A total of 36 parturient with severe preeclampsia were enrolled, including 18 in the dexmedetomidine (DEX) group and 18 in the saline group. Compared with the saline group, among those in the DEX group, CO was reduced by 1.30 L/min . Additionally, HR , SBP , DBP , and MAP  were reduced in the DEX group compared with the saline group. In contrast, there were no changes observed in SV and ICON between groups. In conclusion, dexmedetomidine reduces cardiac output by inhibiting the acceleration of heart rate without sacrificing myocardial contractility and stroke volume.This study was to investigate the hemodynamic effect of dexmedetomidine among parturient with severe preeclampsia after cesarean section. Parturient with severe preeclampsia were randomly allocated to receive dexmedetomidine (0.2-0.7  Preeclampsia is the most common complication of pregnancy affects 2% to 8% of pregnancies , 2. Thisα-2-receptor agonist, is a first-line sedative medication in ICU and has been increasingly used for obstetric anesthesia. Dexmedetomidine, which provides light sedation, possesses analgesic, sympatholytic, anxiolytic properties and attenuates the stress response without significant respiratory depression  to +4 [combative], as assessed at least every 2 hours. The saline group received equivalent volumes of saline. If bradycardia occurred which was defined as heart rate < 50 beats·min−1, the infusion of dexmedetomidine was terminated and the patient was excluded from the study.In the DEX group, patients received continuous infusion of dexmedetomidine without loading dose at a rate of 0.2-0.7 Severe preeclampsia met the Clinical Management Guidelines for Obstetrician-Gynecologists of ACOG criteria for preeclampsia diagnosis and fulfilled one of the severe features : systoliThe primary outcome was the change of cardiac output (CO) from baseline. Secondary outcomes included the change of other hemodynamic variables from baseline: stroke volume (SV), index of contractility (ICON), heart rate (HR), systolic blood pressure (SBP), diastolic blood pressure (DBP), and mean arterial blood pressure (MAP). Variables were obtained before dexmedetomidine or saline administration as baseline and at 10 hours after dexmedetomidine or saline administration, respectively. Adverse events such as eclampsia, heart failure, bradycardia, and nausea and vomiting (PONV) were monitored and recorded. The hemodynamic variables were obtained by the ICON™ monitor, a noninvasive hemodynamic monitoring device .Study data from Lee et al. showed a decrease of CO from 3.72 ± 1.0 L/min at baseline to 2.90 ± 0.5 L/min at 20 min after administration in the DEX group with no changes (from 3.61 ± 1.1 L/min to 3.87 ± 1.4 L/min) in the saline group. A two-sided 0.05 level of significance and a sample size of 36 patients (18 per group) provided 80% statistical power to demonstrate this difference in CO. To accommodate for a 10% attrition rate, we will recruit a total of 40 patients .t tests were performed for between-group differences. Within-group comparisons from baseline to drug administration were tested with paired-samples t tests. All statistical analyses were conducted with SPSS 25.0 software package, a 2-sided P value of <0.05 was considered statistically significant.The normality of the data distribution was verified by the Shapiro-Wilk tests. For continuous data, means with standard deviations (SDs) were presented. The changes of hemodynamic variables were calculated as the value after drug administration minus the value at baseline. Mean differences were expressed with their 2-sided 95% confidence intervals. Independent-samples From the 64 patients who met the enrollment criteria, 40 patients agreed to participate. Thirty-six parturient with severe preeclampsia completed the study, 18 in each group, respectively .Demographic characteristics and baseline of hemodynamic variables were similar in the two groups .P = 0.019). Similarly, HR was remarkably accelerated in the saline group and significantly reduced in the DEX group compared with the saline group . No changes were observed in SV and ICON in either group , .As P = 0.030) and diastolic blood pressure  as well as mean arterial blood pressure  were also significantly reduced in the DEX group compared with the saline group . No major adverse events such as eclampsia and heart failure were recorded during the study. Three patients reported nausea and vomiting in both groups after intervention.Five patients in the DEX group received antihypertensive medication and 6 in the saline group during study period (27.8% vs. 33.3%, The most important finding of present study is the infusion of dexmedetomidine without loading dose reduce cardiac output by attenuated heart rate in patients with severe preeclampsia postoperative but does not impair stroke volume and index of contractility. It is also suggested that the administration of dexmedetomidine decrease blood pressure.Several researches reported the effects of dexmedetomidine on cardiac function. Lee et al.  assessedPrevious studies have reported that administration of dexmedetomidine provided a significant hemodynamic stability during cesarean section in patients with preeclampsia –28. TherThe first limitation is associated with lack of blinding. The unblinded manner may have introduced subjective bias by clinicians. In addition, the plasma concentration of dexmedetomidine was not measured, which may be useful to determine whether plasma concentration varied hemodynamic effects, especially on cardiac function. Thirdly, we did not monitor the plasma level of cortisol and noradrenaline, which could improve our understanding in the potential mechanism of dexmedetomidine on hemodynamic.In conclusion, dexmedetomidine may reduce the risk of heart failure by reducing cardiac work and cardiac afterload. Sedation should be routinely performed in parturient with severe preeclampsia after cesarean section to cope with adverse events caused by stress, pain, postoperative hypertension, and increased cardiovascular volume.


Furthermore, vital signs and adverse events (AEs) were observed. Both groups had a slower renal function decline after treatment than before treatment. Compared with RG group, more reductions of Scr, BUN, UA, and better improvement of eGFR were observed in RG + CHF group (p < 0.05). Additionally, the levels of urinary AKI biomarkers, renal fibrosis biomarkers, and TCM syndromes were decreased in RG + CHF group versus RG group (p < 0.05). No significant between-group differences were observed of AEs. We thus concluded this novel therapy of CHF combining RG might be a useful method for treating A on C patients.Lack of effective drugs for acute kidney injury (AKI) grades 1–2 is a crucial challenge in clinic. Our previously single-center clinical studies indicated Chuan Huang Fang (CHF) might have nephroprotection in AKI on chronic kidney disease (CKD) (A on C) patients by preventing oxidant damage and inhibiting inflammation. Reduced glutathione (RG) has recently been shown to increase the clinical effectiveness of high-flux hemodialysis among patients with severe AKI. In this multicenter randomized controlled clinical study, we designed a new protocol to assess the efficacy and safety of CHF combining RG in patients with A on C. We also explored therapeutic mechanisms from renal fibrosis biomarkers. 98 participants were randomly and equally divided into the RG and RG + CHF subgroups. The RG and RG + CHF groups received general treatments with RG and a combination of RG and CHF, respectively. The therapy lasted for 2 weeks. In this study, the primary assessment result was a difference in the slope of serum creatinine (Scr) over the course of 2 weeks. The secondary evaluation outcomes were alterations in blood urea nitrogen (BUN), uric acid (UA), estimated glomerular filtration rate (eGFR), urinary AKI biomarkers, renal fibrosis biomarkers (transforming growth factor- Clinical Trial Registration:http://www.chictr.org.cn, ChiCTR2100043311Acute kidney injury (AKI) is described as a sudden deterioration in renal function that encompasses both structural failure and loss of functionality . As a seChronic kidney disease (CKD) is a combination of chronic illnesses attributed to a variety of indicators, particularly, inflammation, oxidative stress, and metabolic abnormalities . AKI andAccording to historical records, Chinese physicians utilized traditional Chinese medicine (TCM) to treat AKI during the era when RRT was absent . In ChinWe undertook a multicenter randomized controlled trial (RCT) for assessing the clinical efficacy and safety of CHF combing RG against A on C. By completing this trial in a timely manner, a unique pharmacological treatment method for AKI grades 1–2 would be developed.A multicenter randomized controlled clinical trial was performed in this study. The study duration for this research were ranged from 26 December2016- 13 April2020. Approval for the trial was granted by the international review board and ethical committees of each participating hospital and the Ethics Committee of Shanghai Municipal Hospital of Traditional Chinese Medicine (No.2020SHL-KYYS-60). This trial was registered in the Chinese Clinical Trials Register (No. ChiCTR2100043311) before the enrolment of the first participant.Three hospitals enrolled participants in this study: 1) Shanghai Municipal Hospital of Traditional Chinese Medicine, 2) Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine, and 3) Minhang Branch of Yueyang Hospital of Integrative Chinese & Western Medicine Affiliated to Shanghai University of Traditional Chinese Medicine. Overall, 98 participants were classified randomly and equally into the RG and RG + CHF groups. This study enrolled patients who had CKD stages 2–4 complicated with AKI grades 1–2 and toxicity stasis inter-combination syndrome, as well as spleen–kidney qi deficit.The following were the criteria for participation in the study: 1) patients satisfied all clinical guidelines for chronic kidney disease stages 2–4 and acute kidney injury grades 1–2, 2) patients satisfied the criteria for a diagnosis for the classification of TCM syndromes, 3) 24 h U-pro of patients ≤2.5 g, 4) ages of patients were range from 18 to 70 years, and 5) patients should be volunteered to participate and signed informed consent.tuberculosis, malignant tumors, or consumption disorders, 3) patients suffering from anorectal disorders who failed to receive enema, 4) patients who had received a kidney transplant, 5) patients who were psychopaths or who had poor compliance, 6) those with an allergic reaction to the therapeutic medication, and 7) those who were enrolled in other clinical trials within the past 3 months.The following were the criteria for excluding participants from the study: 1) patients who were pregnant or lactating, 2) patients with acute primary illnesses of other organs requiring immediate treatment, including active Guidelines for Clinical Research of Chinese Medicine (New Drug)  (Based on the ew Drug) and Diagnrotocol) , particiTo produce random number sequences, independent biostatisticians used the SPSS (version 21.0) program, which was used to establish these sequences using a simple random approach. The subjects agreed to take part in the research and completed a formal informed consent immediately. Participants also gave their informed permission before being randomly assigned to groups. Participants were assigned a random number and equally classified into two groups by the investigators depending on the order in which they were enrolled in this study: the RG and RG + CHF groups. During the random sampling process, closed envelopes labeled with sequential coding numerals were employed to keep track of allocation information during randomization, and participants, biostatisticians, and investigators were kept blind to the group allocation. Since there was no use of a placebo in the control group, it was not feasible to keep individuals completely unaware of the therapy. It was, however, concealed from all laboratory personnel who were involved in the investigations.Participants were recruited for the study after providing formal informed permission. They were categorized at random into the RG and the RG + CHF groups. The participants underwent training for study behavior and health to reduce the possibility of loss and ensure that they could follow the experiment without difficulty. Furthermore, they were informed about potential hazards, their rights, and responsibilities, as well as how to deal with an emergency. All of the patients were given a low-protein, high-quality, and low-salt meal plan to follow (protein consumption of 0.6–0.8 g/kg/day). They received basic therapies with the goal of restoring normal water and electrolyte levels as well as acid-base balance abnormalities, controlling blood pressure, improving anemia, and correcting renal bone illnesses. Certain investigators regularly followed up with participants and assisted them to adhere to treatment procedures and undergo tests as per schedule.Prepared rhubarb (Zhidahuang), Ligusticum wallichii (Chuanxiong), Smilacis glabrae (Tufuling), Coptidis rhizome (Huanglian), Codonopsis pilosula (Dangshen), Salviae miltiorrhizae (Danshen), Rhizoma Pinellinae Praeparata (Zhibanxia), Pericarpium citri reticulatae (Chenpi), Cordyceps sinensis (Chongcaojunsi), etc (The control group received an intravenous injection (IV) of RG 1.8 g mixed with 0.9 percent normal saline or 5 percent glucose in a volume of 250 ml once every day for a period of 2 weeks. The treatment group additionally received 200 ml CHF twice daily for a total of 2 weeks. Simultaneously, they received an enema of the concentrated solution of CHF once a day for 5 days , followed by 2 days of rest. RG was obtained from Chongqing Yaoyou Pharmaceutical Co. Ltd. The quality standards of CHF were controlled as follows: 1) All herbal medicines were from Shanghai Municipal Hospital of Traditional Chinese Medicine pharmacy, which were traceable; 2) CHF decoction was produced by the pharmacy; 3) Our team had professional pharmaceutical staff to detect the main monomer content of CHF  using fingerprints method regularly. The principal pharmaceutical ingredients of CHF are si), etc . Major csi), etc .Enema were performed by a professional clinician. A clinician took 100 ml concentrated medicine solution of CHF, waiting for the medicine solution close to almost human temperature, and put it into an enema bag. The patient was told to lie on the side of the operating bed, the clinician inserted enema tube to the patient’s anus 20–30 cm deep and then injected the medicine solution slowly, each dripping time for 20 min. After enema operation, the patient was instructed to lie on his pillow with buttocks rising 10–15 cm. The medical liquid should be maintained in the patient’s intestine for more than 1 h.A series of related studies were carried out, and data were recorded at baseline and every week throughout the duration of therapy. Patient data was gathered, efficacy-related exams were carried out, and indicators of mechanism and safety, as well as medication administration, were meticulously observed throughout the study.β1 (TGF-β1), and connective tissue growth factor (CTGF) were assessed both before and after treatment. Moreover, 1 week before and 2 weeks following the treatment, safety results and vital signs indices  were documented for each participant. In the meantime, adverse events (AEs) were continuously observed, precisely documented, and appropriately addressed by researchers throughout the trial.The primary assessment result was measured and represented as the alteration in the slope of serum creatinine (Scr). The secondary assessment outcomes were post-treatment alterations in blood urea nitrogen (BNU), uric acid (UA), estimated glomerular filtration rate (eGFR), urinary AKI biomarkers including neutrophil gelatinase associated lipocalin (NGAL) and interleukin-18 (IL-18), as well as TCM syndromes. Besides, levels of renal fibrosis biomarkers from serum samples, notably, transforming growth factor-Statistical analysis of the improvement in clinical symptoms score was performed in strict compliance with the Guidelines for Clinical Research of Chinese Medicine (New Drug) . The priEI = /Total symptom score before treatment × 100%The treatment efficacy was evaluated using EI. Symptom improvement levels were defined as follows: clinical control (EI ≥ 95%), significant effect (94% > EI ≥ 70%), and effectiveness (69% > EI ≥ 30%) to inefficacy (EI < 30%).α = 0.05 (one-sided test) and test efficacy 1–β = 0.90. Premised on a 20% drop-out rate, the predicted sample size for random sampling was 49 cases within every group for 98 cases in the study.In this study, we used baseline Scr change to estimate sample size. Premised on previous studies , we compRegarding the Full Analysis Set (FAS), which is made up of individuals who were randomly assigned into two groups, the principal analysis was carried out. Participants who successfully completed the research protocol and had good compliance were defined as the per-protocol sample. In conformity with the Intend-to-Treat (ITT) principle, all subjects receiving test medications were included in a safety analysis.t-tests or one-way ANOVA were applied for continuous data exhibiting normal distribution; in contrast, Pearson’s chi-square test (or Fisher’s exact test for cell count <5 in any cell) was employed for comparisons involving categorical variables. All statistical analyses were conducted using a two-sided design. p < 0.05 was established as a determinant of statistical significance. The analyses of statistical data were carried out by a biostatistician who was not engaged in the research with the aid of the SPSS (version 21.0) software package. There were no further analyses or interim analyses carried out in this study.Continuous variables were presented as mean ± standard deviation or by the median. In this case, categorical variables were displayed in the form of numbers and percentages. Student’s p > 0.05). It indicated that the baseline characteristics of two groups were balanced and the efficacy of this clinical study was comparable . Compared with RG group, Scr in RG + CHF group was lower and the difference was statistically significant (p < 0.05)  .p < 0.01). Besides, UA in RG + CHF group were lower and the difference was statistically significant (p < 0.05), eGFR was improved and the difference was statistically significant (p < 0.01)  .p < 0.05). Besides, IL-18 in RG + CHF group was lower and the difference was statistically significant (p < 0.01)  .β1 after treatment were 115.2 ± 21.6, 143.9 ± 25.3 in RG group and RG + CHF group respectively. Compared with RG group, TGF-β1 in RG + CHF group was lower and the difference was statistically significant (p < 0.05). Besides, CTGF in RG + CHF group was lower and the difference was statistically significant (p < 0.05)  .p < 0.05)  .p < 0.01)  and 6 (12.2%). While the significantly effective rate and effective rate were 24 (49.0%) and 13 (26.5%) in RG + CHF group. Total effective rate (75.5%) in RG + CHF group were obviously higher than that (36.7%) in RG group, and there was significant statistically difference between two groups ( < 0.01) .The AEs occurred in the RG and RG + CHF group were demonstrated below . There wCompared with RG group, more reductions of Scr, BUN, UA, and better improvement of eGFR were observed in RG + CHF group. Additionally, the levels of urinary AKI biomarkers, renal fibrosis biomarkers, and TCM syndromes were decreased in RG + CHF group versus RG group. RG + CHF group demonstrated better renal protective effects than RG subgroup.AKI is a clinical condition hallmarked by a sudden decrease of renal function, whereas CKD is hallmarked by renal functional or structural impairments. As a consequence of AKI, a significant number of oxygen free radicals are produced, endogenous antioxidants are continually depleted, and high levels of inflammatory substances are secreted, all of which contribute to kidney damage progression . MoreoveAccording to our published papers , the comLigusticum wallichii (Chuanxiong), has the potential to prevent AKI via a variety of processes, including ameliorating oxidative stress damage, suppressing inflammatory responses, deterring apoptotic cell death of intrinsic renal cells, and modulating autophagy . However, there is currently no recognized medication therapy strategy for AKI grades 1–2 . As indiPremised on our previously clinical evidence and recent breakthroughs in clinical therapy of A on C, we designed a new protocol with the combination of CHF and RG. And this study tried to explore the medical mechanism of CHF combining RG treating A on C from the aspect of renal fibrosis. The current research would be beneficial in resolving this clinical dilemma in the future. The combination might successfully minimize renal damage of A on C patients, enhance restoration of kidney functionality, and relieve clinical symptoms in these individuals. As a result, we consider that the clinical application values of this therapy method are satisfactory. Based on the findings of this study, we will continue to explore therapeutic effects of CHF combining RG for patients with A on C to gather more relevant clinical data.CHF combining RG showed better therapeutic effects than RG alone, and its renal protective effects were associated with reducing Scr, BUN, UA, and improving eGFR, as well as preventing renal fibrosis. We thus concluded that this novel therapy of combining CHF and RG might be a useful method for treating A on C patients.


Aim: Hypotrichosis simplex (MIM 146520) is a rare form of monogenic hereditary alopecia. Several genes have been identified as being associated with the disease, including LPAR6, LIPH, and DSG4. LSS encoding lanosterol synthase (LSS) has been shown to cause hypotrichosis simplex, but the related mechanisms have not been elucidated to date. This study aims to find mutations in LSS from a Chinese family, among which a 21-year-old male patient and his 9-year-old sister were affected by hypotrichosis simplex.Methods: Dermoscopy and histological analysis were used to examine patients’ scalps, while exome sequencing was used to find the mutations in LSS.Results: The hair loss was only detected on the scalp of the proband and his sister, while other ectodermal structures were normal with no systemic abnormalities. Further, the exome sequencing identified a new homozygous mutation NM_002340.6 (LSS_v001):c.812T>C (p.(Ile271Thr)) in the LSS gene of the proband, which was also found in his sister. In addition, a heterozygous mutation of LSS was found in their asymptomatic parents. Finally, the possible protein structure of the mutational LSS was predicted.Conclusion: The hypotrichosis simplex reported here could be an autosomal recessive disease in this family. The mutation on LSS might reduce the enzyme activity of LSS, thus leading to the disease. LPAR6, LIPH, and DSG4 (S)-2,3-epoxysqualene to lanosterol. Biallelic mutations in LSS were first reported in families with congenital cataracts  is a rare form of monogenic alopecia. Patients with hypotrichosis simplex usually have normal hair at birth and begin to lose scalp and body hair in early childhood . Severaland DSG4 . Normallataracts . More re disease . NeverthLSS was a causative gene for alopecia and cataracts  was used to examine patient scalps. Skin samples from patients were fixed with 4% paraformaldehyde solution and embedded in paraffin, then cut into 5 μm sections. Hematoxylin and eosin (H&E) staining was performed according to the manufacturer’s instructions .https://genome.ucsc.edu/). Base quality score, single nucleotide variants (SNVs), and indel were analyzed by GATK (https://gatk.broadinstitute.org/hc/en-us). The variants were prioritized based on the following filtering criteria: 1) The variations with minor allele frequencies (<0.1%) were identified from the following databases: exome aggregation consortium (ExAC), 1000 Genomes Project, and Genome Aggregation Database (Genome AD V3). 2) Synonymous and non-coding variants not predicted to affect splicing were excluded. 3) Variants with an embryogenesis-related function. 4) Location in or near a homozygous region higher than 2.0 Mb. The pathogenicity of each genetic variation was classified based on the Online Mendelian Inheritance in Man (OMIM) (https://www.omim.org/), Human Gene Mutation Database (HGMD) (http://www.hgmd.cf.ac.uk/ac/index.php), ClinVar database (https://www.ncbi.nlm.nih.gov/clinvar/), and the American College of Medical Genetics and Genomics (ACMG) and the Association for Molecular Pathology (AMP) guidelines.First, blood samples were collected from the proband. The genomic DNA of the peripheral blood was extracted according to standard procedures followed by sonication, and the fragments were subjected to end-repair and adaptor-ligation. After size selection, the library was constructed and purified by an MGIEasy Exome Capture V4 Probe Set and sequenced on an MGISEQ-2000. Reads were compared with the UCSC hg19 human reference genome by BWA . PCR products were purified by FastPure Gel DNA Extraction Mini Kit , and Sanger sequencing was performed . Primers used for PCR and sequencing were LSS_cds8F: AATGAGCTTCTGGGCAGACC and LSS_cds8R: CTCTGAGCTCCTCCTTCCCT.UCSF Chimera software was used to visualize 3D models and to predict the consequences of the mutated residues . The LSSThe proband was a 21-year-old man who presented with sparse scalp hair. He had normal hair at birth but started to lose his hair at 6 months old and has had hair shorter than 0.5 cm since he was 1 year old. Physical examination revealed lightly pigmented hair with slightly reduced strength . OtherwiDermoscopy on the proband’s scalp showed that the patient had diffuse hair thinning without any morphological abnormalities. In addition, most of his hair follicles contained only a single hair, and vellus hairs were few with thin hair shafts and light color. No black dots, broken hairs or exclamation mark hairs could be found. Histological examination showed that epidermal thickness was normal, and the basement membrane was intact despite the decreased number of hair follicles, shown in LSS gene, NM_002340.6 (LSS_v001):c.812T>C (p.(Ile271Thr)), was identified and confirmed by Sanger sequencing (https://rarediseases.info.nih.gov/diseases/9170/hypotrichosis-simplex). In addition, different LSS mutations have been found in patients affected by hypotrichosis simplex previously :c.812T>C (p.(Ile271Thr)) in the LSS gene of the patient and their sister. A protein structure comparison suggested this mutation might reduce LSS enzyme activity. In conclusion, the hypotrichosis simplex reported here could be an autosomal recessive disease in this family, and the mutation on LSS might reduce LSS enzyme activity, thus leading to alopecia. In the future, the mice model can be utilized to further explore the relationship and mechanisms between the LSS mutation and hypotrichosis simplex.This study aimed to find new mutations in


Morris water maze test (MWM) were used to assess the PDD-like symptoms in rats. Also, the pathological manifestations and mechanisms of BCCAO impairing cognitive functions have been explored via hematoxylin-eosin staining, Nissl staining, immunohistochemistry, immunofluorescence, RNA sequencing analysis, lipidomics, and quantitative real-time polymerase chain reaction. In this study, we found that CCH could aggravate PDD-like cognitive symptoms  and PDD-like pathology . Moreover, a potential relationship between differentially expressed mRNAs and lipid metabolism was revealed by RNA sequencing analysis. Lipidomics showed that CCH could affect the intensity of 5 lipids, including sphingomyelin , cardiolipin, lysophosphatidylcholine, cholesteryl ester, and triacylglycerol. Interestingly, the KEGG pathway analysis of both RNA sequencing analysis and lipidomics suggested that CCH leaded to learning impairment by affecting sphingolipid metabolism. Finally, we found that CCH disrupts the sphingolipid metabolism by affecting the mRNA expression of SMPD1 and SMS2, leading to the accumulation of sphingomyelin in the prefrontal cortex. In summary, CCH, an independent exacerbating reason for impairment in learning and memory within the pathopoiesis of PD, aggravates Parkinson's disease dementia-like symptoms and pathology in 6-OHDA-lesioned rat through interfering with sphingolipid metabolism.Chronic cerebral hypoperfusion (CCH) is a cardinal risk factor for Parkinson's disease dementia (PDD), but this potential causality lacks mechanistic evidence. We selected bilateral common carotid artery occlusion (BCCAO) to simulate chronic cerebral hypoperfusion in the rat model of PD induced by typical neurotoxin 6-hydroxy dopamine (6-OHDA). Four weeks after unilateral injection of 6-OHDA into the medial forebrain bundle, rats underwent BCCAO. Male Sprague-Dawley rats were divided into five groups of ten, including sham, PD+BCCAO 2 weeks, PD+BCCAO 1 week, PD, and BCCAO 2 weeks. Then, open field test (OFT) and  At presseverity , 5. α-Syl cortex . On the l cortex , 8. Sevel cortex , 10. DesMorris water maze test (MWM). Furthermore, the pathological manifestations and mechanisms of BCCAO impairing cognitive functions have been explored via immunohistochemistry (IHC), immunofluorescence (IF), ribonucleic acid (RNA) sequencing analysis, lipidomics, and quantitative real-time polymerase chain reaction (qRT-PCR). Our results would provide new ideas for the study of mechanism of CCH enhancing PDD-like symptoms and pathology.Bilateral common carotid artery occlusion (BCCAO) is a common method to induce CCH in rat , 12. TheMale Sprague-Dawley rats (200-220 g) were purchased from Experimental Animal Center of Guangzhou University of Chinese Medicine  and maintained in cages under standard conditions with a 12 h light/dark cycle with free access to food and water. All experiments were carried out in accordance with the Guide for the Care and Use of Laboratory Animals and were approved by the Ethics Committee for Animal Research of Guangzhou University of Chinese Medicine (No. 20210318006). For all experiments, rats were acclimated to their surroundings for 3 days in the animal room prior to surgery, and they had free access to food and water. Then, rats were divided into five groups of ten. Body weight was measured once a week. The intervention methods of rats in each group are shown in μL of 6-OHDA  solution  was injected into the right MFB at a rate of 1 μL/min. Rats injected with the same volume of saline containing 2 mg/mL ascorbic acid were used as the sham group. Apomorphine  solution  was subcutaneously used to induce the spontaneous rotational movement in rat 4 weeks after model establishment and before sacrificing. The number of rotations contralateral to the lesion during 30 min after apomorphine injection was evaluated [The rat model of PD was established by unilateral injection of 6-hydroxy dopamine (6-OHDA) into MFB as previously described with minor modifications . Rats wevaluated .Rats were anaesthetized with isoflurane and treated with carprofen as an analgesic. Also, surgery was carried on a heating pad to prevent hypothermia. BCCAO was performed as described in the literature . A midliThe autonomic motor of rats was assessed by OFT, and spatial memory capability of rats was assessed by MWM test as previously reported , 19. TheμM thick coronal sections using a RM2255 microtome , mounted on glass slides. Distribution of neural cells in the rat prefrontal cortex and striatum was assessed by microscopy following standard hematoxylin-eosin (HE) staining . Nissl bodies in rat brain tissues were detected by a previously described Nissl staining method  [The animal used for histological evaluation were perfused, firstly with 0.9% saline and then with cold 10% paraformaldehyde. The brains were removed, washed, and protected in the 10% paraformaldehyde. Formalin-fixed brains were dehydrated in a series of graded ethanol and xylene baths and embedded in paraffin. Brains were cut into 6 , China) .After dewaxing and rehydrating and antigen retrieval, the paraffin sections were blocked endogenous peroxidase activity by citric buffer (Pondus Hydrogenii (PH) 6.0). Brain sections were blocked with normal goat serum for 1 h at room temperature. The primary antibody was diluted in phosphate buffer saline (PBS) (PH 7.4) in a certain proportion (tyrosine hydroxylase (TH), Proteintech, 1 : 200) and is added to the sections, and the sections are placed flat in a wet box and incubated overnight at 4°C. Following three washes with PBS, rat brain sections were incubated in the dark with green fluorophore-conjugated secondary antibodies for 1.5 h at room temperature. After counterstaining with 4′,6-diamidino-2-phenylindole dihydrochloride (DAPI), sections were mounted and observed by fluorescence microscopy.α-Syn, Abcam, 1 : 16000; Aβ, Abcam, 1 : 200) and is added to the sections, and the sections are placed flat in a wet box and incubated overnight at 4°C. After the sections are slightly shaken and dried, the tissues are covered with secondary antibody (horseradish peroxidase labeled) from the corresponding species of primary antibody and incubated at room temperature for 50 minutes. 3,3-N-diaminobenzidine tertrahydrochloride (DAB) color developing solution newly prepared is added in the circle after the sections are slightly dried. After counterstaining nucleus, the brain sections were dehydrated and mounted. Then, sections were observed by microscopy.The steps for dewaxing and rehydrating, antigen retrieval, blocking endogenous peroxidase activity, and serum sealing are similar to immunofluorescence. The primary antibody was diluted in PBS (PH 7.4) in a certain proportion (http://ccb.jhu.edu/software/hisat/index.shtml) and Bowtie2 (http://bowtie-bio.sourceforge.net/bowtie2/index.shtml) to align clean reads to the reference genome sequence  to get the alignment results. Differential expressions of mRNAs between the two groups were defined as the combination of a log2 Ratio (PD+BCCAO 2 weeks/PD) value of ≥1 and a P value ≤ 0.05 based on reads per kilobase million using DESeq2 software. Hierarchical clustering and scatter plots of differentially expressed mRNAs were constructed using R language. Enrichment of Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways for differentiation expressed mRNAs was performed using the Database for Annotation, Visualization, and Integrated Discovery system . The GO and KEGG pathways with P values (P) < 0.05 were considered significant.Differential expressions of messenger RNA (mRNA) in prefrontal cortex of the sham group and PD+BCCAO 2 weeks were characterized by RNA sequencing (RNA-Seq). Total RNA was prepared from prefrontal cortex using TRIzol reagent  according to the manufacturer's instructions. The concentrations of RNA were assessed by NanoDrop 2000 spectrophotometer , and the quality of RNA was analyzed calculating RNA integrity number with Agilent Bioanalyzer 2100 (Agilent Technologies). Transcriptome sequencing was performed on RNA samples that passed the quality control indicators (2100 RNA integrity number (RIN) ≥ 7.0 and 28S/18S > 1.0). Subsequently, the RNA-Seq libraries were constructed and sequenced on the DNBSEQ platform . In order to display the number of genes in different expression intervals of each sample more intuitively, we carried out statistics on the number of genes in three cases of expression amount . Following adapter trimming and other processing, we used HISAT . An ACQUITY UPLC BEH C18 column  was used for the reversed phase separation. The column oven was maintained at 35°C. The flow rate was 0.4 mL/min, and the mobile phase consisted of solvent A  and solvent B . Gradient elution conditions were set as follows: 0~0.5 min, 5% B; 0.5~7 min, 5% to 100% B; 7~8 min, 100% B; 8~8.1 min, 100% to 5% B; and 8.1~10 min, 5%B. The injection volume for each sample was 4 μL. A high-resolution tandem mass spectrometer Q-Exactive  was used to detect metabolites eluted from the column. The Q-Exactive was operated in both positive and negative ion modes. Precursor spectra (70–1050 m/z) were collected at 70,000 resolution to hit an AGC target of 3e6. The maximum inject time was set to 100 ms. A top 3 configuration to acquire data was set in DDA mode. Fragment spectra were collected at 17,500 resolution to hit an AGC target of 1e5 with a maximum inject time of 80 ms. In order to evaluate the stability of the LC-MS during the whole acquisition, a quality control sample  was acquired after every 10 samples. MetaX software was used to quantify and screen the differential metabolites. Those features that were detected in less than 50% of QC samples or 80% of biological samples were removed, the remaining peaks with missing values were imputed with the k-nearest neighbor algorithm to further improve the data quality. PCA was performed for outlier detection and batch effect evaluation using the preprocessed dataset. Quality control-based robust LOESS signal correction was fitted to the QC data with respect to the order of injection to minimize signal intensity drift over time. In addition, the relative standard deviations of the metabolic features were calculated across all QC samples, and those >30% were then removed.The collected samples in the prefrontal cortex were thawed on ice, and metabolites were extracted with 50% methanol buffer. Pooled quality control (QC) samples were also prepared by combining 10 https://hmdb.ca/) and KEGG, to explain the physicochemical properties and biological functions of metabolites. Student's t-tests were conducted to detect differences in metabolite concentrations between 2 groups. The P value was adjusted for multiple tests using an false discovery rate (Benjamini–Hochberg). Supervised PLS-DA was conducted through MetaX to discriminate the different variables between groups. The variable important in the projection (VIP) value was calculated. Differential metabolic ions between the two groups were defined as the combination of a ratio ≥ 1.5 or ≤1/1.5, a P value ≤0.05, and VIP ≥ 1. Enrichment of KEGG pathways for differentiation lipids was performed using the DAVID.The raw data of mass spectrometry was converted into readable data mzXML using MSConvert software. Peak extraction was performed using XCMS software, and peak extraction quality control was performed. The extracted substances were annotated by summation ions using CAMERA and then identified by MetaX software. The MS primary information was used for identification, and the MS secondary information was matched to an in-house standard database, respectively. Metabolite annotation was performed on candidate identification substances using databases such as HMDB  was synthesized from 500 ng of RNA from each group by reverse transcription using a RevertAid First Strand cDNA Synthesis Kit . In CFX connect Real-Time System , a GoTaq qPCR Master Mix kit  was used for quantitate expression levels by quantitative PCR. Relative expressions of mRNAs were calculated using the standard 2plicates . Also, βP value of the Shapiro-Wilk test > 0.05) were presented as the mean ± standard deviation and analyzed by one-way analysis of variance (ANOVA). Data that did not conform to a normal distribution were presented as median (mix-max) and analyzed by nonparametric tests. P < 0.05 was considered significant. GraphPad Prism 6.0 software was used to draw images.Statistical tests were carried out using SPSS 26.0 software. Mauchly's test of sphericity was used to analyze repeated measurement data. Data considered to conform to a normal distribution   and decomposition (SMS2) of sphingomyelin. Compared with the sham group and PD group, we found the relative expression of SMPD1 was significantly decreased in the prefrontal cortex of PD+BCCAO 2 weeks group, but SMS2 was significantly elevated, while SMPD2 did not change Figures . These rAccording to the diagnosis and treatment guidelines, the two important symptoms of PDD are motor deficits  and cognitive deficits in memory, attention, executive function, and visuoconstructive ability . Meanwhiα-Syn and Aβ  and sphingomyelin synthase 2 , respectively. Thus, high-level of sphingomyelin might be related to SMPD1, SMPD2, and SMS2. Our results showed that mRNAs of low-expressed SMPD1 and high-expressed SMS2, leading to accumulation of sphingomyelin in the prefrontal cortex . On the function , 42. On of α-Syn . Mutatioysosomes , 45. Theα-Syn, which is related to cognitive functions. Thus, maintenance of adequate cerebral perfusion and stable sphingolipid metabolism may be critical to prevent further cognitive impairment in PD patients.In present study, the effects and mechanisms of CCH on PDD-like symptoms and pathology were evaluated in the classical PD rat model of 6-OHDA-lesions. In summary, we found that CCH is an independent exacerbating reason for impairment in learning and memory within the pathopoiesis of PD. Also, CCH disrupts the sphingolipid metabolism by affecting the mRNA expression of SMPD1 and SMS2, leading to the accumulation of sphingomyelin in the prefrontal cortex. Elevated levels of sphingomyelin could aggravate the aggregation and expression of


Dear editor,Vaccines against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have become the most vital countermeasure against the coronavirus 2019 (COVID-19) pandemic. However, numerous complications have been reported, such as fever, headache, myalgia, and rash, after receiving an anti-COVID-19 vaccine [Thrombocytopenia probably occurred in many causes in dialysis patients . A speciAs VITT and HIT are both mediated by platelet-activating antibodies against PF4 , VITT li3 to 450,000/mm3) as indicated in routine blood tests before the second dose of the AZ vaccine. Notably, the patient did not suffer from any thrombocytopenia or thromboembolism but only mild fever and myalgia after his first dose of the AZ vaccine, which was administered 3 months prior to the second dose. In addition, there was no previous history of any autoimmune or thrombotic disease.An 85-year-old male with a history of end-stage renal disease undergoing regular HD with low-molecular-weight heparin since 2016, presented to the emergency department with a headache 7 days after a second dose of the AZ vaccine . Tracing back his medical history, his serum platelet counts had been within normal limits  on the 7th day after vaccination. Given the unexplained thrombocytopenia, he was admitted to our hematology ward for further survey.On physical examination, no neurologic deficits were evident and computed tomography of the brain was unremarkable . A compl3 and a D-dimer peak greater than 4.5 mg/L. Nevertheless, the prothrombin time (9.5 s), activated partial prothrombin time (26.1 s) and international normalized ratio (0.78) were within normal ranges. He was afebrile and leukocytosis was not found . Both Helicobacter pylori. and virus screening gave unremarkable results. Stable hemoglobin levels (10.1 g/dL), preserved ADAMTS 13 activity and no schistocytes found on his blood smear did not support atypical hemolytic uremic syndrome or thrombotic thrombocytopenia (TTP). Serum antinuclear antibody and antiphospholipid antibody were also examined, and evidence was insufficient to support the possibility of rheumatic disease or other immunologic disease. His medication history was reviewed and other drug-related thrombocytopenia was excluded. An abdominal ultrasound showed no evidence of liver cirrhosis and no splenomegaly. Given his symptoms, vaccination history, dialysis history and the laboratory findings, VITT or HIT was highly suspected. The autoantibody of PF4 was checked by enzyme-linked immunosorbent assay (ELISA) on his hospital admission day and found to be positive with an optical density (OD) value of 0.428 (positive as OD >0.4), confirming our hypothesis.A review of his dialysis chart indicated that heparin was suspended due to the development of gastrointestinal bleeding, 3 weeks prior to his second vaccination dose. After the bleeding episode resolved, unfractionated heparin was administered 16 days later. As thrombocytopenia occurred 7 days after the second dose vaccination, non-heparin HD was performed after hospital admission. No obvious thrombotic event was recorded, apart from one time of dialysis circuits clotting in trace status after admission. A series of examinations were done . Initial3) within 4 weeks and no clinical evidence suggested that thrombosis was increasing. The pattern of laboratory parameters observed along with clinical regimens are shown in Initially, he was treated with intravenous (IV) methylprednisolone  for a week. Due to poor response, treatment of IV immune globulin (2 mg/kg/daily for 5 days) was initiated, and combined with dexamethasone (5 mg daily). Anti-PF4 antibody level was rechecked and showed an OD value of 0.702. Given that the platelet counts did not rise to an acceptable level despite previous treatment, oral cyclophosphamide (50 mg daily for 10 days) was prescribed as an immunosuppressant. IV dexamethasone was continued in tapering doses. The platelet count was normalized , a D-dimer level greater than 4 mg/L and the presence of antibodies to PF4 detected, but the fifth criterion, thrombosis, was not documented. The case was classified as a highly probable VITT based on these criteria. Other possible causes of thrombocytopenia, including previous infection, thrombophilia, TTP, autoimmune disease and other medications were specifically excluded.The epidemiology of VITT is rapidly evolving, with several formulated guidelines for diagnosis and management under development. The case we present fulfilled four of the five diagnostic criteria from the UK Hematology Expert Groups , includSupplementary Item S2, [For thrombocytopenia in patients on HD, HIT must be a differential diagnosis. Dialysis patients are at risk of HIT 7–10 days after receiving heparin, with clinical presentations of acute thrombocytopenia and thrombotic events . The widItem S2, ). TreatmItem S2, . In our As shown in this case, a rapidly progressive thrombocytopenia developing on dialysis patients after the AZ vaccination may clinically mimic HIT. Our findings expand upon prior reports in which anti-PF4/heparin antibodies, referred to as heparin-induced antibodies (HIA), were shown to be associated with both VITT and HIT . In addiAll current treatment recommendations for VITT are based on extrapolation from autoimmune thrombotic thrombocytopenia. The consensus of most experts is that a second-line therapy of immunomodulators be applied when first-line therapy (e.g. steroids) does not raise the platelet count to a safe level. Since our study indicated a beneficial effect of cyclophosphamide, we suggest it could be considered in severely-affected patients who do not respond to standard therapy. The optimum treatment schedule of cyclophosphamide remain to be determined.It is challenging to prove definitive causality in the case we present since VITT is a novel syndrome and not yet fully understood. An additional survey of thrombosis or microthrombosis could further verify a definitive diagnosis for VITT.VITT has emerged as a rare but devastating complication. The presented case highlights the need for timely recognition of VITT to prevent devastating complications in an HD patient receiving an anti-COVID-19 vaccination.Click here for additional data file.


To determine whether dizziness can contribute to stroke as a main cause still remains challenging. This study aims to explore clinical biomarkers in the identification of ischemic stroke patients from people with dizziness and the prediction of their long-term recovery.+ T cells, CD8+ T cells, T−/− cells (DNTs), CD4+ regulatory T cells (Tregs), CD8+ Tregs, B cells and regulatory B cells (Bregs) were examined to identify biomarkers with clinical value.From January 2018 to June 2019, 21 ischemic stroke patients with a main complaint of dizziness, 84 non-stroke dizziness patients and 87 healthy volunteers were recruited in this study. Then, their peripheral blood samples were collected, and the percentages of circulating lymphocytes T cells, CD4p = 0.0009). The Bregs proportion in ischemic stroke patients with dizziness was lower than that in non-stroke dizziness patients (p = 0.035). In addition, the percentage of Bregs and DNTs within lymphocytes in patients’ peripheral blood exhibited a significant negative correlation with stroke occurrence . Moreover, the Bregs and DNTs within lymphocytes were negatively related to participants’ age, while presented a weak relationship with clinical risks like smoking, hypertension, and diabetes. Then, area under the receiver operating characteristic curve (AUC) of Bregs and DNTs together was 0.768, the risk factors and Bregs or DNTs ranged from 0.795 and 0.792, respectively, and the AUC value of risk factors, Bregs and DNTs combination was further increased to 0.815. Furthermore, the Bregs percentage within lymphocytes at admission was also a potential predictor of repair at discharge and the following 3 months.According to our data, a significant difference in the DNTs proportion was detected between non-stroke dizziness and ischemic stroke patients with dizziness (Bregs and DNTs could be the clinical biomarkers together in the identification of ischemic stroke patients from people with dizziness. As a major cause of mortality and morbidity worldwide, stroke has been prioritized by World Health Organization in their efforts to reduce the burden of non-communicable diseases , 2017. SThe immune response plays a key role in the pathophysiology of ischemic stroke . PreviouHerein, the aim of the present study is to analyze the characteristics of non-stroke dizziness patients and stroke patients with dizziness, and investigate the difference of the peripheral blood lymphocyte subpopulations in the two groups for reliable biomarkers in the identification of ischemic stroke patients from all dizziness patients.The clinical study was approved by the Ethical Review Board of Central Hospital of Minhang District . From January 2018 to June 2019, 21 stroke patients with a chief complaint of dizziness, and 84 patients with non-stroke dizziness were included according to following criteria: (1) more than 18 years old; (2) stroke patients with a final diagnosis of “cerebral infarction” confirmed by head CT or MRI; (3) non-stroke dizziness patients with a final diagnosis of “dizziness and vertigo”; (4) first onset; (5) onset time no more than 5 days; (6) informed consent was obtained from the patients or their legal representatives. Patients with brain hemorrhage, neoplasms, or acute infections after stroke were excluded. Clinical information of stroke patients was collected at admission, including age, gender, risk factors . Control subjects were 87 age- and gender-matched healthy volunteers without a history of stroke or other vascular diseases.g and 20°C, PBMCs were finally collected from the plasma-Ficoll interphase.After the informed consent was obtained from participants or their legal representatives, peripheral blood samples were collected through forearm veins from the participants, and Ethylene Diamine Tetraacetic Acid (EDTA) vacutainer blood collection tube was used to keep those samples. For non-stroke dizziness and stroke patients, blood samples were collected within 24 h from admission. For healthy volunteers, blood samples were collected when they attended the physical examination. PBMCs were isolated by density gradient centrifugation within 3 h after receipt of peripheral blood samples. Technically, the blood samples were firstly diluted in a 1:1 ratio with PBS buffer, and then layered over Ficoll-Paque  in a 15 ml centrifuge tube. After 30 min of centrifugation at 4006–107 cells were labeled with fluorochrome-conjugated antibodies (BD Biosciences), including FITC Mouse Anti-Human CD3, APC-H7 Mouse Anti-Human CD4, APC Mouse Anti-Human CD25, Percp-cy5.5 Mouse Anti-Human CD19, PE Mouse Anti-Human CD24, PE Mouse Anti-Human CD127, APC Mouse Anti-Human CD27, Percp-cy5.5 Mouse Anti-Human CD8, PE Mouse Anti-Human CD45RA, APC Mouse Anti-Human CD62L, and PE-cy7 Mouse Anti-Human CD183. All samples were run on LSRFortessa (BD Biosciences) using FACSDiva software (version 8.0), and FlowJo (version 9.9.6) was used to analyze the data.Trypan blue exclusion method was used to quantify PBMCs, then 10U test and Spearman’s correlation analysis as appropriate. The statistical significance of differences between experimental groups was assessed with the Brown-Forsythe and Welch ANOVA tests. All above statistical analyses were performed with GraphPad prism software (version 9.0). A forward stepwise binary logistic regression was performed with SPSS 25.0 software package . Variables showing significant association with stroke occurrence in univariate analysis (p < 0.1) and important confounders were included in the multivariate logistic regression analysis. Area under the receiver operating characteristic curve (AUC) was calculated and visualized with R-3.5.2 softwarepROC (receiver operating characteristic curve). If not specified above, a two-tailed p < 0.05 was considered statistically significant.Patient characteristics and experimental data were presented as means ± standard deviations (SDs) or numbers (%). Correlations between cell proportion and multiple characteristics were analyzed using Mann–Whitney p = 0.23), diabetes (p = 0.30), smoking (p = 0.08), and alcohol consumption (p = 0.17). Furthermore, no significant group difference existed between the stroke patients and non-stroke dizziness patients in the laboratory tests, namely white blood cell count (p = 0.42), neutrophil proportion (p = 0.42), lymphocyte proportion (p = 0.43), monocyte proportion (p = 0.05), eosinophil granulocyte proportion (p = 0.84) and basophil granulocyte proportion (p = 0.72). The specific final diagnosis and detailed radiological characteristics of stroke patients were described in A total of 105 patients and 87 healthy controls were enrolled in our study, whose demographic characteristics were described in + cells (B cells), CD3+ cells (T cells), CD3+CD4+ cells (CD4+ T cells), CD3+CD8+ cells (CD8+ T cells), CD3+CD4−CD8− cells , and immunoregulatory lymphocytes, namely CD19+CD24+CD27+ cells (Bregs), CD3+CD4+CD25+CD127− cells (CD4+ Tregs) and CD3+CD45RA−CD8+CD183+CD62L+ cells , T cells , CD4+ T cells  and CD8+ T cells , while there was no statistically difference between non-stroke dizziness patients and health controls . However, there was no group difference found in CD4+ Tregs  and CD8+ Tregs . Similarly, the percentage of DNTs within lymphocytes also showed a significant negative relationship with stroke occurrence (p = 0.05). These results suggested that the lower level of Bregs or DNTs in dizziness patients at admission, the higher likelihood of stroke occurrence.To further explore the potential value of regulatory lymphocytes in distinguishing stroke patients from dizziness patients, 5 independent variables  were included in the forward stepwise binary logistic regression analyses. As shown in the The occurrence and progression of stroke are associated with multiple factors, including age, gender, and basic diseases. Blood leukocytes are also known to be influenced by a series of diseases or lifestyle habits. Thus, the relationships between regulatory lymphocytes and clinical risks were further assessed in order to clarify the dependability of Bregs and/or DNTs as biomarkers in predicting stroke occurrence. As shown in the In order to evaluate the dependability of Bregs and DNTs in distinguishing stroke patients from non-stroke dizziness patients, the percentages of these two cells along with the clinical features were used to calculate ROC curves and AUC. The AUC of the Bregs and DNTs combination was 0.77, which was larger than that of clinical risk factors that related to stroke including age, gender, past history of hypertension, diabetes, habits of alcohol consumption and smoking (AUC = 0.72). Additionally, the AUC values of risk factors combined with Bregs or DNTs were 0.795 and 0.792, respectively. Moreover, the combination of Bregs, DNTs and clinical risk factors further elevated the AUC to 0.82 . These sR = −0.45, p = 0.04; mRS: R = −0.5, p = 0.04] and the 3rd month , while DNTs were not related the repair process.The values of Bregs and DNTs in predicting neural repair of stroke patients were further evaluated by Pearson correlation analysis . The resAccording to previous studies, dizziness is a common reason for the emergency department (ED) visit, and stroke patients only account for about 3%–5% of all ED patients with dizziness . Accurat+ Tregs and DNTs between patients and controls, but also the differences of Bregs and DNTs between non-stroke dizziness and stroke groups were statistically significant.Routine blood test is an easy and convenient test in clinical practice. In the present study, 84 non-stroke dizziness patients and 21 stroke patients with dizziness were recruited, and the clinical data including white blood cells count, percentage of neutrophils, lymphocytes, monocytes, eosinophils and basophils were collected for further analysis. Although a significant difference of these inflammatory cells was found between non-stroke dizziness patients and controls, and stroke patients and controls, no significance difference existed between non-stroke dizziness patients and stroke patients. These results revealed that these inflammatory cells could reflect the unhealthy state of patients, but were unable to distinguish non-stroke dizziness and stroke dizziness. Then, we further collected the peripheral blood samples of patients within 24 h at admission after onset, and used flow cytometry to detect the change of different lymphocytic subpopulations. Notably, the changes in lymphocytic subpopulations were significant. Not only the differences of B cells, Bregs, CD8Bregs are a subset of B cells that secrete interleukin-10 IL-10; . Recent Further, we explored the relationship of regulatory lymphocytes, stroke occurrence and clinical risks. The results indicated that both Bregs and DNTs exhibited a significant negative relationship with stroke occurrence. It means that the lower the level of Bregs or DNTs, the higher the risk of stroke in those patients with a chief complaint of dizziness. Additionally, Bregs or DNTs had a weak correlation with smoking, hypertension and diabetes, while they were closely related to age and alcohol consumption. As alcohol consumption status was unrecorded in controls and the percentage of alcohol consumption person was very low in both non-stroke dizziness and stoke patients, their relationship was not further explored. As for age, Bregs and DNTs showed a negative relationship with it, which indicates that young dizziness patients with low level of Bregs and/or DNTs are high-risk group. In addition, we assessed the abilities of Bregs and DNTs in predicting neural repair of stroke patients. The results indicated that the percentage of Bregs within lymphocytes at admission was a potential predictor of neural repair at discharge and the following 3 months. Further studies are needed to investigate the underlying mechanisms of Bregs in affecting the long-term prognosis of ischemic stroke patients.Next, ROC curves reveled that both Bregs and DNTs combined with the clinical risk factors can be used to distinguish non-stroke dizziness and stroke patients with dizziness to some extent. Previous studies indicated that clinical risk factors related to stroke included age, gender, past history of hypertension, diabetes and habits of alcohol consumption and smoking . In presThis study has some limitations. First, only lymphocyte levels in the peripheral blood were analyzed in the present study, not including lymphocyte levels in the brain. The evaluation of peripheral markers can mask events coming from the central nervous system due to the contamination from systemic sources. Second, the stroke risk factors of healthy persons in control group are not recorded as they are volunteers without intact medical records. Third, the sample size of our study is small. Prospective studies with larger samples are needed to further validate our findings.Bregs and DNTs can be the clinical biomarkers for the diagnosis of ischemic stroke among dizziness patients, and Bregs could also be a potential predictor of neural repair. These findings will lay scientific foundation for Bregs and DNTs application in clinic, which facilitates the diagnosis and subsequent recovery of ischemic stroke patients.The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.The studies involving human participants were reviewed and approved by The Ethical Review Board of Central Hospital of Minhang District . The patients/participants provided their written informed consent to participate in this study.YW, JZ, and YG conceived and designed the experiments. YH and SL performed the experiments. YW and JL analyzed the data. YW and YL prepared the figures. YW wrote the paper. YW and YH revised the manuscript. All authors contributed to the article and approved the submitted version.This work was supported by the National Nature Science Foundation of China , the Chinese State Ministry of Science and Technology grants (2021ZD0201704), and Supported by Shanghai Municipal Science and Technology Major Projects (22ZR1413700 and 2018SHZDZX01), and Shanghai Center for Brain Science and Brain-Inspired Technology.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


We found that a significant increase in butyrate acid was observed in mouse NTDs brains. In addition, aberrant key crotonyl-CoA-producing enzymes acyl-CoA synthetase short-chain family member 2 (ACSS2) levels and lysine crotonylation (Kcr) were elevated high in corresponding low folate content maternal serum samples from mouse NTD model. Next, proteomic analysis revealed that folate deficiency led to global proteomic modulation, especially in key crotonyl-CoA-producing enzymes, and dramatic ultrastructural changes in mouse embryonic stem cells (mESCs). Furthermore, we determined that folate deficiency induced ACSS2 and Kcr in mESCs. Surprisingly, folic acid supplementation restored level of ACSS2 and Kcr. We also investigated overall protein post-translational Kcr under folate deficiency, revealing the key regulation of Kcr in glycolysis/gluconeogenesis, and the citric acid cycle. Our findings suggest folate deficiency leads to the occurrence of NTDs by altering ACSS2. Protein crotonylation may be the molecular basis for NTDs remodeling by folate deficiency.Maternal folate deficiency increases the risk of neural tube defects (NTDs), but the mechanism remains unclear. Here, we established a mouse model of NTDs  Neural tube defects (NTDs) are serious birth defects of the central nervous system caused by the failure of neural tube closure (NTC) during embryonic development. NTDs are the most common birth defect affecting population quality in China and countries worldwide. NTDs mainly occur in the spinal cord and brain (1991). Data from 2015 showed that NTDs occurred in 18.6 cases per 10,000 fetuses worldwide on average; ~ 50% of families chose to terminate the pregnancy, and 75% of NTD patients survived beyond the age of five . NTDs reThe metabolism of carbohydrates, lipids, and proteins is integrated through common intermediate metabolites, specifically the intermediate products of the convergence of two metabolic pathways: the tricarboxylic acid cycle (TCA) and bio-oxidation and then anabolism and catabolism. Current findings suggest that some intermediate metabolites, such as acetyl-CoA, A-ketoglutarate, and S-adenosylmethionine, serve as substrates or cofactors for enzymes that regulate histone modification . These iSome evidences imply that metabolic programs are related to epigenetic states in specific embryo development . More reIn this study, we used a quantitative proteomics approach to obtain a global view of the proteome and crotonylome alterations in response to folate deficiency in mouse embryonic stem cells (mESCs). Proteomics analysis showed that folate deficiency caused global proteomic changes in folate-free mESCs, particularly induced key crotonyl-CoA-producing enzymes ACSS2 and Kcr. We also investigated global protein post-translational Kcr under folate deficiency in mESCs, which revealed key regulatory roles of Kcr in glycolysis/gluconeogenesis, and the citrate cycle (TCA cycle). Furthermore, we found low folate levels and higher ACSS2 levels in the corresponding maternal serum samples in mouse NTD model. This finding was validated in folate deficiency-induced mouse NTD models, indicating that abnormal ACSS2 expression and crotonylation may participate in neural development and contribute to the occurrence of NTDs.via intraperitoneal injection of MTX (1.5 mg/kg) at E7.5 under low-folate diet conditions  were remarkably modulated in folate-free mESCs  and folate-free  through LC–MS/MS . The solns was 1 . The idens was 1 . In totans was 1 . To idenee mESCs . Heatmapee mESCs . To explp < 0.05; in vivo.In KEGG pathway enrichment, we found that most identified proteins are involved in amino sugar and nucleotide sugar metabolism, pentose phosphate pathway, arginine and proline metabolism, and cysteine methionine metabolism, as shown in n = 3) and normal folate mESCs (n = 3). High-performance liquid chromatography (HPLC) separation, immunoaffinity enrichment, and mass spectroscopy (MS) detection was applied to investigate Kcr substrates in folate-free mESCs. We identified 22,277 crotyl sites in 16,340 proteins, which constituted 70% of the whole mESC proteome (p < 0.05) compared with the normal folate group  than total protein lysine residues  based on the fold-change values . Using tTo learn more about the functions of the crotonylated protein, we performed pathway and KEGG analysis. For the upregulated and downregulated sites, the most enriched metabolism pathways shared by these crotonylated proteins were related to citrate cycle TCA cycle) and glycolysis/gluconeogenesis, respectively ,C. ExampA cycle ap < 0.05). The crotonylated lysine contexts generated 62 conserved motifs, as shown in In this study, 22,277 Kcr sites were recognized in 16,340 proteins through a quantitative proteomics approach. It expands the understanding of the Kcr proteome and provides a comprehensive mESC landscape of Kcr. In folate-free mESCs, we identified 354 down-regulated Kcr sites on 292 proteins and 149 up-regulated Kcr sites on 127 proteins  are caused by the genetics and environmental factors . Many stin vitro human embryonic stem cell differentiation  were purchased from the Jinmuyang Animal Laboratory. Sexually matured mice were mated overnight, and were separated and transferred to different cages at 8:00 am the next day, which was recorded as E0.5. On E7.5, female mice with vaginal plug detected were intraperitoneal injected with 1.5 mg/kg (body weight) methotrexate . On E13.5, mice were sacrificed under euthanasia. All routines to handle the animals complied with the care of laboratory animals’ guidelines. Other detail as produced in previously study .We sectioned the embedded paraffin sections and then performed immunohistochemical staining with primary antibodies for Kcr, ACSS2, acyl-CoA dehydrogenase (ACADS), and peroxisomal acyl-coenzyme A oxidase 1 (ACOX1). We incubated tissues with Kcr  ACSS2 , ACADS , and ACOX1  at 4°C for 12 h . Gray-scUPLC-ESI-MS/MS analysis was performed to qualitative and quantitative detection of SCFAs with 30 mg brain samples. Metabolite quantification was analyzed using MRM mode of triple quadrupole mass spectrometry. Data obtained from Mass spectrometry in different samples were chromatographic peaks and the peak areas were integrated on all chromatographic peaks. The peak areas denotes the relevant metabolites .Cell culture medium was prepared under aseptic conditions with 0.1 mM β-mercaptoethanol , 0.1 mM non-essential amino acids , 0.1 mM glutamate , 15% fetal bovine serum , 4 mg/l folate  and 1,000 U/ml mouse leukemia inhibitory factor . Folate treatments of ESC cultures as produced in our previously studies .m/z 100–1,700. The secondary spectra were collected in a scan mode termed the parallel cumulative serial fragmentation (PASEF) mode straight after a first-order MS collection. Charge numbers of the parent ions in the secondary spectra were ranged from 0 to 5. Collection repeated ten times. To avoid repeated scanning of parent ions, the dynamic exclusion time was set to 30 s. Data were processed in MaxQuant with the integrated Andromeda search engine (version 1.5). False discovery rate (FDR) thresholds for protein, peptide, and modification sites were specified at 1% for targets . Cutoffs for significant fold changes between cells by histone crotonylation were set as quantification ratios above 1.5 or below 0.67 . Mobile phase B was 100% acetonitrile solution with 0.1% formic acid. The liquid phase Gradients of the liquid phase were as follows: 0–70 min, 3–22% B; 70–85 min, 22–33% B; 85–87 min, 33–80% B; and 87–90 min, 80% B. The flow rate was set to 300 nl/min. Voltage of the ion source was 1.5 kV. Parent ions and their secondary fragments of the targets were detected in high-resolution time-of-flight mass spectrometer. The scanning range of secondary mass spectrometry was low 0.67 .The original mass spectrometry data were imported into the search database software, and the corresponding analysis parameters were set according to the experimental scheme. The secondary MS data was processed in Maxquant (V1.6.15.0). Trypsin/P was given to the digestion mode and only two missing cleavages were allowed. The smallest length of the peptide was limited to seven and the largest modification number of peptides was five. The mass error tolerance of primary parent ions in the First search and Main Search was set at 20 PPM and 20 PPM, respectively, and that of secondary fragment ions was set at 20 PPM. Fixed modification was positioned for alkylation of cysteine carbamidomethyl, variable modification for oxidation of methionine and n-terminal acetylation. The FDR value was 1% for protein identification and PSM identification. The database underwent further filtering to achieve high quality and trustworthy results. The FDR values of the spectrogram, peptide, and protein identification were all less than 0.01. Identified protein had at least one unique peptide .p value of 0.05 to be significant. We considered the sequence context to be the motif of the crotonylated peptide when the characteristic sequence number exceeded 20 and the p value was 0.000001. The sequence model composed of specific amino acids was analyzed by Motif-x (10 amino acids downstream and upstream of crotonylation sites). Once protein enrichment was completed, function x = 2log10 ran to convert the p value and one-way hierarchical clustering to category (Data bank successfully helped us to annotate Gene Ontology (GO) regarding cell compositions, biological processes, and molecular functions. GO annotation is to annotate and analyze identified proteins using eggnog-Mapper software (V2.0). The software is based on the EggNOG database, and the latest version is the fifth version, covering 5,090 organisms  and 2,502 viruses. Here, the GO IDS in the annotation results of each protein are extracted, and then the proteins are functionally classified in line with cellular components, molecular functions and biological processes. Protein pathways were annotated based on KEGG pathway database, and basic local alignment search tool (BLAST) comparison  was performed to identify proteins., The result of the highest score was chosen for annotation to compare results in each sequence. InterProScan software is used to annotate protein domain of recognized modified proteins. We used WoLF PSORT to annotate the subcellular location of the identified proteins. We considered categories with a category .p < 0.05 and used DAVID 6.8 to perform GO term enrichment for Tandem mass tagging-based proteomics analysis. The detnrichment.ab133664) and mouse anti-Kcr monoclonal antibody . Then, we incubated the membranes with the corresponding secondary anti-rabbit horseradish peroxidase-conjugated antibody  for 1 h at room temperature. Finally, we visualized the membrane using SuperSignal West Pico Chemiluminescence Substrate  and quantified protein bands using Quantity One software.Proteins extractions were separated on sodium dodecylsulphate (SDS) polyacrylamide gel through electrophoresis and transferred to a nitrocellulose membranes. The membrane was soaked in 5% milk to block the non-specific proteins for 5 min and incubated in the primary antibodies at 4°C overnight, including a mouse anti-ACSS2 monoclonal antibody  was used as permeabilizing reagents. Primary antibodies, including Kcr  and ACSS2 , were added to cells and incubated overnight at 4°C. Next, cells were incubated with TRITC-conjugated or FITC-conjugated secondary antibody  at 37°C for 1 h. Nuclei were stained with DAPI , and images were visualized with a Zeiss LSM 510 Element inverted confocal microscope.t-test was performed to give statistical significance. p value of < 0.05 was considered statistically significant and presented as *p< 0.05, **p< 0.01 or ***p< 0.001.All experiments with our final conclusion were completed in triplicate, and the data were presented as the mean ± standard deviation. The Student’s The datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found in the article/The study was conducted according to the guidelines of the management and utility of experimental animals and approved by the Capital Institute of Pediatrics Ethics Committee (DWLL2021011), and all animal manipulations were strictly performed following the relevant laws of China. Human samples were collected and analyzed in accordance with the Capital Institute of Pediatrics Ethics Committee approval (SHERLLM2021024).SW conceived and designed this study. SW, YZ, and PP participated in laboratory work. SW, YZ, FL, and XH performed the data analysis. SW and PP participated in the writing of the manuscript. SW and TZ participated in advising and revising the manuscript critically. All authors contributed to the article and approved the submitted version.This work was supported by the National Natural Science Foundation Projects (82071690 and 81971390), Research Foundation of Capital Institute of Pediatrics (FX-2020-05 and CXYJ-2-21-09), Beijing Hospitals Authority’s Ascent Plan (DFL20221102), Public Service Development and Reform Pilot Project of Beijing Medical Research Institute (BMR2021-3).The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.The reviewer QX declared a shared parent affiliation with the author FL to the handling editor at the time of review.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Dysregulation of adipogenesis is related to diabetic peripheral neuropathy (DPN) pathogenesis, which may be mediated by immune infiltration. Nevertheless, the expression patterns of multiple adipogenesis-related genes and the differences of immune infiltration in different lipid metabolism levels remain unknown. GSE95849, a gene expression matrix containing DPN patients and healthy participants, was downloaded from Gene Expression Omnibus (GEO) database. Differentially expressed adipogenesis-related genes (DEARGs) were screened by overlapping the adipogenesis-related genes with differentially expressed genes (DEGs). DPN patients from GSE24290 and GSE148059 were divided into two adipogenesis subgroups according to the expression of DEARGs. The single-sample gene set enrichment analysis (ssGSEA) was used to estimate the abundance of the immune cells between two subgroups. The analysis of immune infiltration suggested that a variety of immune cells and immune processes were elevated in the high expression group of DEARGs. The differentially expressed genes of the two subgroups were mainly enriched in biological processes and signaling pathways related to lipid metabolism. PPARG, FABP4, LIPE, FASN, SCD, DGAT2, PNPLA2, ADIPOQ, LEP, and CEBPA were identified as the hub genes of the two subgroups, whose related transcription factors (TFs) and miRNAs were predicted. An immunohistochemical assay was used to verify the expression of hub genes in DPN nerve tissues. Our comprehensive analysis of adipogenesis subgroups in DPN illustrated that different expression patterns of DEARGs may lead to different immune and inflammatory states. The identification of DEARGs may help to further distinguish the different characteristics of DPN patients and lay the foundation for targeted treatment. Our findings may bring a novel perspective to the diagnosis and treatment of DPN patients. The “limma” package was used to remove their batch effect. Finally, 112 DPN patients from GSE24290 and GSE148059 were merged as a dataset for further analysis.All the microarray bulk RNA sequencing data we used were obtained from Gene Expression Omnibus (GEO) database (https://www.gsea-msigdb.org/gsea/msigdb/), which is one of the most widely used and comprehensive databases of gene sets for performing gene set enrichment analysis. Log2|fold change (FC)| > 1 and adjust p value <0.05 were used as filter criteria to screen DEGs of GSE95849. Duplicated genes were excluded. Then, the intersection of adipogenesis-related genes and DEGs was analyzed by Venn diagram (FunRich 3.1.3), and 74 DEARGs were finally selected.Adipogenesis-related genes of the HALLMARK_ADIPOGENESIS gene set were obtained from the MSigDB Team (k was chosen according to the following criteria: (i) a higher intragroup correlation and a lower intergroup correlation; (ii) the cumulative distribution function (CDF) curve increased smoothly while the delta area increased gradually; (iii) no subgroups have a small sample size. According to the above criteria, k = 2 was chosen as the appropriate number of clusters. Principal component analysis (PCA) was analyzed by the “states” R package. The “pheatmap” R package was used to visualize the expression of DEARGs in two adipogenesis subgroups.The “ConsensusClusterPlus” R package was used to identify adipogenesis subgroups with an unsupervised clustering method according to the expression of DEARGs. The optimal value of Chronic inflammation and immune infiltration were considered important processes which were induced by excessive adipogenesis . To estip value <0.05 and false discovery rate (FDR) < 0.25 were deemed as significant difference between two subgroups.GSEA 4.2.3 was used for gene set enrichment analysis. The gene sets associated with 50 well-defined biological states or processes were obtained from Hallmark gene sets in MSigDB. A p value <0.05. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) were performed by the “clusterProfiler” R package using a q value <0.05 as statistically significant enrichment.To make a further understanding of the two subgroups, the “limma” package was performed to identify DEGs with |FC| > 1.5 and adjusted https://cn.string-db.org/) and Cytoscape (version 3.9.1) were used to analyze the interaction of DEGs. In order to find the hub genes among DEGs, the “cytoHubba” plugin of Cytoscape was utilized, and the top ten genes were chosen.Search Tool for the Retrieval of Interacting Genes (STRING) database ; monoclonal rabbit anti−FABP4 ; polyclonal rabbit anti−LIPE . The IHC assay was conducted as previously reported .t-test was used for comparison of variables that obey normal distribution while the Wilcoxon rank-sum test was used for nonnormally distributed variables. A two-sided p value <0.05 was deemed statistically significant. The “ggpubr” and “ggplot2” R packages were applied for visualization.R software (version 4.1.2) and Perl (version 5.16.2) were conducted to process, analyze and visualize data. Shapiro-Wilk test was applied for the normality test. The Student The analysis process of this study can be seen in Figure n = 66, 46 in cluster 1 and cluster 2, respectively). PCA analysis indicated that cluster 1 and cluster 2 were distinguished into two parts as shown in GSE24290 and GSE148059 were obtained from the GEO database. After merging these two datasets, the “limma” R package was applied to remove the batch effect and normalized the gene expression matrix Figure . PCA diaImmune infiltration and low-grade inflammation are both hallmarks of DPN . We firsα signaling, and reactive oxygen species pathway were enriched in cluster 2  via IHC . The resDyslipidemia is an important factor in DPN progression. Reduction of blood lipid and weight loss helps to delay the occurrence of DPN. More and more research suggested that adipogenesis can promote the development of DPN, but most of them focus on a single gene , 34, 35.α and IL-6, which were known as inflammatory factors [Excessive lipid synthesis often leads to chronic low-grade inflammation and immune infiltration, which may further cause other tissue damage. In pathological states such as obesity and metabolic excess, adipocytes often recruit more proinflammatory macrophages and other immune cells, and cytokines (such as TNF and IL-1) will significantly elevate to inhibit excessive adipogenesis . DPN is  factors . Persist factors . There w factors . The inf factors . Studies factors . In our  factors . A quant factors . An imba factors . Thus, aγ and is known to be important for ameliorating DPN. Besides, multiple drugs or small molecules could improve DPN by modulating the PPAR pathway [γ and then have an affection on DPN [γ is a key factor in regulating, at least some aspects of macrophage lipid metabolism. The study had shown that Brd4 binds to the promoter and enhancer of GdF3 to promote PPARγ-dependent expression of GdF3 in macrophages and modulated lipid metabolism and diet-induced obesity [γ ligand treatment and found that upregulation of these genes could be blocked by PPARγ-specific antagonist suggesting a PPARγ-dependent regulation [To further explore the application of adipogenesis subgroups in DPN, we identified ten hub genes between two subgroups. Relevant studies had shown that these hub genes were associated with DPN and related metabolism. Peroxisome proliferator-activated receptor gamma (PPARG) is a regulator of adipocyte differentiation, which can express PPAR pathway , 46, 47. pathway . Notablyn on DPN . Stearoyn on DPN . Lipase n on DPN . Metformn on DPN . Interlen on DPN . Fatty an on DPN . Interes obesity . In DCs,gulation . The rolgulation . In addigulation . SCD1 exgulation . LPS-indgulation . There wgulation . Anothergulation . Meanwhigulation . These hAlthough our findings seem encouraging, this study is still insufficient. First, the results of immune infiltration in adipogenesis subgroups were analyzed by ssGSEA, which uses a gene signature-based method to estimate the relative abundance of immune cells. Tissue-based flow cytometry may provide more accurate results. Besides, we applied immunohistochemistry to verify the expression of five hub genes in the DPN sciatic tissues, and the intensive mechanistic studies of these genes still need to be further improved in future studies. This will be the direction of our research in the future.Briefly, our integrated analysis of adipogenesis subgroups revealed the relationship between adipogenesis, immune cell, and inflammatory process and identified PPARG, FABP4, LIPE, FASN, SCD, DGAT2, PNPLA2, ADIPOQ, LEP, and CEBPA as auxiliary diagnostic indicators for adipogenesis subgroups in DPN. These findings emphasized the importance of adipogenesis and would provide a new perspective for immunotherapy of DPN patients. However, future studies with larger samples and clinical information using flow cytometry analysis are warranted to validate these findings.


The aging population is expected to reach 2 billion by 2050, but the impact of somatic symptom disorder (SSD) on the elderly has been insufficiently addressed. We aimed to clarify the prevalence of SSD in China and to identify physical and psychological differences between the elderly and non-elderly.In this prospective multi-center study, 9020 participants aged (2206 non-elderly adults and 6814 elderly adults) from 105 communities of Shanghai were included . The Somatic Symptom Scale-China (SSS-CN) questionnaire was used to measure SSD. Depressive and anxiety disorders were assessed by the Patient Health Questionnaire-9 (PHQ-9) and Generalized Anxiety Disorder-7 (GAD-7), respectively.p < .001) than the non-elderly. Comorbidity of depressive or anxiety disorders was 3.7 times higher than would be expected in the general population. Additionally, the results of adjusted multivariate analyses identified older age, female sex, and comorbid physical diseases as predictive risk factors of SSD in the elderly group.The prevalence of SSD in the elderly was higher than that in the non-elderly (63.2% vs. 45.3%). The elderly suffered more severe SSD (20.4% moderate and severe in elderly vs. 12.0% in non-elderly) and are 1.560 times more likely to have the disorder , the elderly in Shanghai are more vulnerable to have SSD and are more likely to suffer from comorbid depressive and anxiety disorders. SSD screening should be given more attention in the elderly, especially among older females with several comorbid physical diseases.The online version contains supplementary material available at 10.1186/s12888-022-03907-1. As the world population ages rapidly, elderly adults  now accPsychological problems are an important factor affecting the quality of life. Published in 2018, the “White Paper on Mental Health of Chinese Urban Residents” surveyed 1.13 million people from 556 medical institutions  on physical and psychiatric questions [Since the elderly have grown into a larger entity, a special focus on the older population is needed. Compared with the non-elderly, the elderly population has unique social and biological characteristics. Generally, the elderly have a higher prevalence of hypertension , diabeteTo the best of our knowledge, there is no research comparing the prevalence of SSD, depressive disorders, and anxiety disorders between elderly and non-elderly populations. Current data are not sufficient to determine the prevalence of SSD under DSM-5 criteria. As the clinical relevance of SSD in the elderly is uncertain, these studies are urgently required for the optimal allocation of medical resources and health management of elderly community members.To screen for SSD, we designed a prospective cross-sectional study to assess physical and psychological characteristics in elderly people from 105 communities in Shanghai, China. Differs from SSD-12 , we previously developed the Somatic Symptom Scale-China (SSS-CN) questionnaire, a somatic and psychological symptom scale based on the DSM-5 that can be used to assess a combination of psychological, behavioral, and somatic symptoms and which is designed to assess both presence and severity of symptoms . We valiThis prospective cross-sectional study  is a multicenter registry conducted in Shanghai, China, under the supervision of the Shanghai Association of Chinese Integrative Medicine.A multistage and stratified systematic sampling technique was used to choose representative districts from sixteen districts in Shanghai, which were then divided into low, medium, and high economic levels according to the per capita gross domestic product (GDP) of each district in 2018. Eleven districts were selected proportionally and randomly and represented by 105 community health service institutions. Inclusion criteria were (a) voluntary participation and provision of written consent to participate in the study’s evaluation and assessment and (b) completion of the SSS-CN questionnaire, Patient Health Questionnaire-9 (PHQ-9) and Generalized Anxiety Disorder-7 (GAD-7) questionnaires. Exclusion criteria were (a) patients lacking self-assessed abilities or who refused to participate, (b) patients who have been previously confirmed to have serious mental disorders, mental retardation, or dementia, (c) patients with cancer or central nervous system diseases, (d) patients with any missing data within questionnaire items or with more than one item missing from sociodemographic information. The study was approved by institutional review committees at each center. The study is compliant with the 1975 Declaration of Helsinki guidelines.Elderly populations were defined according to the World Health Organization recommendations . People The SSS-CN questionnaire is self-administered with an abbreviated 20-item measure. Briefly, it is composed of four dimensions: physical disorders, depressive disorders, anxiety disorders and depressive and anxiety disorders. Half of the items ask about physical complaints (one item per body system). We validated the reliability and validity of this instrument in a previous study . The totSupplementary material.All participants were assessed for depressive and anxiety disorders according to the PHQ-9 and GAD-7, which evaluate the frequencies at which certain symptoms had been experienced over the last two weeks, ranging from 0  to 3 (‘nearly every day’). These questionnaires were self-administered. The cutoff values are listed below. For PHQ-9, scores of 5–9 indicate mild depressive disorder, 10–14 indicate moderate depressive disorder, 15–19 indicate moderately severe depressive disorder and ≥ 20 indicate severe depressive disorder . For GADSociodemographic data, including age, gender, level of education and marital status, were also collected. A semi-structured questionnaire concerning clinical characteristics was administered to survey participants’ medical and medication histories.The survey was conducted by trained surveyors. A community pilot study was conducted, and surveyors developed unified explanations of each item based on this pilot study to ensure sufficient understanding. Native Chinese language was used. Surveyors read the items and helped to fill out the questionnaires if the participants were illiterate. Surveyors and supervisors were trained to understand questionnaire contents and study procedures. Data were collected by trained clinical researchers, and data collection was supervised by specialist mental health workers.Data were entered into the EpiData 3.1  database using dual data entry. Over 10% of the database was further audited, including logical consistency checks and extraction of information from participants’ community health records to ensure accuracy and completeness. A unique identifier number was assigned to each participant. Proper categorization and coding of data were performed during data cleaning phases. Database access was password-protected, and access was restricted to key team personnel.Descriptive statistics for each group, such as counts and percentages (%) for non-continuous variables, were reported for all sociodemographic and clinical characteristics. The prevalences of SSD of various severities for different genders and for participants diagnosed with depressive and anxiety disorders were calculated.In order to assess whether SSD, depressive and anxiety disorders are associated with aged groups and whether depressive and anxiety disorders are associated with the severity of SSD, binary logistic regressions were performed, adjusting for gender, educational level , marital status , hypertension (yes/no), diabetes mellitus (yes/no), CV disease (yes/no), endocrine and metabolic disease except DM (yes/no), other diseases (yes/no), surgery (yes/no) and antipsychotic intake (yes/no) in all models.Univariate analyses and binary logistic regression models were used to assess the effects of sociodemographic factors and clinical characteristics on SSD in the non-elderly and elderly. Categorical sociodemographic and clinical variables were analyzed by chi-square test. Binary logistic regressions were performed using SSD (yes/no) as the dependent variable, adjusting for gender, educational level, marital status, and antipsychotic intake. Odds ratios (ORs), 95% confidence interval (95%CI) values, and significance values  were reported in tabular form and forest plots. The number of risk factors for each participant was calculated to assess the association between the number of risk factors and SSD prevalence. Subgroup analysis by gender in the elderly group was also conducted.p-value < 0.05 (two-tailed). Missing data were imputed with a multiple imputation approach. All statistical analyses were performed with SPSS 22 .Fisher’s exact test was used instead of the chi-square test only when one or more expectations for each cell of the 2-by-2 tables was below 5. A significance level was set at a Eleven thousand two hundred community dwellers people between March 2019 and June 2019 were approached, and the rejection rate was less than 1%. Then, all collected questionnaires were checked for completeness and quality by trained surveyors based on inclusion criteria and exclusion criteria. Based on inclusion and exclusion criteria, 9020 subjects between 18 to 100 were included in this prospective multicenter registry, composed of 6814 elderly (age ≥ 60 years) and 2206 non-elderly participants (18–59 years). The baseline characteristics of study participants are presented in Table p < 0.001). Furthermore, we calculated the prevalence of SSD separately for participants with (n = 1865) and without (n = 7155) depressive or anxiety disorders, and the prevalence of SSD in patients with combined depressive or anxiety disorders was 3.7 times higher than would be expected in the general population. In elderly participants with SSD, 96.7% exhibited accompanying depressive disorders, and 95.7% exhibited anxiety  and 11.2% (n = 1014). In the elderly, the prevalence of anxiety disorder was 10.9% (n = 741), compared to 12.4% (n = 273) in the non-elderly. For depressive disorders, these values were 16.8% (n = 1142) and 17.8% (n = 393), respectively. Supplementary Table Sp = 0.001) and were also less likely to have anxiety disorders  . Interestingly, educational level was found to be associated with SSD in the non-elderly, but not in the elderly. With regard to clinical characteristics, cardiac disease and cardiovascular risk factors were significantly associated with SSD in elderly participants (p < 0.05).Table p = 0.002). Similarly, participants over 80 years of age were 1.291 times more likely to have SSD . The risk of SSD was 1.566 times higher in elderly females compared to elderly males .In order to identify risk factors likely to independently predict SSD, the indices in Table n = 338), and additional risk factors (up to four) were associated with SSD rates of 55.6% (n = 1357), 65.1% (n = 1400), 78.8%(n = 857) and 84.5% (n = 355). For non-elderly participants, respective SSD rates by number of patient risk factors were 27.0% (n = 131), 45.7% (n = 584), 60.8% (n = 203), 71.4% (n = 65) and 88.9% (n = 16). An elevated number of risk factors corresponded with a higher likelihood of SSD. The prevalence of SSD in the elderly was 1.554 times higher than that of the non-elderly group when controlling for the number of risk factors . Notably, when the number of risk factors reached four, SSD prevalence in the non-elderly surpassed that observed in the elderly  was far smaller than the number of elderly participants meeting the same criteria (n = 420), a potential source of bias. Secondly, non-elderly participants suffering from many (≥ 4) physical diseases may exhibit relatively higher stress levels.As expected, SSD occurrence was positively correlated with the number of risk factors, and prevalence was higher in the elderly  is of choice , 41. HowTo our knowledge, this is the first study to investigate the clinical characteristics of SSD in elderly and non-elderly with the largest population. The relevant demographic, clinical features, and risk factors of the elderly with SSD are emphasized. We believe that our findings could provide a reference to economically developed regions in China in dealing with SSD in the elderly and prepare for the more severe aging in the future.Our study also has several limitations. First, although our study represents the largest elderly and non-elderly population for which SSD has been studied to date, it is, nevertheless, a cross-sectional study for the elderly. No information was obtained about disorder progression or treatment requirements. Secondly, participants in our study were all enrolled from Shanghai, a city with better medical care, better education , 28 and Our findings demonstrate that, compared to the non-elderly, the elderly have a higher prevalence of SSD and exhibit more severe comorbidities with physical and psychological problems. Older females with several comorbid physical diseases were identified as an extremely vulnerable group; attention should be given to screening these outpatients.Additional file 1.