While an association between postoperative acute kidney injury (AKI) and adverse events exists, the incidence and impact of postoperative AKI after pancreaticoduodenectomy for pancreatic ductal adenocarcinoma remain unclear. This study aimed to diagnose AKI and investigate the risk factors for and prognostic value of postoperative AKI.Clinical characteristics of patients who underwent pancreaticoduodenectomy between 2013 and 2020 at a high‐volume centre were collected retrospectively. The Kidney Disease Improving Global Outcomes criteria were used to diagnose AKI. A 1:2 propensity score matching (PSM) was used to minimise bias between the AKI and non‐AKI groups. Short‐term surgical and long‐term survival outcomes were compared between groups. Multivariate logistic regression analysis assessed the independent risk factors for AKI development, major complications, and 30‐day mortality.p < 0.001), estimated glomerular filtration rate < 60 ml/min/1.73 m2 , and intraoperative estimated blood loss >1000 ml  were independent risk factors for postoperative AKI. After PSM, higher incidences of major complications (p < 0.001) and 30‐day mortality (p < 0.001) were observed in the AKI group than in the non‐AKI group. There was no difference in long‐term overall survival outcomes between both groups (p = 0.535). AKI was an independent predictor of major complications  and 30‐day mortality .Postoperative AKI occurred in 10.7% of 1312 patients. Total bilirubin level > 250 μmol/L (odds ratio [OR]: 3.24; Postoperative AKI is common after pancreaticoduodenectomy for pancreatic ductal adenocarcinoma and has a predictive effect on major complications and 30‐day mortality. Therefore, prevention and proper management of postoperative AKI are required in clinical practice. Acute kidney injury is an independent predictor of major complications and 30‐day mortality after pancreaticoduodenectomy for pancreatic ductal adenocarcinoma. Using different diagnostic standards, the prevalence of postoperative AKI varied greatly. In 2012, the Kidney Disease: Improving Global Outcomes (KDIGO) Foundation proposed a widely used guideline that accepted the existing criteria for defining AKI and implemented simplified criteria for practice.In routine clinical managements after PD, surgeons pay the most attention to high‐risk complications, such as fistula, haemorrhage and infections. Postoperative AKI sometimes can be covert and easily ignored, with only slight symptoms or few level changes in biochemical indicators. There were also few studies highlighting the significance of AKI after PD surgeries. We assumed that the occurrence could increase the risk of postoperative morbidity and mortality, and this study aimed to study its risk factors and impacts on both short‐term surgical and long‐term survival outcomes.22.1Data from patients who underwent the classic Whipple procedure with standard lymphadenectomy for PDAC between 1 January 2013 and 31 December 2020 at our institute were retrospectively collected, and the tumor pathology was confirmed. We excluded patients younger than 18 years and those with pre‐existing pancreatic surgery, evidence of metastasis, locally advanced or borderline unresectable tumors, and additional organ resection (such as the kidney or colon).[DGE], and intra‐abdominal infection) and postoperative hospital stay were also recorded to identify short‐term outcomes.Preoperative clinical data included patient demographics , preoperative laboratory data , and preoperative biliary drainage status. Intraoperative data included the surgical type (open PD or robot‐assisted PD), venous resection , operative time, estimated blood loss, intraoperative transfusion, and stage (T or N stage). Postoperative surgical complications , the time from PD to death attributable to any cause. Survival data were censored at the date of the last follow‐up, and our latest follow‐up occurred in March 2022.2.42, regardless of the medical history.The KDIGO system was used during this study. A blood test was conducted on the admission day to define baseline SCr values,2.5t‐test when they are normally distributed or medians (interquartile ranges) using the Mann–Whitney U test when they are not. Univariate independent risk factors of postoperative AKI, major complications, and 30‐day mortality were first analysed, and those with p < 0.05 were entered into the multivariate logistic regression model, and we recorded the odds ratio (OR) and 95% confidence interval (CIs). Furthermore, preoperative and intraoperative risk factors were analysed for AKI development; AKI status was further included in the analysis for major complications, and 30‐day mortality also included postoperative complications. The entire AKI and non‐AKI cohorts were unbalanced. Patients who developed AKI might have been older and had worse eGFRs, and other different baseline characteristics, leading to increased morbidity and mortality. To reduce selection bias and make the AKI status the only exposure, we used R software (R Core Team), SPSS Statistics Essentials for R, and SPSS PS Matching to perform 1:2 nearest‐neighbour propensity score matching (PSM); calliper widths with a standard deviation of 0.05 were allowed. PSM was created using the following baseline characteristics: age, sex, body mass index, ASA class, diabetes mellitus, hypertension, cardiovascular disease, haemoglobin level, TBil level, albumin, baseline SCr level, eGFR, surgical type, venous resection, operative time, intraoperative estimated blood loss, and transfusion. After PSM, all preoperative and intraoperative factors were well‐balanced between the control and observational groups, and a direct causal relationship of AKI and morbidities was analysed. Subsequently, survival curves were constructed using the Kaplan–Meier (KM) method with the log‐rank test, and OS was analysed after PSM. Patients who died within 30 days postoperatively or were lost during regular follow‐up were excluded from the KM analysis.Statistical analysis was performed using SPSS version 26.0 (IBM Corp.). All the tests with a two‐tailed P < 0.05 were considered significant. Categorical variables were presented as numbers (percentages) and compared using the Pearson chi‐square or Fisher exact test. Continuous variables were presented as means ± standard deviations using Student's 33.1p = 0.002; 11 [9.8%], 7 [33.3%], and 3 [37.5%], respectively, for major complications, p = 0.004). The results revealed no significant association between 30‐day mortality and major complications and the time of postoperative AKI occurrence.During the 8‐year study period, 1312 patients were included according to our selection criteria; 141 (10.7%) developed postoperative AKI. Among the entire AKI group, the AKI severity stage was stage 1 for 112 cases (79.4%), stage 2 for 21 (14.9%), and stage 3 for 8 (5.7%). According to the treatment data, no patient with stage 3 was admitted to the intensive care unit or underwent renal replacement therapy. All AKI patients underwent only clinical observation or fluid control therapy. Overall, 111 (78.7%) of the AKI cases occurred during the first 2 days after PD, and the number continued to decrease to 6 (4.3%) during the final 2 days within a week. Thirty‐day mortality and major complications were significantly correlated with the AKI severity stages 1, 2, and 3 , had a higher ASA class (p = 0.003), and were more likely to have a history of cardiovascular disease than those without postoperative AKI (p = 0.008). They also had lower haemoglobin and serum albumin levels and higher white blood cell counts and eGFR than those without postoperative AKI. Furthermore, the preoperative biliary drainage status significantly differed , which was consistent with the higher TBil level of those with postoperative AKI compared to those without (p < 0.001). There were no significant differences in patient characteristics such as sex proportion, body mass index, comorbidities including diabetes mellitus and hypertension, smoking and alcohol habits, baseline SCr level, surgical type, venous resection, operative time, estimated blood loss, intraoperative transfusion, and tumor stage between both groups. After PSM, a well‐balanced cohort was created, with the non‐AKI group (228 patients) as the control group and the AKI group as the observational group (114 patients). Preoperative and intraoperative characteristics are presented in Table In the entire cohort, patients with postoperative AKI were older  and major complications  were significantly different between groups. Patients also experienced an increased incidence of CR‐POPF , especially grade C, BL , IL , PPH grades B and C , and intra‐abdominal infections  when postoperative AKI occurred. The reintervention rates during the hospital stay  also differed between groups.Comparisons of postoperative short‐term outcomes are presented in Table 3.42, preoperative biliary drainage, operative time > 300 min, and intraoperative estimated blood loss >1000 ml were significantly associated with AKI development. During the multivariate logistic regression analysis, however, only a TBil level > 250 μmol/L , eGFR <60 ml/min/1.73 m2 , and estimated blood loss >1000 ml  were independent risk factors for postoperative AKI.We conducted univariate and multivariate analyses to determine potential risk factors for postoperative AKI development in patients who underwent PD Table . During 3.5p < 0.001), diabetes mellitus, comorbidities, and white blood cell count >10 × 109/L were associated with major complications. After the multivariate regression, only AKI  and diabetes mellitus  remained independent predictors for major complications.Risk factors are displayed in Table 9/L, and haemoglobin level < 90 g/L), intraoperative variables (such as operative time > 300 min), postoperative AKI status, and postoperative complications . The multivariate logistic regression analysis revealed that age ≥ 70 years  was the only independent preoperative risk factor and that postoperative complications such as CR‐POPF and BL were not significantly associated with 30‐day mortality. Postoperative complications such as IL , PPH grades B and C , and intra‐abdominal infections  still showed significant differences. Furthermore, postoperative AKI  was a new independent risk factor for 30‐day mortality.During the univariate analysis, 30‐day mortality after PD was significantly related to preoperative characteristics  in the long‐term OS of the AKI and non‐AKI groups after PSM  patients died, and 6 were lost. During a median follow‐up of 17 months, the KM analysis revealed no difference ; data curation ; formal analysis ; methodology ; writing – original draft . Yiran Zhou: Conceptualization ; formal analysis ; methodology ; writing – review and editing . Ziyun Shen: Data curation ; formal analysis ; writing – review and editing . Haoda Chen: Formal analysis ; methodology ; validation . Shulin Zhao: Data curation ; resources ; software ; validation . Xiaxing Deng: Conceptualization ; project administration ; resources ; supervision . Baiyong Shen: Conceptualization ; methodology ; project administration ; supervision ; writing – review and editing .The authors have no conflicts of interest to declare.None.This study conformed to the provisions of the Declaration of Helsinki (as revised in 2013). All the patients included signed informed consent forms and agreed to data collection. It was approved by the institutional review board of Shanghai Ruijin Hospital.Table S1.Click here for additional data file.


Low serum cobalamin concentrations have been associated with ileal malabsorption in dogs with chronic enteropathy. Increased serum methylmalonic acid (MMA) concentrations indicate cobalamin deficiency on a cellular level. Few studies have evaluated serum cobalamin concentrations or methylmalonic acid concentrations in juvenile dogs with parvoviral enteritis or nonparvoviral acute enteropathies.Evaluate serum cobalamin and methylmalonic acid concentrations in juvenile dogs (6 weeks to 10 months old) with parvoviral enteritis or nonparvoviral acute enteropathy.Thirty‐one juvenile dogs with parvoviral enteritis, 29 dogs with nonparvoviral acute diarrhea (NPVAD), and 40 healthy juvenile control dogs.Single‐center, prospective, observational, cross‐sectional study. Serum cobalamin and, when sufficient serum was available, MMA concentrations were measured.P = .0002; median, 463 ng/L; range, <150‐10 000 ng/L) or dogs with NPVAD . Serum MMA concentrations were not significantly different between groups .Most serum cobalamin concentrations were within the adult reference interval. Serum cobalamin concentrations in healthy dogs  were significantly higher than in dogs with parvoviral enteritis (Juvenile dogs with parvoviral enteritis or NPVAD had lower serum cobalamin concentrations than healthy juvenile dogs. However, based on serum MMA concentrations cellular cobalamin deficiency was not apparent. CPVcanine parvovirusMMAmethylmalonic acidNPVADnonparvovirus acute diarrheaRIreference interval112, is a water‐soluble vitamin required for DNA synthesis and cellular energy production.Cobalamin, also known as Vitamin BDogs affected by Imerslund‐Gräsbeck syndrome  can develop blood cell dyscrasias , central and peripheral neurologic signs, and gastrointestinal (GI) signs such as anorexia, vomiting, diarrhea, lethargy, and failure to thrive.Cobalamin is an essential cofactor of the enzyme methylmalonyl‐CoA mutase that converts L‐methylmalonyl‐CoA to succinyl‐CoA.Enteritis caused by canine parvovirus type 2 is the world's most common infectious disease of dogs.Given the destructive effects of parvovirus on the intestine, a deficiency in cobalamin because of ileal malabsorption is possible. If so, cobalamin supplementation theoretically could be beneficial in affected dogs. At the same time, other acute processes can cause intestinal disease in puppies, and may or may not have a similar effect on cobalamin status. Although a previous study identified decreased serum cobalamin concentrations in dogs with parvoviral enteritis compared to healthy dogs,Therefore, we designed a prospective study to investigate serum cobalamin and MMA concentrations in juvenile dogs with parvoviral enteritis, dogs with nonparvoviral acute diarrhea (NPVAD), and juvenile healthy control dogs. It was hypothesized that serum cobalamin concentrations in dogs with any enteropathy would be lower than in healthy dogs and would be associated with an increase in serum MMA concentrations, indicating cellular cobalamin deficiency.2Ours was a single‐center, prospective, observational cross‐sectional study. Dogs between 6 weeks and 10 months of age that presented to the Texas A&M Veterinary Medical Teaching Hospital Small Animal Emergency Service or Primary Care service between October 2019 through June 2022 were eligible for inclusion. The study was approved by the Institutional Animal Care and Use Committee (2019‐0258). Written consent was provided by the owner of each dog before enrollment into the study.Eligible dogs had to weigh at least 2 kg, have had at least 2 episodes of diarrhea with or without vomiting within the last 12 hours or diarrhea with or without vomiting or inappetence lasting over 24 hours, and a commercial, rapid, patient‐side canine parvovirus (CPV) SNAP ELISA  performed at the facility or at a referring veterinarian's practice. These dogs then were divided into 2 groups. One group included dogs that tested positive on ELISA, called the CPV group. The other group consisted of dogs that tested negative on ELISA, called the nonparvovirus acute diarrhea (NPVAD) group. Fecal samples were required for inclusion into the latter group for fecal CPV PCR testing. Dogs that tested positive on PCR were considered part of the CPV group. Dogs that were diagnosed with a foreign body or intussusception after the study were excluded. Dogs with diarrhea lasting >3 weeks were excluded from the study. Age, breed, sex, reproductive status, vaccination status, and duration of reported clinical signs were recorded for each dog.Clinically healthy dogs >2 kg between the ages of 6 weeks and 10 months that presented for routine vaccination or an elective surgical procedure were included as controls. Dogs in this group were considered healthy based on history and physical examination findings with no evidence of vomiting or diarrhea within the 72 hours before presentation. After sample collection, each dog was treated for its primary reason for presentation at the discretion of the attending clinician.2.1Whole blood was collected from each dog and stored in no‐additive collection tubes at 4°C within 24 hours of presentation. If not processed for cobalamin measurement within 8 hours, whole blood was centrifuged within 24 hours and serum stored at −20°C.When required, fecal samples were collected using swab, digital rectal examination, or floor collection and stored in plastic containers at 4°C. Real‐time PCR was performed on the samples for CPV DNA as previously described.2.212, Immulite 2000; Siemens Healthcare Diagnostics). The working range was from 150 to 1000 ng/L, with a reference interval (RI) of 251 to 908 ng/L. When the cobalamin concentration exceeded the upper detection limit, the serum sample was diluted and rerun to obtain a numerical value. Results >10 000 ng/L were entered as 10 000 ng/L for statistical analysis. The surplus serum for MMA measurement was stored at −80°C. Serum MMA concentration was measured using the stable isotope dilution gas chromatography‐mass spectrometry method in batches as described previously.Serum cobalamin measurements were performed in all dogs within 48 hours of blood collection using a solid‐phase competitive chemiluminescent immunoassay . Differences in serum cobalamin and MMA concentrations between groups were assessed using Kruskal‐Wallis tests followed by Dunn's post‐test as appropriate. The correlation between serum cobalamin and MMA concentrations was assessed using Spearman's rank correlation coefficient. Statistical analysis was performed using a commercial software package . Values of 3During this period, 102 dogs with acute GI signs were screened. One dog was excluded because it tested positive on SNAP CPV, but was negative on concurrent fecal PCR. This patient was treated for CPV and recovered but was excluded from the analysis because of the discordant test results. Another dog was excluded for diarrhea lasting >3 weeks. One hundred dogs were included in the analysis. No dogs were diagnosed with foreign bodies or intussusception. Forty healthy dogs also were enrolled.3.13.1.1Thirty‐one dogs were enrolled in the CPV group. Twenty‐four dogs tested positive on a SNAP ELISA. Seven dogs tested negative on SNAP testing but tested positive on fecal PCR. All 7 dogs were reported to have previous vaccinations, ranging from receiving 1 vaccine 1 day before presentation (2 dogs) to finishing 3 rounds of vaccinations 2 months before (1 dog). Four of these dogs had clinical signs consistent with canine parvovirus  and were hospitalized. The remaining 3 dogs were treated on an outpatient basis and were all doing well at the time of the follow‐up call regarding the positive PCR result. All 3 dogs had a parvovirus vaccine administered at least 7 days before.One dog tested negative for both SNAP CPV and fecal PCR but tested positive on PCR run on jejunal tissue taken on necropsy. This dog was included as part of the CPV group.Thirteen dogs presented with signs of inappetence in addition to reported diarrhea. Thirteen dogs had signs of vomiting, diarrhea, or inappetence lasting for >1 day before presentation, with the longest duration of signs being 4 days.American Pit Bull Terriers (5) and mixed breeds (4) were the most common breeds represented. The remaining breed distribution was as follows: German Shepherd (3), Golden Retriever (3), Australian Shepherd (2), Doberman Pinscher (2), Labrador Retriever (2), Black Mouth Cur, Blue Heeler, Border Collie, Boxer, Catahoula, Great Dane, Great Pyrenees, Heeler mix, Miniature Poodle, and Walker Hound (1 each). Twelve females  and 19 males  were included. The median age was 3.5 months .3.1.2Twenty‐nine dogs were enrolled in the NPVAD group. Three dogs presented with signs of inappetence in addition to reported diarrhea. Twelve dogs had signs of vomiting, diarrhea, or inappetence lasting for >1 day before presentation, with the longest duration of signs being 3 weeks. This latter dog was diagnosed with a whipworm infection.Mixed breed dogs (6) and Australian Shepherds (5) were the most common breeds represented. The remaining breed distribution was as follows: Blue Heeler (3), American Pit Bull Terrier (2), German Shepherd (2), Labrador Retriever (2), Shih Tzu (2), Anatolian Shepherd, Basset Hound, Border Collie, Maltese, Pembroke Welsh Corgi, Rottweiler, and Samoyed (1 each). Twelve females  and 17 males  were included. The median age was 4.8 months .3.1.3Forty dogs were enrolled in the healthy group. Mixed breed dogs (7) and Golden Retrievers (6) were the most common breeds represented. The remaining breed distribution was as follows: French Bulldog (4), Labrador Retriever (3), Pug (2), Border Collie (2), English Bulldog (2), American Pit Bull Terrier, Australian Shepherd, Basset Hound, Bernese Mountain Dog, Bichon Frise, Catahoula, Chihuahua, Cocker Spaniel, German Short‐haired Pointer, Great Dane, Maltese, Pembroke Welsh Corgi, Pomeranian, and Standard Poodle (1 each). Nineteen females  and 21 males  were included. The median age was 5 months .3.1.4P = .7). A significant difference in age among groups was found; healthy dogs were older than the CPV group . One dog had a serum cobalamin concentration >4000 ng/L, but an exact value could not be determined because of insufficient sample volume for further dilution. Ten dogs had serum cobalamin concentrations <400 ng/L. Of these, 3 dogs had serum cobalamin concentrations below the lower limit of the reference interval.The median concentration in the healthy group was 848 ng/L . Four dogs had serum cobalamin concentrations <400 ng/L. No dog had a serum cobalamin concentration below the lower limit of the reference interval.P = .0002). Most results were within the adult reference interval across all 3 groups. Serum cobalamin concentrations in the healthy dogs were significantly higher than in both the CPV group (P = .0002) and the NPVAD group (P = .02). However, no significant difference was found between the CPV and NPVAD groups (P = .8).Serum cobalamin concentrations were significantly different among all 3 groups . The median serum MMA concentration in the CPV group was 858 nmol/L . The median serum MMA concentration in the NPVAD group was 764 nmol/L . The median concentration in the healthy group was 796 nmol/L . The single dog with a serum MMA concentration of 3424 nmol/L was a Labrador Retriever.Serum MMA concentrations were not significantly different among the groups : 2 from the CPV group, 1 from the NPVAD group, and 5 from the healthy group.3.4No significant correlation was found between serum cobalamin and MMA concentrations  as the diseased dogs, a small but significant difference in age was found between normal dogs and dogs with CPV. Serum cobalamin concentrations in human neonates are different than those of adults. This observation could explain the difference in serum cobalamin concentrations between the healthy dogs and dogs with CPV. A significant difference however was not found between NPVAD dogs and other groups, nor was an overall difference noted between serum cobalamin concentrations in juvenile dogs and adult reference ranges noted in a previous study.Also, we only evaluated serum MMA concentrations as a marker of cellular cobalamin deficiency. Methylmalonic acid is excreted by the kidneys, and serum MMA concentrations are thought to be more stable and have a 40‐fold higher concentration in urine.Other causes for increased serum MMA concentrations include hypovolemia, renal insufficiency, and genetic disorders.Dogs that tested positive on fecal PCR were placed in the CPV group. However, recent vaccination can cause false positives and dogs also can test positive on fecal PCR without clinical signs consistent with parvoviral infection.In conclusion, at the time of presentation, juvenile dogs with parvoviral enteritis or NPVAD had lower serum cobalamin concentrations than healthy juvenile dogs. However, based on serum MMA concentrations, this finding was not associated with cellular cobalamin deficiency. Future studies longitudinally assessing serum cobalamin and MMA concentrations as well as clinical outcome in similar groups of dogs are needed.Drs Lidbury, Steiner, and Sucholdolski are employed by the Texas A&M Gastroenterology (GI) Laboratory, which offers measurement of serum cobalamin and methylmalonic acid concentrations and detection of parvovirus DNA in feces by PCR on a fee‐for‐service basis.Authors declare no off‐label use of antimicrobials.Approved by Texas A&M IACUC, number 2019‐0258.Authors declare human ethics approval was not needed for this study.


Ulcerative colitis, a typical subtype of inflammatory bowel disease, can cause many serious complications. Burdock fructooligosaccharide (BFO), a linear inulin with a purity of 99.439% and a molecular weight of 2345 Da, demonstrates anti‐inflammatory and immunomodulatory properties.The Kunming mice were divided into two experimental models: a normal pretreatment model and a colitis experimental model. During the experimental treatment period, we assessed changes in weight and disease activity index (DAI), quantified the intestinal index, and determined myeloperoxidase (MPO) activity and reactive oxide species (ROS) levels in colitis mice. We also photographed colon morphology to investigate alterations in the integrity of the intestinal barrier function. Finally, we performed ELISA and qRT‐PCR to evaluate the anti‐inflammatory effect of BFO treatment on colitis mice.The long‐term oral administration of BFO alone exhibited protective effects by preventing disruption of the intestinal functional structure and increasing the colon index in mice. However, in a dextran sodium sulfate (DSS)‐induced colitis mouse model, BFO administration facilitated quick recovery of body weight and effectively reduced the DAI, especially in the BFO‐H group (500 mg/kg/day). BFO treatment maintained the integrity of the intestinal barrier by attenuating the crypt distortion and increasing the goblet cells count It restored the DSS‐induced colon shortening and reduced the symptoms of colitis. These effects may be attributed to the appropriate concentrations of BFO effectively inhibiting MPO activity, clearing excessive ROS, and relieving spleen abnormalitie. BFO also attenuated the overexpression and excessive secretion of inflammatory cytokines  induced by DSS, reduced intestinal inflammation, and consequently protected the intestinal barrier function.BFO effectively alleviated the symptoms of DSS‐induced colitis by mediating anti‐inflammatory effects and protecting the intestinal barrier integrity, thereby potentially facilitating the utilization of safer and more efficacious polysaccharides for managing chronic inflammatory diseases. Burdock fructooligosaccharide (BFO) attenuated the overexpression of inflammatory cytokines induced by dextran sodium sulfate (DSS). BFO treatment prevented the disruption of the intestinal functional structure. BFO can effectively reduce the severity of DSS‐induced colitis. Numerous studies have highlighted the role of biomolecules  from natural extracts in protection of colitis in mice.Arctium lappa L.) belongs to the Compositae family and is consumed as a food and medicinal product,A. lappa, showed anti‐inflammatory activity in the intestine of a TNBS colitis model. Ji et al.A. lappa extracts improved DSS‐induced IBD by enhancing immune responses. However, these extracts were complex mixtures derived from either burdock leaves or roots, thus containing multiple substances. In contrast, BFO utilized in the present study was extracted and purified from burdock root. It belongs to the GFn inulin‐type with a linear chain structure, consisting of d‐fructose and d‐glucose at a molar ratio of 12:1. The linkage among d‐fructose is established by a β (2 → 1) glucosidic bond, while d‐glucose is connected though an α (1 → 2) glucosidic bond at the end of the backbone. The average molecular weight is approximately 2357 Da.A. lappa was found by Wang et al.Burdock ‐induced colitis in mice. DSS, a polysaccharide sulfate widely used to establish UC models, induced clinical symptoms and histological characteristics similar to those observed in human UC.22.1Fresh burdock Root was purchased from Jimo vegetable farm center and identified at School of Life Science, Shandong University. Dextran Sulfate Sodium Salt (DSS) with a molecular weight of approximately 40,000 were purchased from Yuanye Biotechnology Co. Ltd. TRIzol Reagent was purchased from Beyotime Biotechnology. Experimental mice Kunming were provided by Qingdao Daren Fucheng Animal Technology Co. Ltd.2.2The methods for the preparation and purification of BFO were referenced from Hao et al.The crude BFO was initially purified using gel filtration chromatography on a Sephadex G50 (Solarbio) column (1.5 cm × 60 cm) with distilled water flowing at a rate of 0.5 mL/min. The polysaccharide peak from the chromatogram was collected and freeze‐dried for further purification using a Sephacryl S‐200 h column (1.6 × 60 cm) with distilled water flowing at a rate of 0.35 mL/min. Subsequently, the isolated polysaccharide peak was collected and freeze‐dried for purity assessment and activity determination.The purity assay was conducted using a Shimadzu HPLC system equipped with a refractive index detector (RID) and G3000PWXL chromatographic columns . The analysis was performed with ultra‐pure water at a flow rate of 1.0 mL/min. A sample volume was 10 μL with a concentration of 10 mg/mL was injected, and the temperature was 40°C.2.3Kunming mice  were procured from Daren Fucheng Animal Husbandry Co. Ltd, renowned for their robust disease resistance and adaptability, making them extensively employed in various research disciplines such as pharmacology and toxicology. All animals were housed in a pathogen‐free environment with controlled temperature (25 ± 1°C), humidity (55 ± 5%), and photoperiod (12 h light/12 h dark). After acclimatization for 1 week, the mice were divided into two experimental models:n = 10/group) without DSS administration. The CK (control group), pre‐BFO‐L (low concentration pretreatment), pre‐BFO‐M (medium concentration pretreatment), and pre‐BFO‐H (high concentration pretreatment) groups received deionized water and BFO solution at doses of 100, 250, and 500 mg/kg/day respectively. The mice were fed a standard diet throughout the study period. All mice had ad libitum access to drinking water for a duration of 56 days.Normal experimental model ‐ randomly assigned to four experimental groups (n = 10/group) as illustrated in Figure n = 10/group). In the first stage of the experiment, the DSS group (n = 40/group) were fed with DSS water (3% w/v) for 14 days. Subsequently, in the second stage, DSS water was fasted for 14 days to allow for recovery. During this period, mice in the DSS group of the first stage were randomly assigned to four groups , and correspondingly administered deionized water or 100, 250, and 500 mg/kg/day of BFO, respectively. In the third stage, treatment groups  were readministered 3% DSS water for 14 days, while concurrently supplementing the BFO treatment groups with 100, 250, and 500 mg/kg/day of BFO, respectively. The mice were fed a standard diet. All mice were allowed to freely drink the respective solutions for 42 days. The selection of BFO concentration was based on our preliminary experiment and previous research.Colitis experimental model—following the protocol described by Wang et al.2.4n = 10/group). Fecal blood levels were measured using a fecal occult blood qualitative test kit (pyramidon method) (Yuanye Biotechnology Co. Ltd.) according to the instructions. DAI = /3.DAI was determined following the method of Liu et al.2.5At the end of the experiment, the mice were euthanized. Small intestine, cecum, and colorectal were separated and washed with PBS before being drained using a sterile filter paper. Subsequently, the colon in the experimental model of colitis was photographed and measured using a ruler. Furthermore, the weights of the small intestine cecum, colorectal tissue, and entire intestinal tract were measured. The colon in colitis experimental model was photographed and measured using a ruler. The indices of the small intestine, cecum, colon, and whole intestine were calculated as follows: the index of the small intestine or whole intestine = the weight of the small intestine or whole intestine/the weight of the mouse × 100%. The measurements were repeated seven times in each group.2.6The colon tissues from each mouse in the normal and colitis experimental models were sectioned into 5 mm pieces and fixed overnight in a 4% paraformaldehyde solution. Subsequently, the tissue was embedded in paraffin, cut into 5 μm thick sections, and stained with hematoxylin/eosin  for microscopic observation and photography. Each group was evaluated five times.2.7Approximately 20 mg of colon tissues were homogenized in an ice bath to determine the MPO activity according to the instructions of the myeloperoxidase assay kit (Nanjing Jiancheng). The absorbance was measured at 460 nm by a microplate reader . The measurements were repeated three times in each group.2.8ROS in colon tissues was determined using ROS Assay Kit (Nanjing Jiancheng) according to the manufacturer's instructions. Briefly, the cell suspensions obtained from digested colon tissue were collected and incubated with 2′,7′‐dichlorofluorescein diacetate (DCF‐DA) at 37°C for 1 h. Fluorescence intensity analysis was performed using a microplate spectrometer . The measurements were repeated three times in each group.2.9−ΔΔCt calculation method. Values were normalized to the internal control NADPH. Each group was evaluated three times.The experiment was performed by referring to the method described by Song et al.2.10At the end of the experiment, the contents of the tumor necrosis factor‐α (TNF‐α), interleukin (IL)−6, and IL‐10 in serum were determined following the manufacturer's instructions provided with the mouse ELISA kits (Elabscience Biotechnology Co. Ltd.). Protein concentration was measured with a BCA assay kit (Beyotime Biotechnology Industry).The spleens of the experimental mice were collected and weighed to calculate the spleen index according to the following formula: the spleen index = the weight of the spleen (g)/the weight of the mouse (g) × 100%.2.11T‐test was employed to determine significant differences between two samples, while one‐way analysis of variance (ANOVA) with Tukey's HSD test was used for multiple comparison tests. Significant differences between groups were denoted as *p < .05, **p < .01, ***p < .001 versus CK group; and #p < .05, ##p < .01, ###p < .001 versus DSS group based on the findings obtained from at least three independent measurements for each group.The statistical analysis was performed using GraphPad Prism 8.4 software. Experimental results were presented as the mean ± standard deviation. 33.1The BFO eluted from a gel filtration column was collected as a single peak, concentrated, and freeze‐dried. The resulting product was BFO in the form of a white powder Figure . The pur3.2p > .05).Before the administration of DSS, we studied the effects of BFO on the body weight, colon index, and whole intestine index of normal mice. Long‐term administration of different concentrations of BFO  had no significant effect on the body weight of mice  than those observed for the CK group . H&E staining  could better maintain their body weights than those from the DSS group, especially the mice from the BFO‐H group Figure . Further3.4In our study, the colon length was approximately 11.08 cm for normal mice at the end of the experiment. As expected, the colon lengths of the mice from the DSS group were approximately 38.81% lower than those of the mice from the CK group Figure . The colH&E staining results revealed the normal colonic morphology in the mice from the CK group Figure . DSS tre3.5p < .001). Although the inhibitory effect in the BFO‐L group was slightly lower than that in the other BFO groups, it was higher than that observed in the CK group. There was no significant difference between BFO‐M and BFO‐H group as compared to the CK group. The values of BFO‐L, BFO‐M, and BFO‐H groups were approximately 74.57%, 44.65%, and 53.80% higher than that reported for the CK group, respectively.At the end of the experiment, we found that the MPO activity in colon tissues was extremely significantly higher in DSS groups than that in the CK group Figure . HoweverAs shown in Figure p < .05)  Figure . However3.6IL‐1β, IL‐6, TNF‐α, inducible nitric oxide synthase (iNOS), and cyclooxygenase 2 (COX‐2), by qPCR . Interestingly, BFO treatment significantly downregulated the expression of these inflammatory cytokines  induced by DSS (p < .01); the best effect was recorded in the mice from the BFO‐H group. The administration of BFO significantly reduced the upregulation in iNOS expression induced by DSS in a concentration‐dependent manner , respectively. The level of anti‐inflammatory cytokine IL‐10 in the DSS group was 47.39% lower than that in the CK group (p < .01). BFO treatment significantly restored the IL‐10 level, with an approximate increase of 29.91%, 53.80%, and 54.91% in the BFO‐L, BFO‐M, and BFO‐H groups respectively compared to the DSS group  Figure , as wellp Figure .4Numerous studies have reported that the occurrence and development of UC are closely related to the intestinal barrier function and inflammatory response.The DSS‐induced colitis is frequently employed to investigate the pathogenesis of UC, encompassing manifestations such as weight loss, chronic diarrhea, bloody stool, and colon‐shortening.The shortening of colon length is another significant symptom of colitis, which exhibits a positive correlation with the severity of the disease. Moreover, restoration of colon length can be observed upon alleviation of inflammation.p < .001), with values close to those of the CK group. Furthermore, a similar trend was observed in the colon ROS. Wang et al.Neutrophils infiltrating the inflammatory tissue secrete MPO, an indicator of tissue injury and inflammation, which is excessively produced under DSS stimulation.The spleen is the most important lymphatic organ in the body, which can enlarge in response to infection or inflammation.Scutellaria baicalensis Georgi polysaccharide Pro‐inflammatory cytokines play a pivotal role in the progression of DSS‐induced colitis.Xu et al.5In conclusion, administration of BFO alone did not disrupt intestinal function; instead, it exerted a protective effect. In the experimental model of colitis, BFO was found to effectively prevent colon shortening and preserve the function of the intestinal barrier, thus effectively reducing the severity of DSS‐induced colitis by maintaining barrier integrity, inhibiting increased MPO activity, attenuating splenic abnormalities, and reducing the expression and secretion of pro‐inflammatory cytokines. Furthermore, BFO also exhibited superior ability to maintain IL‐10 levels, thus dampening intestinal inflammation. These findings enhance our understanding of the potential benefits of BFO in alleviating UC and provide a theoretical foundation for utilizing BFO in drug development or functional food products aimed at preventing colitis.Qunfei Ma and Xiujuan Zhang performed the research. Kaoshan Chen, Chunyan Liu, and Qiang Chen designed the research study. Yan Lu and Yiru Chen contributed essential reagents or tools. Qunfei Ma, Xiujuan Zhang, and Xuan Xu analyzed the data. Qunfei Ma and Xiujuan Zhang wrote the paper. Chunyan Liu and Kaoshan Chen revised the manuscript. All authors have thoroughly read and approved the final manuscript.The authors declare no conflict of interest.All experiments were performed in accordance with the principles of the National Guidelines for the Nursing and Use of Laboratory Animals and were approved by the Ethics Committee of Shandong University (No. SYDWLL‐2018‐12).


Drug‐induced side effects, particularly serious adverse events (SAEs), often affect cancer patients enrolled in clinical trials. However, little is known about anxiety and depression in cancer patients who experienced SAEs. This study evaluated the prevalence of anxiety and depression in cancer patients enrolled in clinical trials who experienced SAEs and explored the risk factors.A multi‐center, cross‐sectional survey was conducted in hospitals affiliated with the University of Science and Technology of China from December 2021 to November 2022. A total of 112 cancer patients who experienced SAEs while enrolled in clinical trials, and who completed the informed consent process and study questionnaires, were included in the final analysis.The rate of moderate–severe depression in cancer patients was 38.4% and that of moderate–severe anxiety was 13.4%. Among the patients who had moderate–severe anxiety, 93.3% had concurrent moderate–severe depression. Lower cognitive function and lower global quality of life were risk factors for depression in cancer patients who experienced SAEs. Pain, low emotional function, low global quality of life, and a high Impact of Events Scale score were risk factors for anxiety.Cancer patients enrolled in a clinical trial who experienced SAEs tended to be anxious and depressed, particularly the latter. These results indicate the need to evaluate anxiety and depression, and mental health treatment among cancer patients with SAEs in clinical trials. In addition to being physically painful for cancer patients participating in clinical trials, SAEs impose a psychological burden that may result in anxiety or depression.Anxiety and depression subsequent to SAEs decrease the quality of life of cancer patients, which, for those enrolled in clinical trials, can lead to poor adherence and,Emotional distress is a key component of cancer treatment in clinical trials. The American Society of Clinical Oncology (ASCO) has published guidelines for screening and treating depression and anxiety symptoms in cancer patients and has identified risk factors for depressive and anxiety symptoms including sex, marital status, and age, among others.Psychological evaluation is not routinely conducted in cancer clinical trials in China. Reports of anxiety and depression among patients in our center were brought to our attention, where these conditions can affect clinical trials and the rehabilitation of patients.22.1A multi‐center, cross‐sectional survey was conducted in three hospitals affiliated with the University of Science and Technology of China from December 2021 to November 2022. The participants were cancer patients enrolled in oncology clinical trials who had experienced SAEs, were aged ≥18 years, were able to understand and complete the questionnaires independently, and were able to provide informed consent. The exclusion criteria were depression/anxiety or a history of depression/anxiety when enrolled in a clinical trial, neurological problems, refusal to consent, and cognitive impairment of any etiology. A total of 112 cancer patients who had experienced SAEs and completed the informed consent process and questionnaires were included in the final analysis.2.2SAEs among the cancer patients were promptly treated following their occurrence. Between 3 and 7 days later, the researchers obtained demographic information after screening and informed consent, and assisted the participants with respect to the questionnaires. The data collectors in all research centers were trained in a standardized manner, including in terms of the research protocol, questionnaire, and communication skills. The meaning of each question was explained to the participants in detail. The questionnaires were sent to the participants, who completed and returned them. The whole process of receiving, completing, and returning the questionnaires was confidential.The demographic information collected included sex, age, marital status, occupation, education level, living conditions, frequency of acquisition of medical information, whether the individual was participating in a clinical trial for the first time, type of cancer, and other characteristics. Patients were provided with study questionnaires including the Charlson Comorbidity Index (CCI), which is a 19‐item instrument that measures various chronic illnesses and has a total score of 1–6 .The main indicators of depression and anxiety were the PHQ‐9 and GAD‐7 scores. The PHQ‐9 is a nine‐item measure of symptoms of depression.2.3U‐test/Kruskal–Wallis H‐test for non‐parametric variables and Fisher's exact/chi‐squared test for categorical variables. Spearman's correlation analysis was performed to assess the relationships between EORTC QLQ‐C30 and the PHQ‐9/GAD‐7. Associated factors were explored using logistic regression analysis, and were expressed as odds ratios (OR) with 95% confidence intervals (CIs). Multivariate models were constructed using the stepwise selection technique including all potential variables such as demographic characteristics and previously supported risk factors. The ORs and two‐sided P‐values were calculated. A p‐value <0.05 was considered significant for all tests.Descriptive statistics for sex, marital status, income, life events, fatigue, cancer type, depression, and anxiety were generated using SPSS 24 software (SPSS Inc.), which was also used for subsequent analyses. Normality was evaluated in a Kolmogorov–Smirnov test. Demographic characteristics were summarized as a percentage for categorical variables and as the median and inter‐quartile range (IQR) for non‐parametric variables. Demographic characteristics and the CCI, IES, life events, FSI, EORTC QLQ‐C30, and pain intensity numerical rating scale scores were analyzed using a Mann–Whitney 3n = 23 [20.5%]), breast cancer (n = 25 [22.3%]), liver cancer (n = 19 [17.0%]), and gastric cancer (n = 18 [16.1%]). The other cancer types are shown in Table Table n = 39 [90.7%]) was higher in the moderate–severe group than in the none‐mild group (50 [72.5%]) (p = 0.020). The median pain score of the moderate–severe depression group  was also higher than that of the none‐mild depression group . The median IES score was considerably higher in the moderate–severe group (33.0 [22.0–48.0)] than in the none‐mild group , while the mean FSI score was substantially lower in the none‐mild group (14 [7–21]) than in the moderate–severe group . Among patients with moderate–severe depression, 14 (32.6%) also experienced moderate–severe anxiety (Table Table oup (50 7.5%)  had none‐mild anxiety and 15 (13.4%) had moderate–severe anxiety. The median GAD‐7 score was 2 (0–6.5) and 14 (11–14), respectively. Patients in the moderate–severe anxiety group had a higher median pain score (3 [2–4] than those in the none‐mild anxiety group (2 (0–2), The median IES score in the moderate–severe anxiety group (48 [39–60]) was significantly higher than that in the none‐mild anxiety group (22 [12–31]), as was the FSI score: 47 (34–53) versus 18 (9.5–28). In the moderate–severe anxiety group, 14 (93.3%) patients also had moderate–severe depression (Table p = 0.022). Lower cognitive function  and lower global quality of life  were associated with depression in cancer patients who had experienced SAEs. Pain , low emotional function , low global quality of life , and a high IES score  were associated with anxiety in cancer patients with SAEs , role function score , emotional function score , cognitive function score , social function score , and global quality of life score , role function (100.0 [100.0–100.0]), emotional function (100.0 [83.3–100.0]), cognitive function (100.0 [83.3–100.0]), social function (100.0 [66.7–100.0]), and global quality of life (66.7 [66.7–83.3]) scores were significantly higher in the none‐mild than the moderate–severe depression group (00.0), rop < 0.001; Table r = −0.667, p < 0.001), role function , emotional function , cognitive function , social function , and global quality of life , role function (100.0 [83.3–100.0]), emotional function (91.7 [70.8–100.0]), cognitive function (83.3 [66.7–100.0]), social function (83.3 [66.7–100.0]), and global quality of life (66.7 [58.3–83.3]) scores were significantly higher in the none‐mild than in the moderate–severe anxiety group .Psychological evaluation is not a routine part of most oncology clinical trials in China. Our findings demonstrate the need to assess anxiety and depression in clinical trials, especially in cancer patients who have had SAEs. The risk variables determined in this study may aid clinicians in identifying patients who need closer monitoring. Previous studies of cancer patients have shown that education level is related to depression, but that was not the case in this study.This study also revealed that cancer patients with SAEs who did not receive therapy for the first time exhibited considerably higher levels of depression than those undergoing first‐time treatment, probably as a result of the ineffectiveness of prior therapies. Previous research has demonstrated that anxiety and depression are more common in cancer patients who work full‐time, and that both conditions may be made worse by work‐related stress.Similar to previous studies, pain was associated with anxiety and depression in the cancer patients who had experienced SAEs in this study.Poor quality of life can cause various psychiatric disorders.The ASCO guidelines state that in screening cancer patients for anxiety and depression, sex, other chronic medical conditions, and marital status should be taken into account.More than 50% of patients with hematological cancers and gastric cancer had moderate–severe depression, and the incidence of depression in those with lung cancer was higher than in previous studies.This study also had several limitations. Due to its cross‐sectional nature, the causal relationship between SAE and anxiety/depression could not be determined; prospective studies with appropriate control groups are therefore needed. Additionally, there was no follow‐up of the patients, so changes in anxiety and depression were not tracked. According to some studies, continuous monitoring of anxiety and depression is important for accurate assessment thereof.5Cancer patients enrolled in clinical trials are at particular risk of drug‐induced side effects, and thus SAEs. This study examined the anxiety and depression levels of cancer patients who experienced SAEs while participating in oncology clinical trials. The results showed that cancer patients who had experienced SAEs had high rates of anxiety and depression, particularly the latter. The prevalence of moderate–severe depression was noticeably greater than previously reported. Poor quality of life, cancer‐specific stress and pain were associated with depression and anxiety symptoms in patients who had experienced SAEs. It is possible that the risk of anxiety/depression among the patients was due to their expectations about or experiences with the clinical trial, suggesting the need to evaluate anxiety and depression, and education about clinical trials among cancer patients with SAEs.Zhen Peng: Conceptualization (lead); formal analysis (lead); investigation (lead); resources ; writing – original draft (lead); writing – review and editing . Chongwei Wang: Data curation ; investigation (lead); project administration (supporting); resources ; writing – original draft ; writing – review and editing . Yubei Sun: Formal analysis ; investigation ; resources (lead); supervision ; writing – original draft ; writing – review and editing . Yan Ma: Formal analysis (supporting); investigation ; project administration (supporting); resources ; writing – original draft ; writing – review and editing . Jumei Wang: Formal analysis (supporting); investigation ; project administration (supporting); resources (supporting); writing – original draft ; writing – review and editing . Fei Xu: Formal analysis (supporting); investigation ; project administration (supporting); resources ; writing – original draft ; writing – review and editing . Xiaoling Xu: Investigation (supporting); project administration (supporting); resources ; writing – original draft ; writing – review and editing . Yin Chen: Conceptualization (lead); formal analysis ; investigation ; methodology (lead); project administration (lead); resources ; supervision ; writing – original draft (lead); writing – review and editing .The authors declare that they have not competing interests.This study was performed in line with the principles of the Declaration of Helsinki. Approval was granted by the Medical Research Ethics Committee of the First Affiliated Hospital of University of Science and Technology of China .


Background: Coronavirus disease 2019 (COVID‐19) is a pandemic disease caused by severe acute respiratory syndrome CoV type 2 (SARS‐CoV‐2). COVID‐19 is higher in men than women and sex hormones have immune‐modulator effects during different viral infections, including SARS‐CoV‐2 infection. One of the essential sex hormones is progesterone (P4). Aims: This review aimed to reveal the association between P4 and Covid‐19. Results and Discussion: The possible role of P4 in COVID‐19 could be beneficial through the modulation of inflammatory signaling pathways, induction of the release of anti‐inflammatory cytokines, and inhibition release of pro‐inflammatory cytokines. P4 stimulates skew of naïve T cells from inflammatory Th1 toward anti‐inflammatory Th2 with activation release of anti‐inflammatory cytokines, and activation of regulatory T cells (Treg) with decreased interferon‐gamma production that increased during SARS‐CoV‐2 infection. In addition, P4 is regarded as a potent antagonist of mineralocorticoid receptor (MR), it could reduce MRs that were activated by stimulated aldosterone from high AngII during SARS‐CoV‐2. P4 active metabolite allopregnanolone is regarded as a neurosteroid that acts as a positive modulator of γ‐aminobutyric acid (GABAA) so it may reduce neuropsychiatric manifestations and dysautonomia in COVID‐19 patients. Conclusion: Taken together, the anti‐inflammatory and immunomodulatory properties of P4 may improve central and peripheral complications in COVID‐19. Progesterone (P4) inhibits inflammatory signaling pathways and mineralocorticoid receptors (MRs), activates regulatory T cells (Treg)/forkhead box P3 (FOXP3) and glucocorticoid receptors (GRs), leading to activation release of anti‐inflammatory cytokines and inhibition release of pro‐inflammatory cytokines with inhibition development of cytokine storm. These changes inhibit the development of acute lung injury (ALI) and acute respiratory distress syndrome (ARDS), reducing Covid‐19 severity SARS‐CoV‐2 exploits angiotensin‐converting enzyme type 2 (ACE2) and other receptors to bind host cells. SARS‐CoV‐2, when it binds ACE2, persuades a sequence of inflammatory events causing cell injury, hyper‐inflammation, and cytopathic effects.It has been reported that hospitalizations and mortality from COVID‐19 are higher in men than women due to less susceptibility of women to viral infections with low immune response and renin‐angiotensin system (RAS) activity.There is mounting evidence that sex hormones affect RAS and the expression of ACE2 since ovariectomy increases the expression of ACE2 in females.1.1Progesterone (P4) is a sex hormone of endogenous steroids involved in controlling the menstrual cycle, pregnancy, and embryogenesis.P4, in combination with estrogen, is used as hormone replacement therapy in menopause and as contraceptives during the fertility period. George and Willard discovered P4 in 1929, and in 1932 pure P4 was isolated from the corpus luteum. Adolf Butenandt in 1934 described the chemical structure of P4, and it was first prescribed for clinical use in 1934.A) receptors.P4 is regarded as a strong agonist of nuclear progesterone receptor (PR) and membrane progesterone receptors (mPRs). In addition, P4 acts as a ligand for progesterone receptor component 1 (PGRMC1), regulating body metabolism, tumor growth, and neuron viability.P4 has a moderate plasma protein binding activity, about 50%–54% for albumin and 43%–48% for transcortin. P4 is metabolized by hydroxysteroid dehydrogenase in hepatic and extra‐hepatic tissues like the skin and brain. Both 5β‐reductase and 5α‐reductase are the major pathways in the metabolism of P4 into inactive metabolites which are excreted by kidneys.+2 channels, reducing excitotoxicity, upregulation of GABA, and attenuation of neuronal apoptosis. In addition, P4 attenuates neurodegeneration by antioxidant effects and enhancement of remyelination of injured neurons.A high P4 serum level reduces aldosterone activity resulting in a reduction of extracellular fluid by natriuresis, though a low P4 serum level causes fluid retention due to compensatory elevation of aldosterone.Prolong use of P4 is associated with the development of many adverse effects which are classified as very common (10% or more) including breast enlargement (40%), breast tenderness (27%), perineal pain female (17%), breast pain (16%), nocturia (13%), urinary problems (11%), vaginal discharge (11%), Common (1% to 10%) including ovarian hyperstimulation syndrome, breast pain, vaginal dryness, pruritus genital, uterine spasm, vaginal bleeding, altered periods, amenorrhea, intercurrent bleeding, and uncommon (0.1%–1%) including vulvovaginal disorders, vaginal mycosis, breast disorders, pollakiuria, incontinence, ovarian enlargement, pelvic pain, vulvovaginal pruritus, galactorrhea.1.28+, but not in CD4+, T lymphocytes. P4 binding to T and Jurkat cell membranes of steroid membrane receptors. P4 activated an inhibitory G‐protein suggesting that mPRs are coupled to inhibitory G‐protein in Jurkat cells. These results suggest a potential novel mechanism for P4 immunoregulatory function through activation of mPRs.+ T cells during normal pregnancy. As well, mPRs may correlate with the immunomodulatory properties of P4 on T cells. Variation in the expression of mPRs may influence P4 regulatory performance during pregnancy. PIBF increases in pregnant normal women compared to women who have experienced abortion.PRs are widely distributed in the immune cells and play a crucial role in the immunomodulation effects.Moreover, P4 reduces the risk of autoimmune diseases by counteracting the effects of estrogen on the immune cells.A.A, can inhibit toll‐like receptor 4 (TLR4) signaling.Furthermore, P4 active metabolites like allopregnanolone are mainly produced in the brain and act as a positive modulator of GABAThese verdicts revealed the immunological effects of P4, which is primarily anti‐inflammatory, preventing immuno‐inflammatory complications in different inflammatory and autoimmune disorders Figure .1.31.3.1It has been reported that P4 had differential effects on the outcomes of viral infections that could be beneficial or harmful. In influenza, A virus infection that infects epithelial cells of the respiratory system can induce immune cell activation and release of pro‐inflammatory cytokines.Similarly, P4 induces the expression of IL‐22, which promotes the regeneration of pulmonary epithelial cells following infection with the influenza A virus.2Different studies documented that the acquisition of HIV‐1 infection in females was increased in females during the luteal phase of the menstrual cycle when P4 is highest and during the use of hormonal contraceptives.On the other hand, there is a close relationship between pregnancy and the risk of HIV‐1 infection regarding the potential role of P4.Herein, there are potential controversies regarding the role of P4 in the acquisition risk of HIV‐1 replication during the reproductive period and hormonal replacement therapy during menopause.2.1Infection with hepatitis E virus (HEV) is linked with high feto‐maternal mortality; a prospective study included 100 pregnant women with acute HEV and 43 with fulminant HEV compared to 50 healthy pregnant women regarding the expression of PRs and PIBF showed that viral load was higher in patients with fulminant HEV compared to acute HEV.Concerning the role of P4 in HBV, it induces the production of pro‐inflammatory cytokines in patients with chronic HBV infection.Regarding HCV, P4 inhibits INF signaling by suppressing the expression of TLR7 in peripheral blood mononuclear cells in patients with HCV.These explanations revealed a controversial role of P4 in different hepatitis viral infections, which could be beneficial or harmful.2.28+, cytokine production, and impairment of expression of co‐stimulatory molecules on the dendritic cells.Moreover, P4 increases the susceptibility to herpes simplex virus infection (HSV) and reduces protection after HSV vaccination.On the other hand, viral infections may affect the P4 level by different mechanisms. For example, the proliferation of Zika virus (ZV) in the antral follicles promotes follicle degeneration with subsequent distortion of ovarian steroidogenesis and the development of ovarian failure.2.3It has been shown that P4 improves innate immune response and mitigates the release of pro‐inflammatory cytokines by promoting immune tolerance and T cells‐mediated anti‐inflammatory response.In addition, Th17 cells, which produce IL‐17, IL‐21, and IL‐22, can induce neutrophil recruitment, systemic inflammation, upregulation of fibrinogen, and development of cytokine storm. High pro‐inflammatory cytokines and cytokine storms in COVID‐19 patients promote the skewing of naïve T cells toward Th1 and Th17 with successive inflammatory changes. Hu et al.Preclinical findings indicated that P4 rescued the body weight loss of the SARS‐CoV‐2‐infected hamsters in a dose‐dependent manner.These findings suggest that P4 therapy is effective in the management of severe hypoxemic Covid‐19 patients. The underlying protective effect of P4 in COVID‐19 is mainly due to the inhibition release of pro‐inflammatory cytokines with augmentation of anti‐inflammatory cytokines through inhibition of IL‐6 and increment of IL‐10, respectively.Of note, a low dose of P4 inhibits the NF‐κB signaling pathway and prevents the development of endometriosis.Indeed, P4, through stimulation of GRs, reduces the expression of IL‐1β and COX‐2 by inhibiting MAPK signaling pathways.Besides, SARS‐CoV‐2 infection is associated with the activation of MAPK due to the downregulation of ACE2 and upregulation of AngII. Also, SARS‐CoV‐2 can directly activate MAPK to promote its replication. Activated MAPK induces vasoconstriction and immuno‐thrombosis with subsequent COVID‐19 complications, including lung fibrosis and ALI/ARDS.Notably, P4 is regarded as a potent antagonist of MR activated by stimulated aldosterone from high AngII during SARS‐CoV‐2 infection. Aldosterone exerts immunological effects through expressing MRs on the immune cells.Furthermore, P4 hinders the expression of TLR4 and CD80/CD86.Likewise, nod‐like receptor pyrin 3 (NLRP3) inflammasome is highly activated in SARS‐CoV‐2 infection and, together with TLR4, MAPK, and NF‐κB signaling pathways, leading to macrophage activation and release of pro‐inflammatory cytokines.Interestingly, P4 reduces STAT3 activation in response to IL‐6.Interestingly, autophagy is an essential intracellular mechanism for eliminating and recycling toxic unfolding proteins during inflammatory diseases, including COVID‐19.A.A neurotransmission. Therefore, P4 therapy may reduce neuropsychiatric manifestations and dysautonomia in COVID‐19 patients through modulation of the central GABA pathway.On the other hand, Covid‐19 is associated with neuropsychiatric manifestations including confusion, headache, seizure, stroke, and dysautonomia due to alteration of brain neurotransmitters, mainly GABA inhibition and development of sympathetic storm. It has been revealed that P4 active metabolite allopregnanolone is regarded as a neurosteroid that acts as a positive modulator of GABAMoreover, P4 is regarded as an allosteric modulator of the nAchR.A neurotransmission, may improve central and peripheral complications in COVID‐19 (Figure Therefore, P4, through modulation of inflammatory signaling pathways, pro‐inflammatory cytokines, anti‐inflammatory cytokines, immunological tolerance, and GABAThe present narrative review study had several limitations, including the paucity of clinical studies regarding the use of P4 in treating COVID‐19. Most of the mechanisms related to the potential effects of P4 on Covid‐19 are speculative depending on previous experimental and clinical conditions studies. However, this study opens a new way regarding the beneficial effects of P4 in the management of COVID‐19 that need to be confirmed by clinical trials and prospective studies.3P4 has anti‐inflammatory and immunosuppressive effects; thereby, it is effective in different inflammatory disorders. The regulatory effects of P4 on various viral infections are controversial as it may reduce or ameliorate these viral infections. However, the possible role of P4 in COVID‐19 could be beneficial through the modulation of inflammatory signaling pathways, induction of the release of anti‐inflammatory cytokines, and inhibition release of pro‐inflammatory cytokines. However, P4 is capable of shifting the immune response from an anti‐viral Th1 response to a Th2 response, which is likely to favor the virus. This effect may reduce the risk of SARS‐CoV‐2‐induced cytokine storm, but may enhance the proliferation of SARS‐CoV‐2. Therefore, the role of P4 could be beneficial or detrimental to the course of SARS‐CoV‐2 infection and COVID‐19 severity. This review cannot conclude the final role of P4 in COVID‐19. Herein, clinical trials and prospective studies are recommended to substantiate the potential role of P4 in the management of COVID‐19 regarding sex and the reproductive period.Hayder M. Al‐Kuraishy: conceptualization; resources. Thabat J. Al‐Maiahy: conceptualization; resources. Ali I. Al‐Gareeb: conceptualization; resources. Athanasios Alexiou: writing—original draft. Marios Papadakis: funding acquisition; writing—review & editing. Omnya Elhussieny: writing—review & editing. Hebatallah M. Saad: supervision; writing—review & editing. Gaber El‐Saber Batiha: validation; visualization; writing—original draft.The authors declare no conflicts of interest.


Ursus arctos) was used as a model species for which dietary ingredient and nutrient concentrations as well as nutrient ratios were determined annually, seasonally and per reproductive class. Brown bears had a vertebrate‐ and ant‐dominated diet in spring and early summer and a berry‐dominated diet in fall, which translated into protein‐rich and carbohydrate‐rich diets, respectively. Fiber concentrations appeared constant over time and averaged at 25% of dry matter intake. Dietary ingredient proportions differed between reproductive classes; however, these differences did not translate into a difference in dietary nutrient concentrations, suggesting that bears manage to maintain similar nutrient profiles with selection of different ingredients. In terms of nutrient ratios, the dietary protein to non‐protein ratio, considered optimal at around 0.2 (on metabolizable energy basis), averaged around 0.2 in this study in fall and around 0.8 in spring and summer. We introduced the minimal non‐fat to fat ratio necessary for efficient maintenance metabolism. This ratio varied across seasons but never fell beneath the theoretically estimated minimum to ensure metabolic efficiency. This population thus managed to ingest diets that never exerted a lack of glucogenic substrate, suggesting that metabolic efficiency may either be a driver of active diet selection or that natural resources available to bears did not constitute a constraint in this respect. Given the considerable proportion of fiber in the diet of brown bears, the relevance of this nutrient and its role in foraging behavior might be underestimated.The dietary nutrient profile has metabolic significance and possibly contributes to species' foraging behavior. The brown bear ( Dietary nutrient concentrations (incl. fiber) and ratios of free‐ranging Swedish brown bears showed that: the diet is protein‐dominated in spring and early summer, carbohydrate‐dominated in fall and constant in fiber throughout seasons; bears managed to obtain similar dietary nutrient profiles with different ingredients; the protein to non‐protein ratio only averaged around 0.2 in fall and the fat to non‐fat ratio varied across seasons but never fell beneath the theoretically estimated minimum to ensure metabolic efficiency. The area can be characterized as rolling terrain with altitudes ranging from 250 to 650 m above sea level  and the Swedish Environmental Protection Agency (NV‐00741‐18). The GPS collars are programmed to record at least one GPS location per hour. Part of the SBBRP monitoring program focusses on documenting diet composition through scat analysis. For this reason, scats from tracked bears were collected throughout the active period (April/May until late October) in a frequency of 1 scat/individual/week (up until 2015) or 1 scat/individual/2 weeks (from 2016 on). Brown bears commonly defecate close to their resting sites or ‘beds’  tags 40–50 brown bears with GPS‐GSM collars . The procedures required for capturing and handling brown bears ; other fruit (unidentified); invertebrates ; vertebrates ; vegetation ; miscellaneous ; bird eggs.Every scat was weighed, and scat volume was determined by water displacement. Dietary ingredients were determined following the methods of Hamer and Herrero , Dahle e2.4ith diet ingredient ingested and CFi equal to the dry matter of diet ingredient i ingested (g)/fecal volume of the residue of diet ingredient i (mL). Scats with more than 5% EDC miscellaneous were omitted from the dataset to avoid any leveraging effect because no nutrient composition can be attributed to this category : diet ingredients have a different digestibility; hence the proportion of easily digestible diet ingredients may be underestimated in the feces in comparison to the actual biomass ingested. Establishing diet ingredient composition solely based on percent volume  in the scats is therefore biased , i.e. DM, crude protein (CP), crude fat , ash, fiber as crude fiber (CF) and total dietary fiber (TDF) (as % DM). Digestible carbohydrates (NfE) were calculated following the formula:Priority was given to subtracting TDF instead of CF given the fact that it is a more complete measure for fiber. For vertebrate diet items (i.e. whole prey), TDF values were extracted from Pritchard and Robbins , where tThe nutrient composition of the diet was calculated with the formula:The nutrient composition of the diet as ingested was used to calculate the protein to non‐protein ratio CP:(EE + NfE) as well as the non‐fat to fat ratio (CP + NfE):EE both on DM and ME basis.The percent volume of diet ingredients was used to calculate the diet and nutrient composition of the diet consumed per scat as follows:2.5The ingredient and nutrient composition of Swedish brown bears was calculated for all scats during the time period 2015–2018. Per scat the following data were listed: GPS coordinates, start and end date/time at the bear bed, bear ID, bear name, sex , age, age group (subadult [until 3 years] or adult [from 4 years on]), reproductive class, season of scat deposition, number of years the bear was tracked (the amount [1–4] and consecutive years or not [Cons or NonCons]), scat weight, scat volume, Vf diet items in scat, EDC diet and nutrient composition of the diet (online Dryad repository). The ingredient EDC's were grouped in six primary categories: fruit, insects, vertebrates, natural vegetation , anthropogenic vegetation  and other (miscellaneous [<5 %EDC] and eggs). Reproductive class was divided into the categories adult (4 years and older) lone female, adult male, adult female with yearlings, adult female with cubs of the year, subadult (1–3 years of age) female, and subadult male. Subadult females or males still accompanying their mother were excluded from the dataset. Seasonal division was based on annual pivot points that influence bear diets according to Stenset et al. : spring 2.6https://rdrr.io/github/timnewbold/StatisticalModels/) which performs backward stepwise selection of fixed effects to reach a minimal adequate model containing only variables and interactions statistically significant at the α ≤ 0.05 level. Model residuals were normally distributed, as evidenced by Shapiro–Wilk W values ≥0.95, allowing a Gaussian error distribution. Homogeneity of variance was checked by plotting standardized residuals against fitted values. For post‐hoc identification of significant differences between factors in the linear mixed effects models, we used ‘lsmeans’ from R package ‘emmeans’  and nutrient concentrations , on DM basis and ME basis, as well as the ratios CP:(EE + NfE) and (CP + NfE):EE on DM and ME basis, were analyzed using linear mixed models (LMM) for the effects and all possible two‐way interactions of the factors year, season and reproductive class using RStudio (R4_0_2), specifying bear id as a random effect to account for non‐independence of scats produced by the same individual. Variable selection was performed using the R ‘GLMERSelect’ function of the R package ‘StatisticalModels’  from 55 bears  were analyzed for ingredient and nutrient profiles and ratios , although summer diets also contained a considerable percentage of fruits (22.3 ± 33.3% EDC), except for the year 2015 when fruit contribution was lower than in other study years . Adult males had a lower fruit content in their diets than subadult females . The insect share in the bear diets was highest in summer (30.1% ± 34.7% EDC) compared to fall , with the largest summer–fall difference in the year 2015  and no significant difference in 2017 . Overall, adult males had the lowest share of insects in their diet (13.6 ± 26.7% EDC) and subadult males the highest share . Vertebrates were always higher in the summer diet (21.0 ± 34.6% EDC) compared to the fall diet . Females with cubs of the year had the lowest share of vertebrates in their diet (8.0 ± 18.8% EDC) compared to females with yearlings  and adult lone females . Natural vegetation in the fall diet was low and constant over the years (6.0 ± 18.4% EDC) compared to higher levels in summer , except for the year 2016 where fall and summer both showed low levels . Anthropogenic vegetation consumption  was higher in fall diets (14.2 ± 31.8% EDC) compared to summer diets  and was very high in the fall diet of adult males (43.5 ± 44.8% EDC) compared to all other classes ranging from 3.6% to 14.2% EDC; vs. adult lone female , versus adult female with yearlings , versus adult female with cubs of the year , versus subadult female  and versus subadult male .In fall diets, the amount of fruit consumed (70.4 ± 37.2% EDC) was always higher than in summer diets , with the largest summer–fall difference in the year 2015 . The dietary fat content was always higher in summer (11.1 ± 6.0% DM) compared to fall  with a slight decrease in the difference between summer and fall from 2015 to 2017 (delta 7.6 to delta 3.3%), after which this difference increased again in 2018 . The concentration of carbohydrates (NfE fraction) in the summer (27.4 ± 19.8% DM) only reached about half of the carbohydrate concentration of the fall diet , and in the year 2015 summer only had one third (19.6 ± 15.3% DM) of the fall carbohydrate concentrations . Fiber concentrations were only slightly higher in fall (25.5 ± 6.9% DM) compared to summer , with no difference at all in the years 2015 and 2017   with the biggest fall–summer difference in 2015 . No significant effects of reproductive class were observed on any of the nutrient concentrations on DM basis. Protein, fat and carbohydrate concentrations on ME basis showed similar annual and seasonal patterns as on DM basis.The protein content of the ingested diet in summer was almost three times higher 38.0 ± 21.8% DM) compared to the fall diet  with the biggest difference between seasons in the year 2015 . Similarly, on a ME basis, the CP:(EE + NfE) in fall generally corresponded to the ratio determined by Erlenbach et al. , with the biggest difference between seasons again in the year 2015  and adult males . On a ME basis, the (CP + NfE):EE ratio was higher in fall (7.5 ± 3.6) compared to summer , with the biggest difference between fall and summer in the year 2015 . The (CP + NfE):EE ratio only sporadically fell beneath the theoretical 3.55 limit on DM or 1.58 on ME basis  on DM basis was low in fall for all years 0.27 ± 0.40) compared to summer, when this ratio was higher  dominated in summer and carbohydrate‐dominated in fall. The latter can be considered a logical consequence of the seasonal ingredient intake: a strong presence of vertebrates (predominantly moose) and ants in summer with a gradual increasing share of berries (predominantly bilberry) toward fall , where the alternative use of marine versus terrestrial foraging habitats incurred similar energetic costs to parents , nor a class x season interaction, which indicates that bear groups obtain similar nutrient intake profiles by means of different ingredients, and thus regardless of foraging strategy. The latter strengthens more recent evidence in ecology in which animals can obtain a similar fitness with differing foraging strategies, for example in Lesser black‐backed gulls  and stone martens (Martes foina) have been observed to forage for an overall high annual protein ratio  seem to experience negative effects of summer diets rich in fruits on their survival and fecundity        under human care showed that high protein diets  led to higher reproductive success in hamsters post‐hibernation compared to high fat diets , although when we calculate the protein ratio, it is rather similar to the low protein ratio observed in fall for brown bears in both cases (0.30 and 0.17) , the diet in the pre‐hibernating phase mainly consists of fatty seeds :EE, that is non‐fat to fat ratio, which we suggest as a tool to evaluate whether bears feed for efficient maintenance energy metabolism. The process of combusting nutrients for energy is subject to biochemical limitations that may prevent animals from going for the ‘highest energy resource’ possible. This consideration is based on the function of the citric acid cycle. The citric acid cycle is the final common step in aerobic oxidation of the three nutrients  or ‘fuels’ that leads to ‘free’ available energy  compared to protein  and carbohydrates   Figure , the queOne final note on fiber intake concentrations and the inclusion of animal fiber in the latter is that including animal fiber ; however, in other seasons, other processes – metabolic or ecologic – might drive nutrient selection. Fiber had a considerable share in the diets of brown bears, hence should be given more attention in the future.Annelies De Cuyper: Conceptualization ; data curation (lead); formal analysis ; funding acquisition ; investigation ; methodology ; project administration (lead); resources ; validation ; visualization ; writing – original draft (lead); writing – review and editing . Diederik Strubbe: Conceptualization ; data curation ; formal analysis (lead); investigation ; methodology ; software (lead); validation ; visualization ; writing – review and editing . Marcus Clauss: Conceptualization ; data curation ; formal analysis ; investigation ; methodology ; supervision ; validation ; writing – review and editing . Luc Lens: Conceptualization ; funding acquisition ; investigation ; methodology ; project administration ; resources ; supervision ; validation ; writing – review and editing . Andreas Zedrosser: Data curation ; investigation ; methodology ; project administration ; resources ; supervision ; validation ; writing – review and editing . Sam Steyaert: Data curation ; investigation ; methodology ; supervision ; validation ; writing – review and editing . Leen Verbist: Conceptualization ; data curation ; methodology ; validation ; writing – review and editing . Geert P. J. Janssens: Conceptualization ; formal analysis ; funding acquisition ; investigation ; methodology ; project administration ; resources ; supervision (lead); validation ; visualization ; writing – review and editing .The authors declare that they have no conflict of interest.Appendix S1.Click here for additional data file.Appendix S2.Click here for additional data file.


Neoadjuvant chemotherapy followed by radical cystectomy (RC) is the standard of care for patients with muscle‐invasive bladder cancer (MIBC). However, treatment outcomes are suboptimal. Camrelizumab, a PD‐1 blockade, has shown benefits in several tumors. This study aimed to investigate the efficacy and safety of neoadjuvant camrelizumab in combination with gemcitabine plus cisplatin (GC) followed by RC for MIBC patients.2 on day 1 and 8, and cisplatin 70 mg/m2 on day 2, followed by RC. The primary endpoint was pathologic complete response .This was a multi‐center, single‐arm study that enrolled MIBC patients with a clinical stage of T2‐4aN0‐1M0, and scheduled for RC. Patients received three 21‐day cycles of camrelizumab 200 mg on day 1, gemcitabine 1000 mg/mFrom May 2020 to July 2021, 43 patients were enrolled and received study medications at nine centers in China. Three of them were deemed ineligible and excluded from efficacy analysis but included in safety analysis. In total 10 patients were unevaluable as they declined RC (two due to adverse events [AEs] and eight due to patient's willingness). Among 30 evaluable patients, 13 patients (43.3%) achieved pCR, and 16 patients (53.3%) achieved pathologic downstaging. No AEs leading to death were observed. The most common AEs were anemia (69.8%), decreased white blood cell count (65.1%), and nausea (65.1%). Immune‐related AEs were all grade 1 or 2. Pathologic response was not correlated with PD‐L1 expression status or tumor mutation burden. Individual genes as a biomarker for pathologic response were not identified.Neoadjuvant treatment with camrelizumab and GC regimen demonstrated preliminary anti‐tumor activity for MIBC patients with manageable safety profiles. The study met its primary endpoint, and the following randomized trial is ongoing. In this multicenter, single‐arm study, we investigated the anti‐tumor activity and safety of neoadjuvant camrelizumab plus gemcitabine and cisplatin for MIBC patients. The pCR was 43.3% in evaluable patients. Our study met its primary endpoint, and a randomized trial is currently underway. The study consisted of two stages: a single‐arm pilot study in the first stage to investigate the combination regimen's efficacy and safety in patients with local advanced resectable MIBC, followed by a randomized controlled study in the second stage to further compare the combination regimen with chemotherapy alone. The second stage would initiate when the first stage met the primary endpoint. Institutional review boards of all participant centers approved the study, and all patients provided informed consent to participate. This study was registered on Eligible patients were at least 18 years old, diagnosed with histologically confirmed local advanced resectable bladder cancer without metastasis , naïve to platinum‐based systemic therapy, had at least one measurable bladder lesion according to the RECIST 1.1 criteria, clinical stage of T2‐4aN0‐1M0, scheduled for RC, Eastern Cooperative Oncology Group (ECOG) performance score of 0 or 1, anticipated living longer than 3 months, and had normal hematologic test and sufficient organ function at baseline. Patients with autoimmune or immunodeficiency disease, requiring immunosuppressive therapy in the past 2 weeks, or any other conditions unsuitable for this study, as evaluated by investigators, were excluded.2.2During the screening process, all patients underwent a transurethral resection of bladder tumor (TURBT) and underwent imaging (CT or MRI) to assess local and metastatic lesions. Patients were required to have pathologic evidence of muscle invasion in the TURBT specimen prior to enrollment. Tumor tissue and peripheral blood were collected for biomarker analysis.2 IV once on day 1 and 8, and cisplatin 70 mg/m2 IV once on day 2 every 3 weeks. In the event of treatment‐related adverse events (TRAEs), the related study treatment could be discontinued, while the other two treatments could continue at the discretion of the investigator. RC was performed within 3–4 weeks after the completion of neoadjuvant treatment. As per the standard of care, a standard lymph node dissection was conducted during RC.In the single‐arm stage, patients received neoadjuvant treatment 2 weeks after TURBT. To minimize the time to surgery and potential AEs, three cycles of camrelizumab plus GC were administered. Each cycle consisted of camrelizumab 200 mg intravenously (IV) once on day 1, gemcitabine 1000 mg/m2.3The primary endpoint was pathologic complete response (pCR), which was defined as no viable tumor cells identified in resected tissue (pT0N0). Secondary endpoints included pathologic downstaging (defined as pathologic stage < pT2N0), event‐free survival , OS (defined as the time from initiation of chemo‐immunotherapy to death from any cause), and safety. In addition, the study conducted PD‐L1 expression status and whole exoneme sequencing (WES) to explore the potential biomarkers for efficacy. The AEs were reported and graded by National Cancer Institute Common Terminology Criteria for AEs (version 5.0), and toxicities were assessed for 90 days after dosing was complete.2.4Tissue samples obtained from patients during TURBT and peripheral blood samples were collected for assessing programmed cell death ligand 1 (PD‐L1) expression and WES prior to treatment. PD‐L1 expression was evaluated with the IHC 22C3 pharmaDx (Agilent Technologies). DNA was extracted from TURBT tissue and peripheral blood samples, and invasive tumor content was assessed by pathologists to ensure sufficient tumor cells. DNA was extracted using the DNeasy Blood and Tissue Kit . Targeted capture pulldown and exon‐wide libraries were created from native DNA using NadPrep® Hybrid Capture Reagents  and NadPrep® DNA Library Preparation Kit  E96. Paired‐end sequence data were generated using Illumina NovaSeq machines with an average sequencing depth of 100 × for controls and 200 × for tumors.The DNA damage response and repair (DDR) gene set, consisting of 276 genes, was built by assembling relevant gene lists, including MSigDB v5.0 , and knowledge‐based curation of information on specific DNA repair pathways or sub‐pathways, as previously reported.2.5Previous studies have shown that patients with MIBC who underwent neoadjuvant GC had a pCR rate of approximately 25%.All eligible patients who received at least one dose of camrelizumab and underwent cystectomy with post‐operative pathologic assessment was included in the efficacy analysis. The pathologic response will be evaluated by a central pathology committee. Safety analysis will include all eligible patients who received at least one dose of camrelizumab.p < 0.05 was used.Statistical analyses were conducted using SAS 9.4 and R 3.6.0, and a significance level of 33.1Forty‐three patients with MIBC were enrolled from May 2020 to July 2021 at nine centers in China, and all received the study drugs. Three of the enrolled patients were found ineligible during the study  and were excluded from the baseline and efficacy analysis . At the time of cystectomy, 16 patients achieved < pT2N0M0, with a pathologic downstaging rate of 53.3% , and none of the patients had disease upstaging , the median EFS and median OS for 30 evaluable patients have not been reached. One‐year EFS and OS rates were 79.2% and 81.7%, respectively  completed three cycles of neoadjuvant therapy. TRAEs of any grade were reported in 95.3% of patients. Grade 3 or 4 TRAEs were reported in 34.9% of patients (Supplementary Table 3.4p = 0.25)  was observed. However, the median TMB in the pCR group was numerically higher than that in the non‐pCR group  Figure . The com) Figure . The top4The neoadjuvant treatment of local advanced MIBC patients with three cycles of camrelizumab plus GC demonstrated preliminary anti‐tumor activity in this single‐arm stage. The results of this pilot study met the primary endpoint, and the subsequent randomized stage is currently ongoing to compare camrelizumab plus GC with GC alone as neoadjuvant treatment for patients with local advanced MIBC. These preliminary findings are consistent with other studies that have investigated the use of ICI‐chemotherapy combination regimens as neoadjuvant treatment for MIBC patients. Two pilot studies involving four cycles of pembrolizumab combined with GC demonstrated pCR rates of 36% and 44%, respectively,The pathologic downstaging rate, as a secondary endpoint, in this study was similar to rates reported in previous studies. Two separate studies that administered four cycles of pembrolizumab combined with GC regimen as neoadjuvant treatment for MIBC patients reported pathological downstaging rates of 56% and 61.1% after resection.The optimal timing and duration of neoadjuvant therapy for MIBC remain topics of ongoing debate. Previous studies have suggested that delaying cystectomy after neoadjuvant therapy may negatively impact patients' outcomes.Patients who achieved pCR in this study had a more favorable 1‐year survival status compared to those who did not achieve pCR. Previous studies have also shown that patients who responded well to neoadjuvant ICI‐chemotherapy had significantly better recurrence‐free survival compared to non‐responders.RC is the standard treatment for MIBC. However, some patients may not be eligible for surgery or may prefer to avoid RC due to concerns about quality of life. Therefore, bladder preservation therapies have emerged as an alternative treatment method.The hematologic toxicity profile of this study was similar to that of other MIBC patients treated with neoadjuvant ICI‐chemotherapy.No significant correlation was found between PD‐L1 expression and pathologic response in this study or in other studies of ICI‐chemotherapy as neoadjuvant treatment for MIBC patients.Recent studies suggest that mutations in the DDR pathway in UC may be associated with improved responses to NAC and prolonged survival.FRG1 has been found to play a role in both tumorigenesis and angiogenesis, and several previous studies have demonstrated an association between FRG1 expression and OS in various types of tumors.There are several limitations to this study that should be considered. First, the results presented are from the single‐arm stage of the study, and therefore, further evidence from the ongoing randomized stage is required to strengthen the findings. Second, the follow‐up period may have been inadequate to capture all instances of recurrence, and longer follow‐up is needed to provide a more comprehensive understanding of the survival benefit of ICI‐chemotherapy as neoadjuvant treatment for MIBC patients. Moreover, some patients opted not to undergo RC, which may result in an overestimation of the efficacy of our analysis, as our analysis primarily focused on patients who underwent RC. Furthermore, patients who declined RC did not have available data for EFS. This issue will be addressed in the ongoing randomized controlled trial. Lastly, toxicities were only assessed for 90 days, and delayed irAEs may not have been detected within this timeframe.5In conclusion, neoadjuvant treatment with short‐course camrelizumab and GC regimen for MIBC patients was found to be tolerable and provided promising anti‐tumor activity.Sujun Han: Conceptualization ; data curation ; formal analysis ; investigation ; writing – original draft ; writing – review and editing . Zhigang Ji: Investigation ; writing – review and editing . Junhui Jiang: Investigation ; writing – review and editing . Xinrong Fan: Investigation ; writing – review and editing . Qi Ma: Investigation ; writing – review and editing . Linjun Hu: Investigation ; writing – review and editing . Wen Zhang: Investigation ; writing – review and editing . Hao ping: Investigation ; writing – review and editing . Jiansong Wang: Investigation ; writing – review and editing . Wanhai Xu: Investigation ; writing – review and editing . Benkang Shi: Investigation ; writing – review and editing . Wei Wang: Investigation ; writing – review and editing . Haifeng Wang: Investigation ; writing – review and editing . Honglei Wang: Investigation ; writing – review and editing . Shouzhen Chen: Investigation ; writing – review and editing . Hai‐Long Hu: Investigation ; writing – review and editing . Jianming Guo: Investigation ; writing – review and editing . Shen Zhang: Investigation ; writing – review and editing . Shuai Jiang: Investigation ; writing – review and editing . Quan Zhou: Investigation ; writing – review and editing . Nianzeng Xing: Conceptualization ; data curation ; funding acquisition ; investigation ; supervision ; writing – review and editing .This work was supported by The Capital Health Research and Development of Special Funding (2022–1‐4021).The authors have no conflict of interest to declare.The study has been approved by institutional review board of all participant centers.All patients signed inform consent to participate this study.Not applicable.https://www.chictr.org.cn (ChiCTR2000032359).This study was registered on Supporting information S1. Supplementary materialClick here for additional data file.