are more disruptive than Telemachus’ so-called identity issues.In their study of ancient family formations of the Mediterranean, Oedipus Rex (the first of his three Theban plays), Oedipus survives his abandonment because he is adopted. In Exodus, the Pharaoh’s daughter sees beyond biological roots to raise a Hebrew child as her own and, in doing so, paves the way for him to become a prophet who leads the people of Israel to freedom. By choosing these examples, Velleman conveniently overlooks the numerous stories that document inadequate parenting by biological fathers. Greek mythology (from which Velleman retrieves the story of Telemachus and Oedipus) has many tales of unacceptable child-rearing by biological parents. One example is Agamemnon who executes his daughter to appease Artemis and quench his army’s bloodlust . The Torah (from which Velleman cites the story of Moses) also contains examples of questionable child-rearing; one is the story of Abraham whose faith in God is more important than his responsibility as a father. While Moses feels like ‘a stranger in a strange land’, I wonder how Isaac, the son of Abraham, feels in Genesis 22 when his father—on the orders of the Lord—ties him to a sacrificial altar and prepares to slaughter him with his dagger.In fact, in Velleman’s examples, offspring would have been at a greater disadvantage—some dead—had they not found alternative families. In Sophocles’ ‘Friendship shows us who we really are’ .Star Wars, an epic space-opera franchise created by George Lucas, is Velleman’s most intriguing case study. I will spend more time unpacking this example because it speaks more directly to the question of donor conception and assisted reproduction. Star Wars looks back to the past and mythical formation but also considers its own time and changing sociological notions of how families are formed. Let us look at Velleman’s reference to Star Wars in full:When people deny the importance of biological ties, I wonder how they can read world literature with any comprehension. […] How can they even understand the colloquy between Darth Vader and Luke Skywalker? Surely, the revelation “I am your father” should strike them as a bit of dramatic stupidity — a remark to be answered “So what?” 2005, 369).69.3The revelation is not ‘dramatic stupidity’; regardless, I do think that ‘So what?’ is an appropriate response to its context within the saga. If we trace the plot—which unfolds over the franchise’s trilogy of trilogies—we see a rejection of blood and the celebration of friendship.The Empire Strikes Back  and the Sith Lord, Vader.kes Back  the reacStar Wars champions the bond forged between Luke and a family that is chosen rather than born. The non-biological relationships between Luke, Han Solo, Obi-Wan Kenobi, Yoda, R2-D2, C-3PO and Chewbacca are the most significant counters to the threat of the dark-side and provide Luke with the support and strength he needs to fight the Empire.Even though—as film critic Andrew Lewis Conn notes—the revelation of Luke’s parentage was shocking to the 1980s audience (a moment which ‘caused a collective frisson’), we—like Luke—need to say, ‘So what?’ and acknowledge the complexity of Luke beyond his genetic line Star Wars is not the story of Luke discovering his roots, but of Luke assisting Anakin’s rebirth. Jedi Master Yoda describes the biological connection between Luke and Vader in Star Wars: Episode VI Return of the Jedi (1983) as a burden because the revelation of parentage has more meaning to Anakin’s journey than Luke’s. There is significance in Charles Taliaferro and Annika Beck’s claim that Vader never actually takes on a fatherly role with Luke despite their genetic link: ‘Luke seems to be acting more like the father than Vader, as Luke tries to turn him back to “the good side,” as if the villain is a lost child.’ As Taliaferro and Beck note, Anakin/Vader as a father operates as a ‘toxic caricature of love’ and ‘a painful portrait of how fatherhood can go wrong’ . ConStar Wars trilogy (Star Wars: Episode VII The Force Awakens (Star Wars: Episode VIII The Last Jedi (2017), and Star Wars: Episode IX The Rise of Skywalker (2019)), sees the introduction of Ben Solo (Kylo Ren), who is the son of Luke’s sister Leia and Luke’s friend Han Solo. Ben is born with the Skywalker predisposition to the Force , which negatively affects his childhood. The weight of identity expectations and biological lineage damages him and he struggles to live up to his namesake ‘Ben’ Kenobi, his mother the Resistance leader, his hero father, his uncle (Luke) and his grandfather . Tormented by the pull to both the light and dark sides of the Force and unable to navigate his complex biological heritage, Ben embraces the identity of Kylo Ren and turns against his family, even murdering his father.So far, I have focused on Velleman’s example: one isolated moment in the second film in the franchise. Now, I want to explore how identity is addressed in the later films because, although they do not figure for Velleman, in the later examples we can see changing perspectives on the family which coincide historically with developments in the field of assisted reproduction. This symbiotic relationship helps us think about how we understand family stories related to donor conception. Awakens , Star WaThe Last Jedi, Supreme Leader Snoke states that when he first encountered the boy, he sensed ‘the potential of your bloodline; a new Vader’, then moments later mocks him by saying, ‘you have too much of your father’s heart in you’ , the Sith Emperor. This revelation is jarring but does not sway her to darkness. She rejects what is presented as her ‘blood right’ to become Empress and instead fights for the Resistance. In a significant scene, Kylo Ren and Rey both try to wield Anakin Skywalker’s lightsabre—a weapon to which Ren believes he has a birthright as Anakin’s grandson. Yet, the weapon favours Rey and is wielded by her. The weapon does not favour the biological Skywalker line, and this exchange symbolically highlights the fallacy of presenting ‘blood’ as superior to any other sort of connection.In the final moments of the last film, a stranger asks Rey for her name and she announces that it is Skywalker. Despite Palpatine’s blood running through her, she adopts the name and heritage that fits her blossoming identity. By the end of the film, the only person who bears the Skywalker name is not a biological descendant. Biology is not all important and consequently, when Palpatine tells Rey ‘long have I waited, for my grandchild to come home!’, the revelation is met with dismissal because Rey is a Skywalker: it is something she is emotionally and psychologically, not something she has in her veins. Luke makes this point himself when he tells her that ‘some things are stronger than blood’ . are not predetermined by “destiny” but are defined and redefined by difficult and continually recurring moral choices’ For Velleman, donor conception is immoral because it produces a disadvantaged child who does not know their biological roots. Velleman equates donor-conceived babies with babies who experience gestational complications, and he suggests that donor-conceived people are ‘handicapped’ 372. VelStar Wars, many characters are unaware of their genetic history for a multitude of reasons: taken as children to be trained by the Jedi (Qui-Gon and Obi-Wan), abandoned by irresponsible parents (Han), orphaned in the galactic war (Jyn Erso), cloned (Boba Fett), misled about their genetic history  or produced through fatherless donor conception (Anakin). Star Wars is populated by heroes who have little or no access to their full ancestral history. This detail reflects those in society who do not have full awareness of their bloodline for various reasons, including abandonment, estrangement, adoption, parental death, family breakdown, domestic violence, relocation, donor conception, displacement, migration and incomplete ancestral history. Coming from a family of academics and teachers has enabled Velleman to imagine that he follows in his ancestors’ footsteps. By leaning heavily on his family history when considering his identity, Velleman glorifies detailed awareness of lineage to which many do not have access. Further, Velleman’s assertion that identity is shaped by ancestral knowledge prioritises nuclear family models as comprising of ‘full’ identity, while other models engender suboptimal partial ones.However, in Beyond Price, Velleman speaks of a ‘standard of adequacy’ to justify procreation , which involve the sudden loss of a biologically linked patriarchal figure expected to assume legal and social responsibility. The sperm provider is not expected, nor obliged, to occupy the role of the father; instead, fathering is assumed by a male raising the child, or non-existent in the case of lesbian couples or solo mothers who form fatherless families. The complexity of fatherlessness in sperm donation narratives is not compatible with the father-absence narratives Velleman offers in his articles.Offspring, Barry Velleman and I also read the donor conception industry very differently. Partly, this is symptomatic of the times about which we are writing. Velleman (despite more recent pieces) is writing about sperm donation within the American model of donor conception pre-2005, which was plagued by unregulated practice, donors with complete anonymity and children raised with no knowledge of their conception history. In Beyond Price, 2013), it is important to note how industry regulation continues to differ across the globe, and in the USA, from state to state. While Velleman refers only to the US model, I also invoke the UK model which, in terms of regulation, has modelled tighter practices since 1991. Since Velleman’s first article on donor conception in 2005, guidance in the USA and the UK has become clearer. For example, the US Food and Drug Administration (FDA), Human Fertilization and Embryo Authority (HFEA), DCN, Donor Sibling Registry (DSR), and American Society for Reproductive Medicine (ASRM), etc advise against the scenarios Velleman articulates as characteristic of the industry. The UK and US industries recommend that prospective parents select a registered donor from a regulated bank with donor-identity release. In contrast to Velleman’s depiction of the industry, the ASRM advises that parents should raise donor-conceived people from infancy with knowledge of their donor conception, parents should select a traceable donor, and the parents should obtain as much data as possible on the donor  accepts only 1% of donor applicants, and competitor Xytex accepts less than 3%. Following the ‘donor eligibility final rule’ enforced in 2005 by the US’s FDA, donor samples are screened for ‘clinical evidence of relevant communicable disease agents and diseases’ . Since tTelling and Talking series includes guides, booklets and pamphlets aimed at sharing the donor story with friends and families, and their Our Story series is aimed at children and depicts different types of conception journeys . While Velleman suggests donor-conceived offspring are lacking narrative structures, there is increasing awareness that narrativising the donor conception journey is crucial to helping children understand themselves and articulate their unique contexts. These children do not lack a narrative; they have a different narrative and the significance of this is addressed within these emergent modes of story-telling.Shifts in regulation, the importance of seeking known donors (or ID release donors), and the recommendation for complete transparency with offspring from an early age have culminated in increased attention being given to how donor conception is narrativised and the importance of story-telling to include new family formations. The UK’s DCN states: ‘The DCN takes a clear stance that supports openness in families and thus the majority of members are parents who have undertaken to tell their children, often from a very early stage in their development’ . By prioritising the act of story-telling in families created through technologised reproduction and donor-assisted conception, the DCN hopes to not only diversify how conception is taught to young people but to normalise this diversity. Their et al) and those who are raised without knowledge of their conception history, without access to data and without the option of donor release . When in 2011 bioethicist Margaret Somerville drew on Velleman’s ‘Family History’ to agree that donor conception is harmful and therefore against the Hippocratic oath, she was only considering examples of bad practice. In her paper she refers to two antidonation case studies by adult donor-offspring, extrapolating from those the ‘fact’ that ‘most don’t even know they’re donor-conceived’ , Damien Riggs argues that the term ‘reproduction’ needs to be defined beyond genetic connections:While Velleman does not engage with regulatory change, the regulations introduced in the UK have encouraged a general shift in cultural attitudes towards gamete conception and related ideas of what constitutes a parent and family. Respected groups like the Nuffield Council on Bioethics (NCB) have attempted to redefine contemporary definitions of terms like ‘kin’ and ‘reproduction’ for use beyond the heteronormative nuclear family. For example, the NCB suggests that the nuances of donor conception require a new way of defining terms like donor and father, which are conflated in Velleman’s work: ‘The “recipient” parent or parents will be the child’s real parents from the beginning […] the law makes provision for the donor to be excluded from the legal status of parent’ . I agree with Booy that the 1970s was a ‘polarising’ time, and this polarity is also reflected in how the ART industry was being perceived following the birth of Louise Brown—the first IVF baby in 1978. Susan Golombok notes that this decade was a pivotal time in Western culture for how families were perceived: ‘Changes to the structure of the family have been taking place since the 1970s’, and these changes coincided with changing attitudes towards people identifying as LGBTQ+ and the growing gay rights and women’s liberation movements (Return of the Jedi). The growth of the industry, as evidenced by the increase in fertility clinics in the 1980s and 1990s, coincided with a torrent of breakthroughs in ART (such as greater success with frozen gametes and embryo transfers) and cultural shifts that welcomed treatment for same-sex couples, co-parents, solo people and unmarried couples.The first film of the saga was released in the 1970s; a decade marked by crisis and hope. Miles Booy traces how the predicaments of the 1970s  informed the making of ovements , 2–3. ThStar Wars franchise, which centred on the secretive birth of the Skywalker twins who were raised without knowledge of their parentage. The twins’ birth featured in the 2005 film, Revenge of the Sith, the same year that the UK introduced compulsory identity release for all gamete donors and banned completely anonymous donations. ART development after 1970 spanned the three Star Wars trilogies and ran in parallel to the saga’s complex treatments of solo parenting, anonymous parentage, donor conception, bioengineering and the welcoming of diverse family formations.By the end of the 1990s, Vader was dead, Anakin Skywalker was born fatherless to a solo mother, and many countries had prohibited completely anonymous gamete donations. The emergence of debate around the problem of anonymity coincided with the development of the Star Wars positions the franchise as not just a work that draws on myth and can be described as mythical, but a work that is itself a cultural artefact, then we should consider how the franchise coincides with ART development as an ‘ideological investment’ (When Vader announces ‘I am your father’ in the 1980 film he signifies the older tradition of glorifying genetic parenthood and blood lineage (that is still with us); however, Luke in the 2019 film signifies changing times by passing the family mantle to Rey, to whom he has no genetic ties. From Shmi’s solo motherhood to fatherless Anakin, to the creation of a clone army, the franchise has responded to the bioethical debate which coincided with its development. If, as estment’ 3.Star Wars responds to the ART industry is complex, perhaps fittingly given that Lucas used ART to have his own child via surrogacy. The control the Empire has over clones may, on the surface, suggest the demonisation of reproductive technology. Star Wars does not present a ‘black and white’ thesis on reproduction but instead ambivalently explores how technology can be both positively and problematically used. Star Wars presents a middle ground that Velleman’s article overlooks, but one that I and many advocates of donor conception welcome: the need for industry regulation to prevent the abuse of ART without imposing such extreme regulation that diverse family formations are oppressed. To explore this ‘middle ground’, I will look at how reproductive technologies are presented in the saga.How Attack of the Clones  articulate anxieties about the increased probability of multiples  following stimulated intrauterine insemination and multiple embryo transfer in IVF. In fact, the link between multiple pregnancies and ART led to the introduction of the One at a Time campaign by the HFEA in 2007 to limit multiple pregnancies following a spike in the 1990s and 2000s. So, on the surface, the clone trope may suggest criticism and concern over ART.In e Clones , Obi WanStar Wars coincided not only with the ART revolution but also with its regulation and the debates surrounding the ethics of donor conception.However, clones do not represent an argument against reproductive technologies and donor conception wholesale but instead represent the urgent need for industry regulation. If anything, Jango’s donor storyline warns against unregulated practices and reflects the ‘Wild West’ nature of American ART in the twentieth century, in which one unregistered donor could anonymously sire hundreds of children. Simply put, Jango would have been able to create a baby army in 2002 in America but not in 2005 in the UK. Attack of the Clones. It is also easy to see how we could misread the saga as advocating for exhaustive regulation. However, while regulation is important (no one wants a million Jango Fetts), the extreme of the Empire’s rule is what happens when regulation is not in the interests of people and does not allow for the formation of diverse family structures.The Empire’s creation of clones, and the monstrous visages of their principal characters  have obvious parallels to societal concern over the use of ART technologies to create hybrids, chimeras and genetically engineered super-people. Concern over irresponsible genetic engineering was a heated topic in the mass media in the 1990s following the ground-breaking news that The Roslin Institute had successfully cloned a sheep, Dolly. A year later, in 1997, Stuart Newman, a member of the Council for Responsible Genetics, alongside biotechnology critic Jeremy Rifkin, attempted to patent the speculative creation of an animal/human hybrid known as Humouse. Their ambition was not to put the patent to use themselves; instead they wanted to prevent others from creating such ‘monsters’ by reserving the technology as intellectual property. We can easily see context that informed the 2002 film Star Wars trilogy were shot in Tunisia—notably, scenes for the Skywalker home planet of Tatooine. In Tunisia, fertility treatment for solo mothers and same-sex couples is illegal. In the 1970s, when the first film was released, Tunisia did not have a licensing body and had only a few clinics in comparison to the exponential growth of the industry in America. However, even now in 2022 Tunisia, ART is restricted to married couples due to religious and cultural beliefs. On Tatooine (described as outside the reach of the Republic), the tyrannical rule of the Empire is felt, and under this control communities are marginalised, families are broken apart and populations are wiped out. The Empire represents a new type of life under the power of the Sith: one of struggle, restriction and the complete loss of freedom. This is what discriminatory regulation looks like.The Empire represents the potential recklessness of reproductive technologies through the construction of a clone army, yet the Empire’s tyrannical control over the population of the galaxy marks the danger of excessive regulation. The Empire presents two extremes at once: irresponsible reproductive science and discriminatory regulation. Many scenes in the first Star Wars presents the audience with a ‘middle-ground’ in which diverse families are welcomed but non-discriminatory regulation of reproductive technology is recommended. This is similar to the ‘middle-ground’ the FDA, HFEA, DCN, DSR and ASRM advocate. The extreme Velleman presents is incompatible with the pro-diversity message Star Wars enshrines. As Obi-Wan warns Anakin, who is falling into the darkness due to his inability to compromise and empathise with the views of others: ‘Only a Sith deals in absolutes’  that Rey was able to choose a family rather than be confined to her genetic one. solutes’ .‘Your focus determines your reality’ .‘Always remember’, Qui-Gon advises a young Anakin Skywalker, ‘your focus determines your reality’ . A prophStar Wars, should propel us towards a deeper appreciation of how our cultural conceptions of family are expanding.In this article, I have used the same texts to suggest another position. We both could be said to be guilty of indulging what Velleman describes in ‘Narrative Explanation’ as ‘the confusion between emotional and intellectual instances of “Aha!”’ when reading mythical stories 22. In hChoosing to be open about donor conception: the experiences of parents advises early and regular ‘telling’ (The Pea That Was Me by Kimberly Kluger-Bell (2012) and Our Story by the DCN (2018) encourage the normalisation of donor conception to children. The dominant cultural narrative of the nuclear family ideal is shifting, and this shift is nicely articulated in Hoffman and Asquith’s, The Great Big Book of Families, which opens with an idealised picture of a nuclear family consisting of a father, a mother and two children; surrounding the picture are the words: ‘Once upon a time most families in books looked like this. But in real life, families come in all sorts of shapes and sizes’ , but a question about the importance of story-telling to our understandings of family and identity. Velleman’s reliance on Oedipus, Moses, Telemachus and Skywalker—and indeed his other philosophical work on narratives—suggests that story-telling is vital to our interpretation of selfhood. Therefore, it is incoherent to overlook the significance of donor conception story-telling and how these stories also articulate contemporary conceptions of what constitutes a family. Looking at ART in contemporary story-telling through memoirs, blogs, podcasts and interviews must inform how bioethicists approach the donor conception debate. For as the NCB argue: ‘Donor conception is first and foremost about people […] any debate about the ethical considerations that should inform public policy on donor conception should start, not with the analysis of abstract principles, but with the people concerned, and the reality of their lives’  will be increasingly marginalised while the heteronormative nuclear family model is further privileged. I am also concerned about the misrepresentation of cultural artefacts like literature, myth and film as historically idealising the nuclear family as this is not a transparent way of presenting the complexity of family models that have existed since antiquity. Velleman’s suggestion that donor-conceived offspring have a fractured narrative does not appreciate the diversification of narratives for different families. The question should not be about the significance of biology to identity construction but about the importance of how we talk about different types of origin stories. Unwillingness to enfold diverse conception narratives into the story of reproduction is what will disadvantage children, no matter how they are conceived, born and raised.