Combination atezolizumab/bevacizumab is the gold standard for first‐line treatment of unresectable hepatocellular carcinoma (HCC). Our study investigated the efficacy and safety of combination therapy in older patients with HCC.191 consecutive patients from eight centres receiving atezolizumab and bevacizumab were included. Overall survival (OS), progression‐free survival (PFS), overall response rate (ORR) and disease control rate (DCR) defined by RECIST v1.1 were measured in older (age ≥ 65 years) and younger (age < 65 years) age patients. Treatment‐related adverse events (trAEs) were evaluated.n = 116) had higher rates of non‐alcoholic fatty liver disease , presenting with smaller tumours  with less portal vein thrombosis , with fewer patients presenting with BCLC‐C stage disease . There was no significant difference in OS  and PFS  between older age and younger age. Older patients had similar ORR  and DCR  compared to younger patients. Atezolizumab‐related  and bevacizumab‐related  trAEs were comparable between groups. Rates of grade ≥3 trAEs and toxicity‐related treatment discontinuation were similar between older and younger age patients. Patients 75 years and older had similar survival and safety outcomes compared to younger patients.The elderly (Atezolizumab and bevacizumab therapy is associated with comparable efficacy and tolerability in older age patients with unresectable HCC. AASLDAmerican Association for the Study of Liver DiseasesAFPalpha‐fetoproteinBCLCBarcelona Clinic Liver CancerCRcomplete responseCIconfidence intervalCTCAENational Cancer Institute Common Terminology Criteria for Adverse EventsDCRdisease control rateECOG‐PSEastern Cooperative Oncology Group Performance StatusHCChepatocellular carcinomaHRhazard ratioORRoverall response rateOSoverall survivalPD‐L1programmed death ligandPFSprogression‐free survivalPRpartial responsePDprogressive diseaseMTDmaximal tumour diameterMVImacrovascular invasionNAFLDnon‐alcoholic fatty liver diseaseNSCLCnon‐small cell lung cancerSDstable diseaseSmPCsummary of product characteristicsTACEtrans‐arterial chemoembolizationtrAEtreated‐related adverse eventVEGFvascular endothelial growth factorAtezolizumab and bevacizumab are effective cancer treatments for patients with liver cancer. However, its effectiveness and safety in older patients with liver cancer are not as clear. Our study demonstrates this treatment is effective in older patients, without an increase in side effects.1Hepatocellular carcinoma (HCC) is the fourth leading cause of cancer mortality and the sixth most prevalent worldwide.Increasing age is an established risk factor for HCC.The impact of immunotherapy in older age patients is less well studied than in younger cohorts, due to an underrepresentation in cancer trials.Our retrospective study aimed to evaluate the real‐world efficacy and safety of combination atezolizumab and bevacizumab in older age patients for unresectable HCC.22.1This was a multi‐centre retrospective cohort study. Study participants were consecutive patients with unresectable HCC receiving atezolizumab plus bevacizumab across eight tertiary centres in Germany, Japan, Austria, United Kingdom, Italy, Taiwan and United States of America. All patients had a histological or radiological diagnosis of HCC in accordance with American Association for the Study of Liver Diseases (AASLD) criteria2.2Patients' baseline demographics and clinical parameters including underlying liver disease aetiology, Child–Pugh class, BCLC stage, Eastern Cooperative Oncology Group Performance Status (ECOG‐PS) were collected. Overall survival (OS) was defined as the time in months from first drug administration to date of death or date of last follow‐up. Profession‐free survival (PFS) was the time from first drug administration to date of progression on radiological imaging or death whichever came first. Treatment response was evaluated using RECIST criteria v1.1 on CT or MRI at 9–12 week intervals.2.32 test for categorical data, and the unpaired Student's t‐test for continuous data. The proportion of emerging TrAEs and ORR/DCR were compared between age cohorts using the χ2 test.For age analysis, patients 65 years or older were classified as older age as per classification from United Nationsp value of less than .05 was defined as statistically significant. Univariate and multivariate Cox regression models for older age and established prognostic factors were conducted for OS and PFS.Time‐to‐event analysis was performed for OS and PFS using the Kaplan–Meier method. OS and PFS were compared between age cohorts using log rank. A Due to the increasing age and life expectancy of patients, survival and safety analysis was performed using patients 75 years and older and patients under 75 years old.2.4This study received ethical approval by Imperial College Tissue Bank (Reference Number R16008) and by the local ethics committee of each treating centre.33.1There were 210 patients receiving atezolizumab and bevacizumab from the 12 centres between January 2020 and December 2021. Nineteen patients had received prior systemic therapy and were excluded from the study. The baseline characteristics of the 191 consecutive patients receiving atezolizumab plus bevacizumab are shown in Table p < .001), with a lower rate of chronic Hepatitis B infection . Older patients had smaller tumours , an inferior proportion of macrovascular invasion (MVI)  and were more likely to commence treatment with BCLC‐B stage disease . Older patients had similar rates of ascites  and varices , but lower rates of hepatic encephalopathy . Rates of Child–Pugh B cirrhosis were similar between the age groups .One hundred and sixteen patients (60.7%) were 65 years or older when receiving the first dose of atezolizumab plus bevacizumab, and 75 (39.3%) were less than 65 years old. The older cohort had a higher proportion of non‐alcoholic fatty liver disease (NAFLD) . At the time of analysis 62 (32.5%) patients had died. Survival was comparable between the age cohorts with older age having a median survival of 14.9 months compared to 15.1 months (p = .67) for those <65 years  and multivariate  analysis (Table p = .69)  (Table p = .27) and DCR  compared to younger patients (Table The median duration of treatment with atezolizumab and bevacizumab was 3.5 months (interquartile range [IQR] 1.5–7.7 months). Treatment duration was comparable between the older and younger groups (3.4 (IQR 1.4–8.1) months vs. 3.5 (IQR 1.6–6.6) months; 2) Table . Diseasep = .80) and PFS  in patients with Child–Pugh B liver disease. Similar comparable survival outcomes were seen in patients with Child–Pugh A disease for older patients.As Child–Pugh class had a significant effect on OS and PFS in multivariate survival analysis, we performed subgroup analysis in patients by Child–Pugh class. Older patients had comparable OS  (Table p = .91). Rates of atezolizumab‐related AEs were comparable between the two groups, the most common being fatigue in both age cohorts . Bevacizumab‐related trAEs were similar between the two groups . No difference was noted in rates of treatment discontinuation with seven patients 65 years and older and five patients under 65 discontinuing treatment due to trAEs . There was no difference in grade 3 or higher atezolizumab‐ and bevacizumab‐related trAEs between age.There was no difference in trAEs of any grade between older and younger age patients  Table . There w3.43.4.1p = .90)   Figure  and medi) Figure , with no) Figure . Age 75 ) Figure  and S4. ) Figure .4The landmark IMbrave150 study resulted in a step change in the management of advanced HCC. Combination atezolizumab/bevacizumab is now a recommended first‐line therapy in patients who have no contraindication to either drug. Recent results from the HIMALAYA trial demonstrate tremelimumab and durvalumab superiority to monotherapy in unresectable HCC, further demonstrating the efficacy of combination immunotherapy.A recent subgroup analysis of the IMbrave150 investigated the efficacy and safety of atezolizumab/bevacizumab in unresectable HCC patients aged 65 years and older.p < .001) compared to systemic chemotherapy.Two large meta‐analyses have reviewed the efficacy of immunotherapy in older patients with non‐HCC cancer types. Elias et al. reviewed 17 randomized controlled trials using nivolumab, pembrolizumab and atezolizumab.In our study, there was a higher proportion of underlying NAFLD in the elderly cohort, and a lower proportion of Hepatitis C compared to the younger age group. A recent meta‐analysis showed that non‐viral HCC may not fully benefit from improved survival after immunotherapy,A concern for the use of cancer therapy in older age patient is significant toxicity. Systemic chemotherapy has been associated with higher rates of trAEs,The relationship between ageing, cancer and immunity is complex. Immune function has been shown to decline with increasing ageOur study has some limitations. This is a retrospective study, and therefore subject to collection and selection bias. Across eight tertiary centres, there may be inter‐site variation in treatment protocols, follow‐up, efficacy and safety assessments. We chose an age of 65 years as a cut‐off between the two groups. This was based on boundaries used in previous large meta‐analyses,5Our study shows atezolizumab and bevacizumab therapy to be efficacious and safe to use in older patients with unresectable HCC. Clinicians should not be deterred in administering combination therapy in older patients, provided eligibility for combination immunotherapy is otherwise met.All authors consented to the publication of the manuscript.MV is supported by the National Institute of Health Research. AD is supported by the NIHR Imperial BRC and by grant funding from the European Association for the Study of the Liver (Andrew Burroughs Fellowship) and from Cancer Research UK (RCCPDB‐Nov21/100008). DJP is supported by grant funding from the Wellcome Trust Strategic Fund (PS3416) and from the Associazione Italiana per la Ricerca sul Cancro (AIRC MFAG Grant ID 25697). We acknowledge support from the NIHR Imperial Biomedical Research Centre (BRC), the Imperial Experimental Cancer Medicine Centre (ECMC) and the Imperial College Tissue Bank.AD received educational support for congress attendance from Roche. JvF received advisory board fees from Roche. HW received lecture fees and advisory board honoraria from Roche, Bayer, Ipsen, Eisai, BMS. AS received research grants (to institution) from AstraZeneca, Merck, Bristol Myers Squibb, Exelixis, Clovis, KAHR medical, Actuate therapeutics, Incyte Corp. and Advisory board fees from AstraZeneca, Bristol Myers Squibb, Merck, Exelixis and Pfizer. PRG reports a consulting or advisory role and received honoraria from AdaptImmune, AstraZeneca, Bayer, Bristol Myers Squibb, Eisai, Ipsen, Lilly, Merck Sharp & Dohme, Roche and Sirtex; has been on a speakers bureau for straZeneca, Bayer, Bristol Myers Squibb, Eisai, Ipsen, Lilly, Merck Sharp & Dohme, Roche and Sirtex; has received research funding from Bayer and Roche; has provided expert testimony for Lilly; and has received travel or accommodation expenses from AstraZeneca, Bayer, Bristol Myers Squibb, Eisai, Ipsen, Lilly and Roche. DB has received lecture and speaker fees from Bayer Healthcare, the Falk Foundation Germany and consulting fees from Boston Scientific. AV reports honoraria for speaker, consultancy and advisory role from Roche, AstraZeneca, EISAI, Bayer, Merck, Bristol Myers Squibb, Merck Sharp & Dohme, Incyte, PierreFabre, Ipsen and Sanofi. BS received travel support from Gilead, Ipsen and AbbVie. NP received consulting fees from Amgen, Merck Serono, Servier; lectures fees from AbbVie, Gilead, Lilly, Sanofi; travel expenses from Amgen, ArQule; and institutional research funding from Basilea, Merck Serono, Servier. TP received consulting fees from Bayer; and institutional research funding from Bayer, Lilly, Roche. RS received consulting fees for EISAI, Roche, Bayer, SIRTEX, Novartis; research funding (to institution) from Incyte, Novartis, Astex Pharmaceuticals, Bayer and Boston Scientific. MP is an investigator for Bayer, BMS, Ipsen, Lilly and Roche; he received speaker honoraria from Bayer, BMS, Eisai, Lilly, MSD and Roche; he is a consultant for Bayer, BMS, Eisai, Ipsen, Lilly, MSD and Roche; he received travel support from Bayer and BMS. AC received consulting fees from MSD, BMS, AstraZeneca, Roche; speakers' fee from AstraZeneca, MSD, Novartis and Astellas. LR received consulting fees from Amgen, ArQule, AstraZeneca, Basilea, Bayer, BMS, Celgene, Eisai, Exelixis, Genenta, Hengrui, Incyte, Ipsen, IQVIA, Lilly, MSD, Nerviano Medical Sciences, Roche, Sanofi, Servier, Taiho Oncology, Zymeworks; lecture fees from AbbVie, Amgen, Bayer, Eisai, Gilead, Incyte, Ipsen, Lilly, Merck Serono, Roche, Sanofi; travel expenses from Ipsen and AstraZeneca; and institutional research funding from Agios, ARMO BioSciences, AstraZeneca, BeiGene, Eisai, Exelixis, Fibrogen, Incyte, Ipsen, Lilly, MSD, Nerviano Medical Sciences, Roche, Zymeworks. DJP received lecture fees from ViiV Healthcare, Bayer Healthcare, BMS, Roche, Eisai, Falk Foundation, travel expenses from BMS and Bayer Healthcare; consulting fees for Mina Therapeutics, EISAI, Roche, DaVolterra, Mursla, Exact Sciences and Astra Zeneca; research funding (to institution) from MSD and BMS. All remaining authors have declared no conflicts of interest. The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed. No writing assistance was utilized in the production of this manuscript.Appendix S1 Supporting informationClick here for additional data file.


Type 2 diabetes (T2DM) is an established risk factor for developing Parkinson's disease (PD), but its effect on disease progression is not well understood.The aim of this study was to investigate the influence of T2DM on aspects of disease progression in PD.We analyzed data from the Tracking Parkinson's study to examine the effects of comorbid T2DM on PD progression and quality of life by comparing symptom severity scores assessing a range of motor and nonmotor symptoms.P = 0.002), and nonmotor symptoms, as assessed by Non‐Motor Symptoms Scale total (38.4 [2.5] vs. 31.8 [0.7] P < 0.001), and were significantly more likely to report loss of independence  and depression . Furthermore, over time, patients with T2DM had significantly faster motor symptom progression (P = 0.012), developed worse mood symptoms (P = 0.041), and were more likely to develop substantial gait impairment  and mild cognitive impairment  compared with the PD group.We identified 167 (8.7%) patients with PD and T2DM (PD + T2DM) and 1763 (91.3%) patients with PD without T2DM (PD). After controlling for confounders, patients with T2DM had more severe motor symptoms, as assessed by Movement Disorder Society Unified Parkinson's Disease Rating Scale, Part III  is associated with more severe Parkinson's disease (PD), affecting motor symptoms, nonmotor symptoms, and cognition. Patients with T2DM develop substantial gait impairment and mild cognitive impairment faster than those without T2DM. Targeting insulin resistance in PD may offer a therapeutic window in which to halt further PD progression. Parkinson's disease (PD) affects 6 million people worldwide, and its prevalence is expected to increase in response to an aging population.Beyond its effects on PD risk, a handful of studies have evaluated the influence of T2DM on disease progression and suggested comorbid T2DM may be associated with faster motorUsing the Tracking Parkinson's cohort, a large long‐term observational study into PD, our aims were, first, to evaluate the association of comorbid T2DM on PD severity in patients recently diagnosed with PD and, second, to determine whether T2DM negatively affects disease progression. In view of accumulating data suggesting antiglycemic treatments may be useful in the treatment of PD, we also conducted an exploratory analysis for our third aim: to determine whether metformin use in patients with diabetes confers any protective effects on the severity and long‐term outcomes.Data from Tracking Parkinson's, including demographic, clinical, imaging, and biospecimen measures, that have been collected for more than 6 years were analyzed. The study setup and design have been previously reported.Due to previous evidence that insulin resistance may impact several aspects of PD progression via multiple molecular mechanisms, we have chosen a number of motor and nonmotor outcomes that could highlight the influence of T2DM on PD. We used several motor and nonmotor features previously established as “severe/advanced” disease markers or clinical milestones,Appearances of motor fluctuations, dyskinesias, impulse control disorders (ICDs), dopamine dysregulation, and hallucinations were regarded as binary endpoints, corresponding to Movement Disorder Society Unified Parkinson's Disease Rating Scale (MDS‐UPDRS) item responses of ≥1.Group comparisons between PD and PD + T2DM groups were performed at baseline, using multivariate analysis of covariance with post hoc Bonferroni correction for multiple comparisons. Categorical variables were compared using Fisher's exact test, and multivariate logistic regression was used to determine the adjusted odds ratios (ORs). Potential confounders were included in the statistical models guided by mechanisms proposed and depicted in directed acyclic graphs, which considers each variable in relation to the exposure and outcome  were excluded from the model.P values presented have been adjusted for multiple comparisons.To assess the effect of T2DM on rate of change of a given symptom , we used separate linear mixed effects models with robust variance estimates, with examination of the interaction effects of group (PD vs. T2DM) and time. Symptom progression was modeled adjusting for age, sex, vascular score, disease duration, ethnicity, baseline LEDD, and the baseline variable value as fixed effects for the intercept and slope. Participant‐specific random effects were included as both a random intercept and a random slope to account for the correlation in repeated measurements within the same participant. Due to the increasing number of patient dropouts over the follow‐up period, we chose a cutoff of 50% missing data as the upper threshold, and thus chose visit 3 as the “endpoint” of the study, representing a mean follow‐up time of 37.8 (SD 4.3) months since study entry. Analyses were performed using SPSS statistical software, version 26.0 . All +), and patients with comorbid T2DM (PD + T2DM).An additional sensitivity analysis was conducted using vascular score as the independent variable to evaluate whether increased vascular disease is the driver of T2DM‐mediated effects on disease progression and allowed novel pathophysiological insights to be made. Patients were categorized into patients with PD with no T2DM and no vascular diseases (PD/VasR), patients with PD with no T2DM but with one or more vascular diseases  had comorbid T2DM (PD + T2DM) and 1763 (91.3%) did not (PD) , were older when diagnosed , and had a higher BMI .Compared with the PD group, patients with comorbid T2DM were 3.9 years older than the PD group (P = 0.004) and NMSS ; worse sleep scores, as assessed by Parkinson's Sleep Scale  and Epworth Sleepiness Scale ; worse cognitive scores, as assessed by MoCA ; more severe motor symptoms, as assessed by MDS‐UPDS Part III ; worse overall quality‐of‐life scores, as assessed by the EQ5D Visual Analogue Scale  and EQ5D Index ; and higher impaired scores on the Schwab and England Activities of Daily Living scale  compared with people without T2DM. Also, patients with T2DM had a higher LEDD usage than participants without T2DM .Despite a similar disease duration of 15.6 months, and after additionally adjusting for differences in age, sex, ethnicity, vascular score, baseline H&Y stage, LEDD, and BMI, patients with T2DM had consistently more severe symptomatology in most aspects of PD (Table P < 0.001), depression , and self‐reported loss of independence  in comparison with patients without T2DM (Table P = 0.029).There were significantly greater numbers of patients with T2DM who had substantial gait impairment , depression , and loss of independence  relative to the PD group, after adjusting for age, sex, disease duration, ethnicity, vascular score, LEDD, H&Y stage, and BMI  and also MCI , compared with the PD (without T2DM) group. There were no significant differences in the time to develop H&Y stage 3, dyskinesia, motor fluctuations, hallucinations, ICD‐RDs, loss of independence, and depression  of 71 patients with PD + T2DM and 340 (34%) of 986 patients with PD without T2DM. Substantial gait impairment developed in 36 (24%) of 147 patients with PD + T2DM and in 232 (13%) of 1737 patients with PD , after adjusting for age, sex, gender, ethnicity, baseline motor score, LED, and BMI, indicating that there was a significant difference in the progression of motor symptoms in patients with T2DM. During the follow‐up, patients with T2DM had significantly more severe motor symptoms as reported by MDS‐UPDRS Part II scores. In addition, this group also had significantly worse MDS‐UPDRS Part III scores at each time point , there was a significant group × time interaction in the mixed model , suggesting that the PD‐T2DM group had a significant difference in progression of mood symptoms. Supporting this, patients in the PD + T2DM group had significantly worse depression scores at each time point (P = 0.023).There was also a significant group × time interaction when modeling change in mood as measured by the NMSS Mood subscore (P = 0.001), with only the PD + T2DM group reporting loss of independence compared with the PD group  after 36 months; however, the group × time interaction failed to reach the conventional threshold for significance (P = 0.077). There were no significant differences in motor fluctuations or dyskinesia  protein in pancreatic beta islets is thought to contribute to the development of T2DM.This study is the first to report the impact of T2DM on nonmotor symptoms in PD. At study entry, in patients with a mean disease duration of 18 months, patients with T2DM already had a greater overall nonmotor symptom burden (as measured by MDS‐UPDRS Part I and NMSS) and reported poorer sleep compared with patients without T2DM. Interestingly, the main drivers for the differences in total NMSS scores were primarily sleep, mood, and memory issues, which were themselves captured on separate scales.15 Dysfunctional neuronal insulin signaling leads to increased aggregation of amyloid‐ß, hyperphosphorylated τ, proinflammatory pathway activation, and impaired brain glucose metabolism,Previous studies suggested patients with PD with T2DM were more likely to have cognitive impairment after a mean of 6 years' disease duration; however, in this study, we show even 18 months after a diagnosis of PD, comorbid T2DM was associated with significantly worse cognitive test scores and a higher proportion of MCI. In addition, in patients without cognitive impairment at baseline, the PD + T2DM group was almost twice as likely to develop MCI subsequently. Although increased leukoaraiosis in these patients has previously been thought to underlie these effects, our study did not necessitate structural imaging, and we were not able to perform a systematic analysis of this topic. However, others demonstrate no difference in leukoaraiosis between patients with PD with and without T2DM.The PD + T2DM group had significantly worse mood scores at baseline and reported consistently lower mood throughout follow‐up than the PD group. Furthermore, T2DM was independently and directly associated with worse longitudinal progression of the NMSS mood subscores throughout the follow‐up period. Depression is a strong predictor of quality of life in PD, and is itself a risk factor for PD, while similarly, there is increased prevalence of depression in patients with T2DM.Contrary to other nonmotor symptoms, patients with T2DM reported fewer symptoms of ICD‐RDs. The pathophysiology of ICD‐RDs like punding is complex but is thought to involve stimulation of D1 and D2 receptors, and studies in animals support the hypothesis that the reward system acts by means of increasing dopamine in the nucleus accumbens and the dorsal striatum (becoming conditioned cues).Given the interest in antiglycemic drugs as potential novel treatments for PD, an exploratory analysis was performed to evaluate whether metformin could reduce or restore the negative impact of T2DM. This study demonstrated that metformin use did not confer any protective effects on the diabetic PD population on any outcomes, and in fact, MoCA scores were significantly lower (worse) at 36 months in the PD + T2DM/metformin group. Interest in metformin as a potential neuroprotective drug is supported by in vivo and in vitro studies demonstrating metformin can restore dopaminergic dysfunction and reduce aggregation of α‐synuclein.The main strength of our study is that this large longitudinal study of patients with PD was deeply phenotyped with a variety of PD scales to characterize a variety of symptoms, in addition to typical motor scales, allowing us to gain a global overview of PD severity, as well as individual symptoms. We also had a large number of relevant vascular risk factor covariate data available. Using these and other data in our directed acyclic graph allowed us to select many covariates (based on a literature review and expert opinion) that allowed us to adjust for any potential differences between groups in our models to help estimate causal effects from the observed data. This was helpful in delineating and understanding confounders and potential sources of bias, and examining the independent effects of T2DM in sequential analyses. The prevalence of T2DM in our PD cohort was approximately 10%, which is in line with other reported studies, and thus the large number of patients with T2DM allowed tentative casual inferences to be made and some generalizability of the findings. Interestingly, at baseline, the PD + T2DM group was significantly older and had later disease onset compared with the PD group. Previous studies have shown the greatest risk of developing PD occurs in those T2DM patients with longer disease duration, and synergistic effects of T2DM on PD tend to occur at later stages, possibly explaining the later age of onset.An important limitation in this study, as is observed in many other longitudinal studies of this nature, is the dropout of patients during follow‐up. Dropout was significant in both groups even by month 32, but higher in the PD + T2DM group, which suggests that data may not be missing at random. One could speculate that this is driven by higher disease severity, which would support our overall findings of the negative impact of T2DM on PD severity. However, because this may introduce a bias regarding surviving patients, a decision was made to include follow‐up data for only the first 36 months for one aspect of the longitudinal analysis. The survival analyses curves may have been influenced by censored data, and further studies will be needed to confirm these findings.We used linear mixed effects models to examine the interaction effects of T2DM on a number of clinical features typically associated with PD, and our results suggested worse motor progression and mood over time were dependent on the presence of T2DM. Thus, multivariable models can simultaneously control for many covariates and also be used to test additive versus interactive effects of different covariates, which can help generate new hypotheses about the effects of T2DM on PD. However, when there are potentially many covariates, a large sample size is required, although a variety of statistical approaches are available to address this.In conclusion, we have identified T2DM as an independent risk factor associated with more severe motor symptoms, nonmotor symptoms, and poorer quality‐of‐life scores in patients recently diagnosed with PD. Furthermore, the presence of T2DM has a detrimental effect on the clinical course of PD, contributes to faster motor and nonmotor symptom progression, and increases the risk of developing MCI and gait impairment. The importance of this is that insulin resistance is potentially a modifiable metabolic state, with multiple peripheral and central targets for intervention. Thus, targeting insulin resistance may represent a novel target for alleviating parkinsonian symptoms, ameliorating neurodegeneration, and slowing progression to disability and dementia.D.A.: conception, data analysis, and manuscript writing and editing. J.E., A.W., G.V., M.L.C., N.V., C.G., T.F., and S.G.: manuscript writing and editing. M.L.: data analysis and manuscript writing and editing. Y.B.‐S., H.M., and D.G.: study design, data collection, and manuscript editing.D.A. has received honoraria from Bial Pharma and grant funding from CureParkinson's, Parkinson's UK, and National Institute for Health Research, outside of the submitted work. N.V. has received unconditional educational grants from IPSEN and Biogen, travel grants from IPSEN and AbbVie, and honorarium from AbbVie and STADA, and has served on advisory boards for AbbVie and Brittania, outside of the submitted work. Y.B.‐S. has received grant funding from the Medical Research Council, National Institute for Health Research, Parkinson's UK, National Institutes of Health, and Economic and Social Research Council. H.M. reports paid consultancy from Biogen, UCB, AbbVie, Denali, Biohaven, and Lundbeck; lecture fees/honoraria from Biogen, UCB, C4X Discovery, GE‐Healthcare, Wellcome Trust, and Movement Disorders Society; research grants from Parkinson's UK, Cure Parkinson's Trust, PSP Association, CBD Solutions, Drake Foundation, and Medical Research Council. D.G. received honoraria from Bial Pharma, Merz Pharma, and consultancy fees from The GM Clinic, Glasgow. T.F. has received honoraria from Profile Pharma, BIAL, AbbVie, Genus, Medtronic, and St. Jude Medical, outside of the submitted work. All other authors have nothing to report.Table S1. Previous studies evaluating the effects of T2DM on PD disease progression.Table S2. Longitudinal follow up. Univariate and multivariate Cox regression analyses for the development of each disease marker/clinical outcome in PD according to the presence of T2DM. Multivariate analyses adjusted for age, sex, vascular score, ethnicity, disease duration, Hoehn & Yahr stage, BMI and LEDD.Table S3. Clinical Features of patients within 18 months of diagnosis of PD compared to patients with co‐morbid T2DM and patients treated with metformin.Table S3. Multivariate Cox regression analyses for the development of each disease marker/clinical outcome in patients with PD according to the use of metformin. Multivariate analyses adjusted for age, sex, vascular score, ethnicity, disease duration, Hoehn & Yahr stage, BMI and LED.Table S4. Clinical Features of patients within 18 months of diagnosis of PD. Patients were categorised into patients with no T2DM and no vascular risk factors (PD/VasR−), patient with no T2DM but increased vascular risk factors (PD/VasR+), and patients with co‐morbid T2DM (PD + T2DM). There were no significant differences between the PD/VasR− and PD/VasR+ groups, but when compared to the increased vascular risk factors group (without T2DM), patients with T2DM had significantly worse scores in depression, quality of life and substantial gait impairment.Table S5. Multivariate Cox regression analyses for the development of each disease marker/clinical outcome in patients with PD according to the use of metformin. Multivariate analyses adjusted for age, sex, ethnicity, disease duration, Hoehn & Yahr stage, BMI and LED.Figure S1. Directed acyclic graph illustrating confounding and mediating factors to determine the causal impact of T2DM on PD severity. This considers each variable in relation to the exposure and outcome, as both the failure to adjust for a confounder, and over‐adjusting for an intermediate variable can lead to biased results16,17. Included associations were based on past literature and expert knowledge, and we used the program DAGitty18, which uses an algorithm to identify a “minimally sufficient adjustment set” containing no redundant variables, to allow us to adjust models for confounders and make causal inferences.Figure S2. Kaplan–Meier curves – no significant differences were observed in time to develop H&Y Stage 3, Dyskinesia, Motor fluctuations, Hallucinations, ICD, Loss of independence, Depression.Figure S3. Linear mixed modelling progression of symptoms of PD.Figure S4. Longitudinal impact of T2DM on symptoms in PD per group. Multivariate Cox regression analyses for the development of each disease marker/clinical outcome in PD according to the presence of T2DM. Multivariate analyses adjusted for age, sex, ethnicity, disease duration, vascular score, Hoehn & Yahr stage, BMI and LEDD.Figure S5. Longitudinal impact of T2DM on symptoms in PD per group.Figure S6. Linear mixed modelling progression of symptoms of PD vs PD + T2DM and PD + T2DM/Met.Figure S7. Longitudinal follow up. Univariate regression analyses for the development of each disease marker/clinical outcome in PD according to the presence of no vascular risk factors (PD/VasR−), increased vascular risk factors (PD/VasR+), and T2DM.Click here for additional data file.


This study aimed to evaluate possible treatment strategies for patients with de novo T790M mutation‐positive (T790M+) non‐small‐cell lung cancer (NSCLC).Patients diagnosed with de novo T790M+ NSCLC and treated with epidermal growth factor receptor tyrosine kinase inhibitors (EGFR‐TKIs) between 2011 and 2018 at a regional hospital in Taiwan were retrospectively reviewed. Their clinicopathological characteristics and subsequent treatment information were collected, and potential prognostic factors were identified using univariate and multivariate analyses.p < 0.01) and overall survival  than gefitinib and erlotinib. Additionally, patients with low T790M ratios had better PFS than those with high T790M ratios, implying that the proportion of T790M+ tumors determined the response to EGFR‐TKIs. Multivariate analysis confirmed that both EGFR‐TKI treatment  and a low T790M ratio  were independently favorable prognostic factors for patients with de novo T790M+ NSCLC. Median PFS was 6.1 (95% CI 4.4–7.8) months. In addition, patients treated with first‐generation (1G)/second‐generation (2G) EGFR‐TKIs followed by osimertinib (n = 8) demonstrated the best OS compared with patients treated with frontline osimertinib (n = 5) or 1G/2G EGFR‐TKIs without osimertinib .All tumors with T790M mutations coexisted with sensitizing mutations. Through the last follow‐up in May 2021, afatinib and osimertinib demonstrated better progression‐free survival (PFS, Sequential TKIs may represent an alternative option for de novo T790M mutation, particularly frontline afatinib and tumors with low T790M ratios. Patients with high T790M ratios had better progression‐free survival than patients with low T790M ratios. Osimertinib, afatinib, and low T790M ratios were favorable prognostic factors. Patients with a particularly low de novo T790M mutation ratio should be treated by afatinib followed by osimertinib rather than frontline osimertinib. A subclone of T790M mutation tumors may influence the response to EGFR‐TKIs. EGFR mutations (EGFRm+).Molecular targeting using epidermal growth factor receptor tyrosine kinase inhibitors (EGFR‐TKIs), including first‐generation (1G) TKIs, such as gefitinib and erlotinib,In contrast to the acquired T790M mutation, de novo T790M mutations are found in treatment‐naïve NSCLC, and the frequency of de novo T790M mutations depends on the sensitivity of the methods used for mutation detection.Previous studies have demonstrated conflicting results, and no direct comparison has been performed among EGFR‐TKIs due to the low prevalence of the de novo T790M mutation. Therefore, we aimed to compare the efficacy of different EGFR‐TKIs in de novo T790M mutation‐positive (T790M+) NSCLC patients to identify an alternative therapeutic strategy for the de novo T790M mutation.EGFR mutation status was retrospectively reviewed.Patient data were obtained from the Cancer Registry System using the Chang Gung Research Database,EGFR mutation, such as concurrent exon 19 deletion, L858R, or uncommon mutation, tumor response, and subsequent treatment, were obtained. The last follow‐up time point collected in this study was May 2021.The clinical data of 44 patients who received EGFR‐TKIs as the first‐line treatment were retrospectively reviewed. Furthermore, the clinicopathological features, including age, sex, smoking history, Eastern Cooperative Oncology Group performance status (ECOG PS) score, tumor involvement, This study was approved by the Institutional Review Board of CGMH (201901395B0). Patient consent to participate was not required due to the retrospective nature of this study.EGFR mutation prior to 2012. The development of the amplification refractory mutation system (ARMS) increased the sensitivity of EGFR mutation detection, and after 2012 most patients' EGFR mutations have been detected using ARMS, such as QIAGEN and TaqMan assays unless the physicians required other methods. Using ARMS, delta CT (dCT) values could be obtained for T790M and other sensitizing mutations. Differences in the dCT values for the T790M mutation and sensitizing mutations (i.e. dCT [T790M] − dCT [sensitizing mutation]) indicated the ratio between the T790M mutation and other EGFR sensitizing mutations (T790M ratio) within the tumor sample. For more than two sensitizing mutations detected within the same tumor, the mutation with lower dCT was used as the reference. Due to the relatively low sensitivity of Sanger sequencing, any T790M mutation detected by Sanger sequencing was categorized as having a high T790M ratio in the present study. The present study evaluated the prognostic value of the T790M ratio for estimating PFS. The T790M ratio was found to be relatively variable, therefore it was classified into two subgroups, high and low T790M ratio, according to the median value.Sanger sequencing was the only method for detecting All patients were administered EGFR‐TKIs once daily until disease progression or intolerable toxicity. The tumor response was evaluated using the Response Evaluation Criteria in Solid Tumors 1.1 criteria. The best clinical tumor response was recorded based on the radiological findings: the complete response (CR), PR, SD, or progressive disease (PD). Any tumor response that was not assessed before death or discontinuation due to intolerance was recorded as not assessed (NA). The objective response rate (ORR) was the sum of CR and PR, and the disease control rate (DCR) was the sum of CR, PR, and SD. In the survival analysis, PFS was defined as the duration from the first day of EGFR‐TKI treatment until the first radiological evidence of disease progression, EGFR‐TKI discontinuance due to toxicity or other reasons, death, or the latest follow‐up time point. Those patients who experienced no progression or death during the treatment were censored during PFS analysis. Patients with radiological progression or death within 1 month after EGFR‐TKI discontinuation and who received no sequential treatment were counted as an event. OS was defined as the duration from the first day of EGFR‐TKI treatment until the date of death or the last follow‐up. Data of patients who did not experience death were censored during survival analysis.p < 0.1 on univariate analysis, to evaluate the independent prognostic factors. The results are presented as the hazard ratio (HR) and 95% confidence interval (CI) based on Cox regression analyses. IBM SPSS Statistics for Windows  was used to perform all statistical analyses, and p < 0.05 was considered significant. We used the R packages “survival” and “survminer” to plot the survival curves.The PFS and OS were estimated using the Kaplan–Meier method and compared using the log‐rank test. Univariate analysis was performed to evaluate possible prognostic factors, including age, sex, staging, T790M ratio, ECOG PS, smoking history, and tumor involvement. Multivariate analysis was performed, including all variables with A total of 2420 patients with EGFRm+ NSCLC treated with frontline EGFR‐TKIs were reviewed. Among them, 2190 (90.5%) patients had a common mutation (exon 19 deletion or L858R mutations), 186 (7.7%) had an uncommon mutation other than T790M, and 44 (1.8%) had de novo T790M mutation. The de novo T790M mutations were detected using the AMRS  in 38 patients, the cobas® EGFR Mutation Test in one patient, Sanger sequencing in four patients, and an unknown assay in one.p = 0.032; Table A total of 44 patients with T790M mutation retained for this study had a mean age of 71 years, and 32 (72.7%) patients were women. All but one patient presented metastatic NSCLC (stage IV). Among this population, 21 (47.7%), six (13.6%), 12 (27.3%), and five (11.4%) patients were treated with gefitinib, erlotinib, afatinib, and osimertinib, respectively, as first‐line treatments. Patients who received osimertinib and erlotinib were more likely to have experienced metastasis to the brain (p < 0.05). Patients who received afatinib  and osimertinib  had significantly longer PFS than patients who received erlotinib  and osimertinib  were significantly associated with lower risks of disease progression relative to the patients who received erlotinib. Similarly, patients with a low T790M ratio were less likely to experience disease progression than those with a high T790M ratio . Moreover, patients with no liver metastasis had a reduced risk of disease progression compared with those who experienced liver metastasis  . Unlike the contribution of the T790M ratio to PFS, the T790M ratio had no impact on OS .TKI, sex, smoking status, and ECOG PS were individually associated with OS in the univariate regression analysis. Patients treated with afatinib , osimertinib , and gefitinib  were significantly associated with a lower risk of death compared with those treated with erlotinib  Figure . Male sep value <0.1 according to the univariate analysis, TKI, ECOG PS, and lung metastasis were independently associated with OS after adjusting for smoking status and sex in the multivariate regression analysis. Patients treated with afatinib  and osimertinib  were significantly associated with lower risks of death, whereas those with lung metastasis  and ECOG PS of 2–4  were significantly associated with a higher risk of death after adjusting for potential confounders .Further analysis of OS was performed according to subsequent treatment. Patients who received subsequent osimertinib treatment following 1G/2G EGFR‐TKIs  had longer OS than patients without subsequent osimertinib  and patients treated with osimertinib alone  experienced better survival than patients who received frontline line osimertinib or 1G/2G EGFR‐TKIs without subsequent osimertinib treatment.p = 0.0185),The rate of de novo T790M mutation detection depends on the sensitivity of the methods used for mutation detection.33 These findings suggest that a high T790M ratio may represent a good prognostic factor for patients with either de novo or acquired T790M mutations, indicating that osimertinib may be more active for T790M mutations than EGFR‐sensitizing mutations, consistent with the findings from in vitro cell lines.Although osimertinib can target both T790M and EGFR‐sensitizing mutations, the activity of osimertinib against EGFR sensitizing mutations may not be as good as 1G/2G EGFR‐TKIs.Therefore, a current dilemma is whether frontline osimertinib or 1G/2G EGFR‐TKIs represent a better treatment for de novo T790M+ NSCLC patients. The latter treatment could present the opportunity for switching to osimertinib following progression on previous EGFR‐TKI treatment and lead to the best survival outcome, based on the results from our cohort Figure . TheoretThis study has several limitations. First, this was a retrospective study enrolling limited cases, which also reflected the relative rarity of de novo T790M detection when using traditional detection methods in clinical practice. Second, limited cases treated with frontline osimertinib were included because osimertinib has only been available for the last few years and is not reimbursed by Taiwan national health insurance. Third, the T790M ratio was only a relative value estimated using ARMS, a semiquantitative method of detection; the T790M ratio evaluation using more sensitive methods, such as NGS or ddPCR (quantitative methods), is important and is warranted in the future. Fourth, only five and eight patients received osimertinib as frontline and subsequential treatment, respectively; hence, the survival benefit should be interpreted with caution. In addition, patients treated with frontline osimertinib tended to have brain metastasis, which might lead to worse OS than patients without brain metastasis. However, despite these limitations, this study was able to provide an alternative view based on the clinical findings when treating de novo T790M mutations with treatments other than osimertinib.Our study provided evidence that sequential treatment with 1G/2G EGFR‐TKIs followed by osimertinib was effective in de novo T790M+ NSCLC patients. Afatinib was the best choice among the EGFK‐TKIs, particularly for patients with a low T790M ratio Figure .The authors declare no conflicts of interest.


Neurogenic erectile dysfunction (nED) is one of the most common and intractable postoperative complications of rectal and prostate cancer surgery and sometimes accompanies patients lifelong. The transplantation of stem cells has been proved a promising way for treatment. However, the therapeutic efficacy is severely impaired by excessive cell loss and death and poor accumulation in the injury site along with the traditional implantation strategy. Herein, an EPO‐loaded multifunctional hydrogel was designed. The hydrogels' adhesive property and mechanical strength were enhanced by adding catechol‐catechol adducts, thus significantly improving adipose‐derived stem cells (ADSC) retention and rescuing cell loss in the injury site. Meanwhile, the sustained release of EPO effectively ameliorated the viability and paracrine activity of ADSC, leading to enhanced migration of Schwann cells and differentiation of PC12 cells in vivo. On a bilateral cavernous nerve injury rat model, the present stem cell‐EPO‐hydrogel implanted strategy could significantly alleviate erectile dysfunction. The higher expression of Tuj1 and lower expression of GFAP in the major pelvic ganglia (MPG) indicated the acceleration of neural differentiation while the suppressing development of astrocytes. Also, the combined therapy restored the expression levels of eNOs, nNOs, and α‐SMA in penile tissues, suggesting the rehabilitation of the penis. Further analysis of Masson trichrome staining and apoptosis evaluation of the corpus cavernosum showed the preservation of vascular endothelium content and the prevention of penile fibrosis after denervation. Overall, we believe that this combined strategy presents a promising way not only for restoring neurogenic erectile function but also for the clinical translation of stem cell therapy. Radical resection of the lesion is one of the most efficient treatment options. Postoperative erectile dysfunction (ED) is a significant complication and frequently occurs in male patients for the sake of inadvertent crush, tension, and resection injuries to the pelvic autonomic nerve (PAN) during the surgical procedure.Unlike ED resulting from aging and diabetes, nED is essentially a neurotraumatic disease. Previous studies have revealed that sufficient penile erection cannot be imagined without a sound neural network, an intact vascular system, and healthy cavernosal tissue.In the past decades, emerging evidence has proved that stem cell therapy such as adipose‐derived stem cells (ADSC) holds great potential for nerve regeneration and tissue engineering.Hydrogels are polymeric networks that bond together by physical and/or covalent crosslinks. Due to its excellent biological properties, biodegradability, and biocompatibility, hydrogels derived from natural biomaterials have been extensively developed in tissue engineering and nerve regeneration. Meanwhile, increasing evidence has demonstrated the advantage of the combined implantation of stem cells with biocompatible hydrogels over utilizing alone, in which hydrogels provide a temporary shelter for cell adhesion, proliferation, and migration, while the implanted cells modulate the local microenvironment to compensate for the damaged tissues.Besides, due to the intricacy of the microenvironment after nerve injury such as Wallerian degeneration and inflammatory infiltration, the addition of exogenous of neuro‐protective‐related cytokines has been revealed to further improve stem cell therapeutic efficiencies, such as nerve growth factor (NGF), platelet‐derived growth factor (PDGF), brain‐derived neurotrophic factor (BDNF), NT3, and NT4.Herein, an EPO‐loaded multifunctional hydrogel was constructed for cell implantation to an nED rat model Scheme . Methacr22.1Gelatin type A (from porcine skin), methacrylic anhydride (MA), and carboxymethyl chitosan (CMC) were purchased from Macklin Biochemical Technology Co., Ltd. . Active erythropoietin (EPO) was bought from Cloud‐Clone Crop. Hyaluronic acid (HA) was obtained from Freda Biopharma Co., Ltd. Dopamine hydrochloride  was bought from Aladdin Biochemical Technology Co., Ltd. All reagents were of analytical grade unless otherwise noted.2.2GelMA was prepared as previously described with slight modifications.HADA was synthesized by an EDC/NHS reaction.To prepare the CMCDA conjugate, 1‐g CMC was first completely dissolved in deionized water (20 ml) at 60°C, followed by the addition of 575‐mg EDC. In addition, 345 mg NHS and 569 mg DA were weighed and dissolved in 20 ml deionized water. Then, the CMC/EDC solution was dropwise added to the DA/NHS mixture. The pH value was kept at 5, and the by‐products were removed by centrifugation after reaction in nitrogen ambient for 6 h. The supernatant was harvested and dialyzed against hydrochloric acid (pH = 5) for 24 h, then dialyzed against deionized water. The CMCDA solution was lyophilized at −80°C and stored at −20°C before use.2.3To construct precursors of hydrogels, the concentration of GelMA was maintained at 10% (wt/vol), with or without the supplement of 2% (wt/vol) HADA and 5% (wt/vol) CMCDA solution (3: 2: 1 by volume), followed by the addition of 0.1% (wt/vol) photo‐initiator LAP. A 365‐nm UV laser was applied for gelatinization.2.42.4.1−1 at a resolution of 4 cm−1. Finally, Gelatin and GelMA were dissolved in D2O, and the nuclear magnetic resonance (1H‐NMR) spectra were recorded on a 600 MHz nuclear magnetic resonance spectrometer (Bruker AVANCE).Briefly, UV–vis spectrophotometer  and Fourier‐transform infrared (FT‐IR) spectroscopy (Bruker VERTEX70) was applied to determine the chemical structures of HA, HADA, CMC, and CMCDA. Samples were dissolved in pure water, and the wavelength was set ranging from 190 to 500 nm to obtain the absorption spectra. As for FT‐IR, the spectra were investigated over a scanning wavelength from 4000 to 399 cm2.4.2m0. Then, the hydrogels were submerged in PBS at 37°C, and the swollen weight was measured at selected time intervals and recorded as m1. The swelling rate was calculated according to the following equation:A conventional gravimetric method was performed to assess the swelling properties. Lyophilized prepared hydrogels were weighed and recorded as 2.4.3Rheological tests were performed using a rotary rheometer (Kinexus) at 25°C with plate geometry (plate diameter: 25 mm). Frequency sweep tests were conducted to analyze the stability of the hydrogels over a range frequency from 0.1 to 10 Hz and a 5% constant strain. Time sweep tests were carried on to observe the stiffness of the hydrogels at a 1 Hz frequency and a 1% constant strain. Besides, a Bose ElectroForce 3200 instrument (Bose) was utilized to analyze the compressive stress–strain and adhesive property. For the compression test, the 6‐mm‐high hydrogels were subjected to compression until crushing. For the adhesion test, 200‐μl hydrogels were applied between two pieces of fresh porcine skin (adhesion area: 1 cm × 1 cm).2.4.4w0 and w1 represent the dry weight of the initial hydrogels and the remaining hydrogels at different time point, respectively.Enzymatic degradation experiments were performed to systematically evaluate the degradation behavior of hydrogels. Briefly, the equal volume hydrogels were immersed in 20 ml PBS, with or without the addition of 1000 U/ml lysozyme. The solution system was kept at 37°C under constant agitation at 100 rpm. At the selected time intervals, samples were taken out and lyophilized. The pore structure and morphology of the lyophilized hydrogels were determined by an S‐3400 scanning electron microscope (Hitachi). The weights were obtained, and the degradation rate was calculated by the following equation:2.5Fifty‐microliter sterile PBS buffer containing EPO  was mixed with 950 μl of GelMA/HADA/CMCDA pre‐gel solution completely at 37°C. EPO 500 IU/ml concentration was reported to display the most potent protective effect to stem cells.2.62. The culture medium was changed every 3 days, and all the cells used in experiments were at Passages 3–5. Flow cytometry was performed to identify the obtained cells. The following PE‐labeled antibodies were used: CD11b, CD45, CD14, CD29, CD44, and CD90. The data were determined by a flow cytometer (BD Bioscience). Additionally, adipocytes and osteoblasts differentiation of ADSC was further observed by culturing in a special medium. Alizarin red S was used for osteoblasts staining, and oil red O was applied for adipocytes staining.As published previously, primary adipose‐derived stem cells (ADSC) were isolated from epididymal adipose tissue of 2‐week‐old male Sprague–Dawley rats.SCs were isolated from the sciatic nerve of 2‐week‐old SD rats. First, the rats were killed, and the bilateral sciatic nerves were collected. Then, the nerve bundle was cut into ≈3‐mm segments and incubated in high‐glucose DMEM supplemented with 10% FBS at 37°C in a humidified atmosphere containing 5% CO2. Five micrograms per milliliter of cytarabine was used to purify SC. Immunofluorescence staining for S100β was performed for the identification of SC.PC12 cells  were obtained from Zhongqiao Xinzhou Biotechnology Co., Ltd., and cultured in DMEM supplemented with 10% FBS for proliferation.2.74 cells/dish). One‐milliliter culture medium was then supplemented and co‐cultured for different time intervals . At each time indicated, the cells were washed with PBS three times, and the live/dead reagents  were added. After incubation for 30 min, the survival and growth of ADSC were visualized under a confocal laser scanning microscope . Meanwhile, to observe the cell morphology, the cell attached‐hydrogels were fixed with paraformaldehyde (4%), followed by added with TRITC phalloidin for staining F‐actin and DAPI for staining the nuclei. Additionally, a CCK‐8 assay was performed to assess the cell viability according to the protocol. The absorbance value was measured at λ = 450 nm.Calcein‐AM/PI double staining, cytoskeleton staining, and cell counting kit‐8 test were utilized to assess the cytotoxicity of hydrogels on ADSC. Before seeding cells, the 35‐mm confocal dishes  were precoated with 100 μl of GelMA, GelMA/HADA/CMCDA, and GelMA/HADA/CMCDA/EPO pre‐gel solution, respectively. After being cured for 20 s by a 365‐nm UV laser, 200‐μl cell suspension of ADSC was seeded on the hydrogels (1 × 102.87 cells/ml) in the logarithmic growth phase were added to 950 μl of GelMA, GelMA/HADA/CMCDA, and GelMA/HADA/CMCDA/EPO pre‐gel solution, respectively. After mixing well by pipetting, 300 μl of the above pre‐gel solutions were coated on the 48‐well plate, and then a hydrogel was polymerized by UV irradiation. The cell‐encapsulated hydrogels were co‐cultured in low‐glucose DMEM. After incubated for different time intervals , the hydrogels were transferred to 35‐mm confocal dishes and gently rinsed with PBS three times. Similarly, the live/dead staining assay and cytoskeleton staining assay were performed the same as described in the Biocompatibility test in vitro section to observe the spatial distribution, viability, and morphology of the cells loaded in the hydrogels. CCK‐8 assay was utilized to determine the viability of the encapsulated cells.ADSC encapsulation into the hydrogels was performed on previously published protocols with slight modifications.2.94 cells/well). Fresh medium without any supplement was set as a control group. After co‐culture for 3 days, 200‐μl cell suspension of SC (1 × 104 cells) was seeded in the upper chamber . After incubation for 48 h, cells remaining on the upper membrane were removed with cotton swabs, and the migrated cells under the membrane were fixed with 4% paraformaldehyde and stained with crystal violet solution for 20 min. Finally, the upper chamber was gently washed in PBS, and the immigrated cells were observed under an inverted microscope. The cell numbers of three randomly chosen fields were counted.The transwell experiment was further employed to determine the effect of different cell co‐culture systems on the chemotaxis of SC. The cells of the 24‐well plate were randomly divided into ADSC, pre‐coated GelMA/HADA/CMCDA + ADSC, pre‐coated GelMA/HADA/CMCDA/EPO, and pre‐coated GelMA/HADA/CMCDA/EPO + ADSC  was determined by ELISA kits (Meimian Biotechnology Co. Ltd.). Three independent experiments were conducted for statistical analysis.The enzyme‐linked immunosorbent assay (ELISA) was performed to investigate the paracrine response of ADSC to hydrogels. Briefly, 100‐μl pre‐gels with or without EPO were added to each well of the 24‐well plate and gelated before the experiment, and the well without any supplement was set as control. Then, ADSC was seeded into the 24‐well plate at the density of 1 × 102.11Good differentiated PC12 cells demonstrate similar morphology, physiology, and biochemistry to nerve cells. Therefore, the supernatants of different cell co‐culture systems mentioned in the 2.9 section were harvested to culture PC12 cells. After different time intervals, PC12 cells were stained with Calcein‐AM and observed under a fluorescence microscope. The neurite length and growth ratio of PC12 cells were counted to assess the neurite growth. Specifically, 100 cells were randomly selected, and the length of neurites was measured by FIJI/ImageJ software. Cells were divided into four types according to the ratio of synapses with cell size: no neurite (L0), neurites length shorter than the size of the cell body (L1), neurites length between the original and twice of the cell body (L2), neurites length between twice and three times of the cell body (L3), and neurites length over three times of the cell body (L4).2.12To investigate the fate of the implanted ADSC in vivo, PKH26 dye  was used to stain the cells according to the protocol. Then, the labeled cells were encapsulated into GelMA/HADA/CMCDA/EPO hydrogel for implantation of BCNI rats. The PKH26 labeled cells in the hydrogels and MPG tissues were determined under a fluorescence microscope at different time intervals.2.13n = 5): (1) PBS group: 50 μl PBS was injected into the crush area on each side; (2) Gel group: 50 μl GelMA/HADA/CMCDA was applied for treatment on each side; (3) Gel/EPO group: 50 μl GelMA/HADA/CMCDA/EPO was used on each side; (4) Gel/ADSC group: 50 μl GelMA/HADA/CMCDA containing 1× 106 ADSC; (5) Gel/EPO/ADSC group: 50 μl GelMA/HADA/CMCDA/EPO containing 1× 106 ADSC was implanted on each crush side. Finally, experimental animals were kept for further evaluation. For the sake of description, Gel represents GelMA/HADA/CMCDA hydrogel.All animal experiments were approved by the Ethics Committee of the Institutional Animal Care and Use Subcommittee of the Third Hospital of Sun Yat‐sen University. To establish the bilateral cavernous nerve injury (BCNI) model, 8‐week‐old Sprague–Dawley male rats were prepared and anesthetized with 2.5%–3% isoflurane. Next, the MPG/CN were exposed posterolaterally on both sides of the prostate.2.14Four weeks after surgery, both intracavernous pressure (ICP) and mean arterial blood pressure (MAP) were recorded continuously to evaluate the erectile function of animals. After anaesthetization, the right carotid artery was exposed through a midline incision from the neck to the upper thorax, and a heparinized (250 U/ml) silastic cannula connected with a three‐way stopcock was inserted to record the MAP. Similarly, a heparinized butterfly needle (24‐G) connected to a pressure transducer was implanted in the corpus cavernosum to measure the ICP. To elicit erection, a bipolar electrode with a 5 V, 20 Hz electric stimulus was applied to stimulate the CN directly. All data were recorded by a BL‐420s Biological Functional System . After the experiment period, penile tissues and MPG/CN were harvested for further analysis.2.15Freshly dissected penile and MPG tissues were harvested and fixed with 4% paraformaldehyde for further histological staining analysis. Masson trichrome staining was first carried out to determine smooth muscle and collagen expression levels in penile tissues. A 5‐μm slice was prepared and stained following the manufacturer's instructions, and the smooth muscle was stained red while the connective tissue appeared blue. For immunofluorescence staining, the penile sections were incubated with primary antibodies including α‐SMA , eNOS , nNOS , Caspase‐3 , and the MPG sections were covered by primary antibodies including Tuj1 , and GFAP  at 4°C overnight. After rinsing the slices with PBS, secondary antibodies were adopted for 1‐h immersion. Nuclei were stained with DAPI. Images were observed, and the interesting areas were captured under a confocal laser scanning microscope (Zeiss LSM 710) and a fluorescence microscope . FIJI/ImageJ software  was used to carry out image analysis.2.16Relative expression levels of the expected proteins in penile tissues were detected by western blot analysis as previously described.2.17t‐test was performed for comparisons between two groups, whereas one‐way analysis of variance (ANOVA) followed by post hoc Bonferroni analysis was applied for comparing multiple groups. Statistical significance is demonstrated as (*) p < 0.05, (**) p < 0.01, and (***) p < 0.001.All data were analyzed using SPSS 26.0 software and displayed as mean ± SD. Unpaired Student's 33.11H‐NMR  indicated the preparation of HADA, while the presence of a peak at 1580 cm−1 corresponding to the benzene ring skeleton vibration (VC═C) suggested the fabrication of CMCDA. Notably, the characteristic peak of the amide bond (VC═O) was not observed in the spectrum of CMCDA, which was considered to be masked by the C═C peak. For UV–vis absorption spectra, a slight blue shift (≈4 nm) was observed both in HADA and CMCDA, confirming the generation of amide linkages. Figure SEM results  was used to simulate the in vivo environment.s Figure . Apparen3.4Compared with other kinds of stem cells, ADSC is easy to isolate from tissues and possesses the properties of higher activity to secrete neuroprotective related cytokines and immunoregulatory factors while lower immunogenicity.Next, the effect of hydrogel materials on ADSC viability and proliferation was assessed. The survival of ADSC spreading on hydrogels was visualized via live/dead staining Figure . Little 3.5The above results displayed the excellent biocompatibility and the potential of GelMA/HADA/CMCDA/EPO hydrogel as a carrier for cell implantation. Hence, we further analyzed the viability and spreading of ADSC in the 3D cell culture hydrogels. As shown in Figures 3.6Peripheral nerve regeneration is a complex biological process, and that immediately occurs after nerve injury. SCs are the primary cells that make up the peripheral nervous system, which is activated to phagocytize the debris in the initial time and enhance neurite growth by producing neuro‐protective related factors in the later period.3.7To verify the capability of the hydrogel complex in nerve regeneration, PC12 cells were incubated with the supernatants, and the morphology was observed to evaluate the effects of hydrogels on cell differentiation Figure . As show3.8max to MAP and the ratio of relative total ICP (area under the curve) to MAP were calculated to evaluate the erectile functions. Generally, no difference was found in MAP among groups. After the crush to CN, rats exhibited the most severe ED (PBS group), while varying degrees of function improvement was achieved after the treatment of different hydrogel complex, with the most effective therapy in the Gel/EPO/ADSC group . In addition, EPO receptor expression was identified and localized in penile tissues and the periprostatic neurovascular bundles responsible for erectile function.max/MAP value and relative total ICP/MAP value of Gel and Gel/EPO group were higher than those of the PBS group, suggesting the benefits of hydrogel materials and EPO in function restoration.Encouraged by the above enhancement of nerve regeneration, the effects of EPO‐loaded GelMA/HADA/CMCDA hydrogel on erectile function improvement and synergism with ADSC were assessed via local implantation in a rat BCNI model. The EPO‐loaded GelMA/HADA/CMCDA hydrogel encapsulated with ADSC (Gel/EPO/ADSC group) was immediately injected to the local nerve crush site and polymerized by UV irradiation, with merely PBS implantation (PBS group), GelMA/HADA/CMCDA hydrogel implantation (Gel group), EPO‐loaded GelMA/HADA/CMCDA hydrogel implantation (Gel/EPO group), and GelMA/HADA/CMCDA hydrogel encapsulated with ADSC implantation (Gel/ADSC group) performed in parallel as controls Figure . Four wep Figure . The str3.9At present, stem cells have been preliminarily explored in nED treatment research, of which the administration routes are IC and IP.Next, the residual hydrogels were harvested and observed under a fluorescence microscope. Interestingly, a large amount of PKH26 labeled ADSC was detected on the first day, and only a small number of labeled cells remained on the 7th day Figure . CompareTo further investigate the fate of the delivered ADSC, the MPG tissues were collected and sliced for the monitor of PKH26 labeled ADSC at 3 d, 7 d, and 14 d. As shown in Figure 3.10Regeneration of an injured nerve is a complex biological process involving the infiltration of neurons and myelination, the chief obstacle of which is glial scar formation. To further understand the underlying mechanism of the erectile function restoration, immunofluorescence staining of MPG/CN was carried out to observe the nerve regeneration elicited among groups. Herein, the expression level of class III β‐tubulin (Tuj1) for neuronal cells and glial fibrillary acidic protein (GFAP) for activated astrocytes were detected, respectively. Fluorescence microscopy images demonstrated significant differences in the density and distribution of Tuj1 and GFAP among groups Figure . The exp3.11A good neural network, an intact vascular system, and healthy cavernosal tissue ensure sufficient penile erection. Damage to CN will lead to a series of pathophysiological changes in the architecture of the penis, such as loss of endothelial tissues, denervation, and decrease of cavernosal smooth muscle ingredient.Taken together, these results showed consistent outcomes with erectile function detection and nerve regeneration assessment, suggesting that the EPO‐loaded hydrogel synergizing with ADSC contributed to restoring nED via enhancing rapid nerve regeneration and penile rehabilitation.3.12Nerve injury can lead to different degrees of dysfunction of target organs. Although the damage is limited to the neuron, the functional recovery involves the neuronal cell body, axon, and target organ. Therefore, the study of nerve injury mainly includes three aspects: the protection of neurons, the promotion of nerve regeneration, and the prevention and treatment of denervated target organ atrophy. The delay or irreversible damage of any of these links will affect the rehabilitation of function, ultimately affecting the curative effect. Apoptosis and fibrosis are essential mechanisms leading to nED, which was reported to occur as early as 1 week after BCNI.4In this study, we designed and fabricated an EPO‐loaded multifunctional hydrogel encapsulated with ADSC for the effective restoration of nED. The improved adhesive property and mechanical strength of hydrogels were achieved by introducing catechol‐catechol adducts HADA and CMCDA, thereby effectively ensuring ADSC accumulation in the injury site. Meanwhile, the hydrogel sustaining release of EPO improved the viability of ADSC, leading to enhanced proliferation and paracrine activity of ADSC. In vitro results showed that the multifunctional hydrogels could promote the migration of SCs and the differentiation of PC 12 cells. On a bilateral cavernous nerve injury rat model, the stem cell‐EPO‐hydrogel combing strategy induced a significant erectile function restoration. Meanwhile, the hydrogels could promote nerve regeneration while inhibiting glial scar formation. More importantly, timely and effective nerve repair could inhibit apoptosis and enhance the rehabilitation of the denervated penile tissues. Our results, collectively, present a promising strategy not only for the restoration of neurogenic erectile function but also for the clinical translation of stem cell therapy.Jun Shao: Conceptualization (lead); formal analysis (lead); investigation (lead); methodology (lead); validation ; writing – original draft (lead); writing – review and editing . Pan Nie: Data curation ; investigation ; methodology (lead). Wende Yang: Conceptualization ; investigation ; methodology . Rui Guo: Conceptualization ; methodology (lead); resources (lead). Dongbing Ding: Data curation ; formal analysis ; validation . Rongpu Liang: Investigation ; writing – original draft (supporting). Bo Wei: Conceptualization ; funding acquisition ; supervision ; validation . Hongbo Wei: Conceptualization ; funding acquisition ; project administration ; supervision ; validation .The authors declare no conflict of interests.https://publons.com/publon/10.1002/btm2.10319.The peer review history for this article is available at DATA S1 Supplementary FiguresClick here for additional data file.


Dear Editor,A woman in her 40s with a history of atopic dermatitis in her childhood and no other medical history consulted with a disseminated, pruritic, papulonodular eruption of 7 months of duration. On examination, the patient had multiple papules and nodules with a crateriform appearance, an erythematous border and a large central necrotic crust, affecting the trunk, arms and legs Figure . There wThe histopathological study showed epidermal invagination with signs of ulceration and transepidermal elimination of collagen, accompanied by a marked intraepidermal inflammation with polymorphonuclear leukocytes without eosinophils. In the dermis, there was a mixed inflammatory infiltrate composed of polymorphonuclear and histiocytic cells.6/L), elevated non‐specific IgE (519 kU/L) and positive antinuclear antibodies (320 URF) without any antigenic specificity.Complete blood count, chemistry profile, and hepatic, renal and thyroid function were normal except for eosinophilia  UVB, and later psoralen plus ultraviolet A (PUVA), was started with a partial response after 8 months. Subsequently, the patient received different treatments: oral corticosteroids, antihistamines, topical corticosteroids under occlusion and cyclosporine with partial improvement and repeated flares. Three years after diagnosis, subcutaneous dupilumab was prescribed with a loading dose of 600 mg and then 300 mg fortnightly, together with NB‐UVB for 6 weeks. After 2 months, there was improvement in pruritus without new lesions. After 12 months, the patient presented a nearly complete response, with only residual hyperpigmentation and marked improvement in her quality of life Figure  which peAcquired perforating dermatoses (APD) are a rare group of skin disorders of unknown aetiology characterised by transepidermal elimination of dermal material of which reactive perforating collagenosis is the most common. Although its pathogenesis remains unclear, pruritus and repeated trauma from scratching are regarded as central pathogenic factors.Treatment for APD can be challenging. The most common treatments are topical and intralesional steroids, oral antihistamines and topical retinoids. Other treatment options include NB‐UVB, PUVA, oral retinoids, allopurinol, tetracyclines, dapsone, hydroxychloroquine, methotrexate and apremilast with variable results.Chronic pruritus found in APD mostly depends on a non‐histaminergic pathwayTreatment of APD is directed towards aetiological factors and pruritus control, with usually poor response to most therapies. Given the role of IL‐4 and IL‐13 in chronic pruritus, dupilumab could represent a potential treatment for patients with APD. Future research should be directed towards targeted therapies such as anti‐IL 4, 13 and 31 or JAK inhibitors in APD.The authors involved have reported no relevant financial relationships with commercial interest(s).Dr. Riquelme has received honoraria as a consultant for Lilly and has received speaker fees from Sanofi, both outside the submitted work. Dr. Mascaró has received speaker fees from Academia Española de Dermatología, Bocemtium Consulting, Clover Soluciones Globales de Marketing, Ferrer Internacional, Fundació Clínic per la Recerca Biomédica, LEO Pharma Spain, Loki & Dimas, Luzan 5 Health Consulting, and M.S.D. de España S.A, S&H Medical Science Service, and Sanofi‐Aventis all outside the submitted work.The patients in this manuscript have given written informed consent to the publication of their case details.


Behavioral and physiological (self-)regulation in early life is crucial for the understanding of childhood development and adjustment. The autonomic nervous system (ANS) is a main player in the regulative system and should therefore be modulated by the quality of interactive behavior of the caregiver. We experimentally investigated the ANS response of 18–36-month-old children in response to the quality of maternal behavior during a mother–child-interacting paradigm.via the pre-ejection period (PEP) and the left ventricular ejection time (LVET), and the parasympathetic nervous system (PNS) activity via the respiratory sinus arrhythmia (RSA) were continuously measured by an electrocardiogram. Maternal sensitivity of interactive behavior was assessed by using the Emotional Availability (EA) Scales.Eighty mothers and their children came to our laboratory and took part in an experimental paradigm, consisting of three episodes: a resting phase (E1), a structured play phase (E2), and a free play situation (E3) between mothers and their child. Children’s and mother’s heart rate (HR), the sympathetic nervous system (SNS) activity Children of mothers with insensitive behavior had a significantly lower RSA at baseline, showed a lack of RSA withdrawal during structured and free play, and had shorter LVET across all episodes compared to children of sensitive mothers.Our findings depict the influence of low-quality maternal interaction on the child’s ANS regulation, in calm and more stressful play situations. The overall higher SNS mode with impaired PNS reactivity may negatively influence child’s ANS homoeostasis, which may result in a long-term impact on mental and physical wellbeing. Further, the maternal sensitivity may function as a buffer for the stress response of their child. These results could serve as a basis for the development of appropriate psychoeducational programs for mothers of low sensitivity in their interaction with the child. The first years of a child’s life represent an essential time for the development of behavioral and physiological regulatory processes associated with healthy socio-emotional and cognitive development . One of Various theoretical perspectives suggest that sensitive and responsive caregiving, defined by an accurate interpretation and prompt response to infant needs, can facilitate the organization of the response systems of infants and children in favor of a regulation that supports processes of more complex social, emotional, and cognitive development .The external regulation of children’s regulatory processes becomes particularly relevant in stressful situations if children have not yet developed a sufficient repertoire of regulatory capacities. There seems to be overwhelming agreement that children who experience a predominantly sensitive and responsive caregiving demonstrate healthier patterns of physiological and behavioral strategies both at rest and when facing a challenge or stressor .Only few research has focused on the quantification of physiological parameters during mother–child interaction . In thisThe ANS, controlled by the central autonomic network, is mainly involved in emotional and behavioral reactions initiated by an attachment stressor. Therefore, it is particularly relevant to investigate emotion regulation during infant development and in different psychopathologies .via the ANS  and the parasympathetic nervous system (PNS)—which consist of two different anatomical structures with distinct pathophysiology . Thus, aFurther, RSA has been widely measured in early childhood differing from calm vs. challenging situations . RSA is via a sharp change in heart rate  by focusing on psychological, biological, and social factors in a prospective design. The study was funded by the Federal Ministry of Education and Research  and approved by the Ethics Committee of Ulm University, performed in accordance with relevant guidelines and regulations. The main aim was to identify mechanisms of stress resilience and processes in the transgenerational transmission of CM on different levels: psychological, biological, and social. Because this is a longitudinal study, data from this study have been analyzed with respect to ANS activity in previous articles: These are From October 2013 to December 2015 1,460 healthy women in the maternity unit of the women’s hospital of the University Hospital of Ulm were asked for study participation 1–6 days after parturition of a healthy child. Based on exclusion criteria , 533 mothers signed an agreement for participation.All participating mother–child dyads were then invited for three follow-up time points . 240 mothers participated in a laboratory and home visit (t1) 3 months post-partum, 158 mother–child dyads took part at a laboratory and home visit around 12 months (t2), and 152 between 18 to 36 months (t3) of child’s age. t3 was the measurement point, where the data for this paper origin from. Sample attrition from t1 to t3 was attributed to personal reasons, lack of interest, and time conflicts for carrying out observations.M = 36.6 years (SD = 4.1 years)] and child’s age varied between 1.6 years and 3.7 years [M = 2.9 years (SD = 0.1 years)]. The level of education showed that 72.5% of the mothers had completed a grammar school degree (13 years of school), 22.5% a secondary school degree (10 years of school), and 5.0% a basic secondary school degree (9 years of school). Furthermore, 63.7% had achieved at least a bachelor’s degree at a university or a university of applied sciences.Mothers’ age at measurement point t3 was between 27 and 46 years  and RSA showed a positive association with child age in E1  and E3 .HR was negatively related to child’s age across all three episodes  and E2 , and showed an inverse correlation in E3 . LVET showed a positive correlation with maternal sensitivity in E1 , E2 , and E3 . Mother’s age correlated negatively with their HR in E2  and E3.RSA was found to be positively correlated with maternal sensitivity in E1 [F = 2.03, p = 0.141, F = 0.00, p = 0.951, F = 0.12, p = 0.863, When comparing children’s HR across all episodes between sensitive and insensitive mothers, the main time, group and group x time interaction effects were not significant [F = 0.70, p ‚ = 0.498, F = 5.10, p = 0.027, F = 34.09, p < 0.001, Whereas the main time effect of children’s RSA across all episodes between sensitive and insensitive mothers was not significant [F = 0.70, p = 0.489, F = 36.43, p < 0.001, F = 0.29, p = 0.747, The main time effect was not significant [F = 0.40, p = 0.669, F = 0.05, p = 0.839, F = 1.06, p = 0.350, When comparing children’s PEP values, there was no significant effect over time [No significant results were detectable in the ANCOVA comparing mother’s HR, RSA, LVET, or PEP across the episodes between sensitive and insensitive mothers. The results are shown in Our results show that children of insensitive mothers had lower overall RSA scores, indicating lower PNS activity and difficulties in maintaining the homeostatic state. This result is consistent with current literature , which ivia the PNS, whereas children of insensitive mothers more via the SNS. Since the PNS is responsible for a person’s regeneration and recovery, and the SNS is responsible for maintaining constant alterness and preparation for potential threats ; BMBF, 2013–2016, additional interim funding 2017.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Quantifiable biomarkers may be useful for a better risk and frailty assessment of patients referred for transcatheter aortic valve implantation (TAVI).To determine if adiponectin serum concentration predicts all‐cause mortality in patients undergoing TAVI.77 consecutive patients, undergoing TAVI, were analyzed. The CT axial slices at the level of the fourth lumbar vertebra were used to measure the psoas muscle area, and its low‐density muscle fraction (LDM (%)). To assess the operative risk, the STS  score, Log. Euroscore, and Euroscore II were determined. A clinical frailty assessment was performed. ELISA kits were used to measure adiponectin serum levels. We searched for a correlation between serum adiponectin concentration and all‐cause mortality after TAVI.p = .004). In the multivariable model, including STS score, vascular access of TAVI procedure, LDM (%), and adiponectin serum concentration, serum adiponectin level, and LDM(%) were independent predictors of all‐cause mortality after TAVI . Adiponectin level was a predictor of all‐cause mortality in females and males .The mean age was 80.8 ± 7.4 years. All‐cause mortality occurred in 22 patients. The mean follow‐up was 1779 days (range: 1572–1825 days). Compared with patients with the lowest adiponectin level, patients in the third tertile had a hazards ratio of all‐cause mortality after TAVI of 4.155 (95% CI: 1.364–12.655) (Adiponectin serum level is an independent and incremental predictor of all‐cause mortality in patients undergoing TAVI. ASaortic stenosisAVAaortic valve areaCTcomputed tomographyLDM(%)low‐density muscle expressed as a percentage of the PMAPMAP[soas muscle areaSTS‐scoreSociety of Thoracic Surgeons Predicted Risk of Mortality scoreTAVItranscatheter aortic valve implantation1Over the last two decades, transcatheter aortic valve implantation (TAVI) has evolved from an experimental procedure to an appropriate treatment of symptomatic aortic stenosis in patients at high operative risk.3, logistic EuroSCORE,It has been demonstrated that conventional clinical cardiac surgery risk assessment, by EuroSCORE IIPlasma adiponectin has been proposed as a potential biomarker of frailty.Therefore, this study sought to determine whether plasma adiponectin levels could predict all‐cause mortality after TAVI. We tested the ability and relative strength of adiponectin levels to predict all‐cause mortality after TAVI in a multivariable model, that included clinical operative risk assessment, frailty assessment, vascular access of the TAVI procedure, CT imaging variables (LMD (%)) and adiponectin serum concentration.22.1Between January 2014 and June 2016, patients referred for TAVR were included in the analysis. The recruitment of patients to this study partially overlapped with another ongoing frailty study.The protocol was approved by the local research ethics board. The study has been carried out in accordance with The Code of Ethics of the World Medical Association (Declaration of Helsinki).2.1.1The UZAntwerp TAVI registry is a comprehensive electronic database of patients undergoing TAVI. Patient baseline characteristics, TAVI procedural information, and outcome data  were retrieved from the registry Table . In addi2.2The CT scanning protocol has been previously described.All CT scans were performed on a dedicated CT scanner . Images were acquired using 80–120 kVp and automated tube current modulation.A triphasic intravenous radiology contrast (Iomeron 350) injection protocol was used: 80 ml contrast agent at 4 ml/s, followed by 60 ml contrast agent at 4 ml/s plus 60 ml 0.9%NaCl solution at 2 ml/s, followed by 28 ml 0.9% NaCl solution at 2 ml/s).2.3The acquisition protocol of the psoas muscle variables has been described previously.2.4The serum concentration of adiponectin was measured using an enzyme‐linked immunosorbent assay (ELISA) from blood samples from the TAVI‐biobank. The Human HMW Adiponectin/Acrp30 Quantikine® ELISA‐kit (R&D Systems) was used to determine the serum adiponectin concentration.The adiponectin concentration was measured twice, and the results were averaged. The coefficient of variation (CV), which is the ratio of the standard deviation/mean of both measurements, was calculated. The results of the adiponectin serum concentration measurements were rejected if the CV exceeded 15%.2.5t‐test and Chi‐square tests were used to compare categorical variables.Continuous variables are presented as mean ± standard deviation, and categorical variables are presented as frequencies. Continuous variables were compared using Student's Univariable Cox hazard analyses were performed to determine the hazard ratios of the clinical frailty assessment variables, the operative risk stratification variables, vascular access of the TAVI procedure, LDM (%), and adiponectin serum concentration for all‐cause mortality after TAVI.p‐value ≤ .2 in the univariable analysis. If several operative risk stratification variables or clinical frailty variables were eligible for inclusion in the multivariable analysis, the variable with the lowest p‐value was selected to represent perioperative risk and clinical frailty assessment, respectively. Multivariable Cox hazard analyses were performed to determine the hazard ratios of the adiponectin levels for all‐cause mortality after TAVI. Subgroup analyses were performed to test whether adiponectin serum concentration predicts all‐cause mortality after TAVI in both sexes. Risk‐adjusted Kaplan Meier analysis was performed for adiponectin tertile groups.The aforementioned variables were eligible for inclusion in the multivariable analysis if their Receiver operating characteristic (ROC) curves of Adiponectin concentration were performed to calculate the area under the curve.p < .05.Pearson correlations were used to test for a correlation between the Barthel frailty score and LDM%. All analyses were performed in SPSS Version 25 (IBM), and statistical significance was defined as 377 patients met the inclusion criteria for the study. However, six patients were excluded from the analyses involving LDM(%) due to insufficient CT data.3.1p = .015).Thirty‐four (44.2%) patients were male Table . The mea3.2Patients were followed for a mean of 1779.0 ± 81.2 days. The median follow‐up was 1825. Twenty‐two deaths occurred during the follow‐up period.3.3p < .05). A trend toward a statistically significant difference between survivors and nonsurvivor was observed for age, LVEF, vascular access route of TAVI‐procedure .Statistically significant differences in sex, hospital stay, operative risk assessment score, and psoas muscle assessment were observed between patients with and without mortality after TAVI Tables , p < .05p = .273, .246, .257, and .953, respectively).Frailty scores  failed to demonstrate statically significant differences between survivors and non‐survivors .The mean serum adiponectin level was 9733.6 ± 7076.3 ng/ml and 7866.6 ± 5847.7 ng/ml in females and males, respectively (3.5p = .526). A subgroup analysis in males and females failed to show a correlation between adiponectin serum level and LDM(%) .No correlation between adiponectin serum level and LDM(%) was found . The multivariable Cox regression survival model, including STS score, vascular access of TAVI procedure, LDM(%) tertiles, and adiponectin tertiles, demonstrated that adiponectin concentration (p = .017) and LDM(%) (p = .042) are independent predictors of reduced survival after TAVI. Compared to patients in the conjugated first and second adiponectin serum level tertile, patients in the third tertile had a hazard ratio (HR) of 4.6995 (95%CI: 1.055–20.924) (p = .004)  Table .p < .001)  curves of adiponectin were performed, with an area under the curve (AUC) of 0.730 (95%CI: 0.60–0.86) Figure .3.6.1p = .012) and 11.549(95%CI: 1.388–96.062) (p = .024), respectively. Figure p = .007 and 0.004, respectively).In both sexes, adiponectin level was associated with survival. Compared to males and females within the conjugated 1st and 2nd adiponectin concentration tertile, males and females within the third tertile had a hazard ratio of death of 3.761 (95%CI: 1.346–10.509) , and LDL cholesterol to the multivariable model did not change the overall conclusion that adiponectin is an independent predictor of all‐cause mortality after TAVI Table .p = .997).A post‐hoc binary logistic analysis, using the 30‐days mortality after TAVI as the marker for TAVI‐procedure‐related mortality, failed to demonstrate an association between adiponectin level and 30‐days mortality after TAVI , vascular access of the TAVI procedure, and a CT‐derived psoas muscle attenuation variable (as a surrogate marker for sarcopenia).Nevertheless, it should be noted that our study results do not align well with the current insight that adiponectin has mainly cardioprotective effects, as patients within the highest serum adiponectin concentration had the highest all‐cause mortality.On the other hand, our study results are consistent with the findings from frailty studies, demonstrating that adiponectin levels are elevated in elderly cognitively impaired patients.p = .012) and 11.549 (p = .024) respectively, compared to their counterparts in the combined first and second tertile.In contrast to earlier frailty studies, showing sex differences between adiponectin levels, frailty, and mental status, our results confirmed a strong and independent association between adiponectin serum concentration before TAVI and all‐cause mortality after TAVI in both sexes. Males and females in the third adiponectin serum level had a hazard ratio of death of 3.761  has been proposed as a surrogate marker for sarcopenia. Several studies have demonstrated that PMA is a predictor of mortality after TAVI.Given that adiponectin plasma concentration and LDM% were proposed to represent frailty and sarcopenia, we tested for a correlation between both variable but did not demonstrate one. This may suggest that adiponectin better represents other aspects of frailty than sarcopenia.Selecting the variables for our multivariable model was challenging. Our model builds further on research by others. Historically, studies confirming an association between clinical operative risk assessment, using the EuroSCORE II, the Logistic EuroSCORE, or the STS score were performed first.Our study results demonstrated that hospitalization duration is a predictor for all‐cause mortality after TAVI. We deliberately did not include hospital stay in our multivariable model as this variable does not contribute to the clinical decision‐making before TAVI implantation.The vascular access route of TAVI was an independent predictor of all‐cause mortality in the univariable analysis but failed in the multivariable analysis. Given the anti‐inflammatory and anti‐atherosclerotic properties attributed to adiponectin, a correlation between adiponectin level and arterial vascular obstruction could be hypothesized. However, our study sample is too small to elaborate on this hypothesis further. Nevertheless, our study could not demonstrate an association between the adiponectin serum concentration and the TAVI‐procedure‐related 30‐days mortality.Age, sex, and cachexia have been associated with mortality after TAVI.4.1Three study limitations need to be discussed. First, it could be argued that the small sample size of our study may have altered the study results. On the other hand, the ability of adiponectin levels to predict all‐cause mortality after TAVI in this limited study population indirectly proves the prognostic strength of serum adiponectin levels. Our sample size is small with 77 patients being included in the analysis. The finding of an increased risk for those in the highest tertile of adiponectin is hypothesis‐generating, and further large‐scale prospective studies are required to validate our observations.All‐cause mortality was used as the outcome variable in our study. This reflected the fact that frailty may lead to death through multiple causes, for example: fracture hip, aspiration pneumonia, and head injury. In this small sample size, further analysis of the potential effects of adiponectin on other causes of mortality such as cardiovascular events or sepsis was not possible.Adiponectin levels may have been influenced by multiple metabolic factors, such as lipid profile, diabetes, smoking status, and medication use. These potential confounders could only partially be extracted from the database. To mitigate this limitation, diabetes, body mass index (BMI), and LDL cholesterol were introduced in a supplemental multivariable model . The overall conclusion that adiponectin is an independent predictor of all‐cause mortality after TAVI remained unchanged. It should be emphasized that strong predictors of mortality after TAVI, such as age and sex are part of the STS Score and thus included in the multivariable model.5Our study, with a mean follow‐up of 1779 days, demonstrates that adiponectin serum concentration is an independent and incremental predictor of all‐cause mortality after TAVI. Further prospective studies are warranted to determine the role of serum adiponectin levels to predict frailty and outcomes in patients with severe AS, who are being considered for TAVI.HH: Dr. Heidbuchel did not receive any personal honoraria. He received unconditional research grants through the University of Antwerp and/or the University of Hasselt from Bayer, Boehringer‐Ingelheim, Bracco Imaging Europe, Abbott, Medtronic, Biotronik, Daicchi‐Sankyo, Pfizer‐BMS, and Boston‐Scientific, all outside the scope of this study. JB: Dr. Bosmans is a proctor for Medtronic CoreValve.Supplemental Figure 1 demonstrates the calculation of the portion of Low‐density muscle (LDM%). The slice portraying the 4th lumbar vertebra is selected. Panel A shows the manually contoured psoas area. Subsequently, the filter selects the voxels with Hounsfield Units (HU), ranging between 0 and 29 HU, attributed to LDM (Panel B).Click here for additional data file.Supplemental Figure 2 shows the Kaplan Meier survival curves for adiponectin serum levels in males and females . Compared to males and females within the conjugated 1st and 2nd adiponectin concentration tertile (green curves), males and females within the third tertile (green curves) had a hazard ratio of death of 3.761 (95%CI: 1.346 – 10.509) (p = 0.012) and 11.549(95%CI: 1.388 −96.062) (p = 0.024), respectively.Click here for additional data file.Supplementary information.Click here for additional data file.