In mass spectrometry-based proteomics experiments,achieving high-throughput and efficientproteolytic digestion is crucial to ensure optimal protein cleavage and enhance the depth of protein identi-fication (including the number of identified proteins and the coverage of protein amino acid sequences) .Trypsin is the most widely used protease in mass spectrometry-based proteomics due to its ability to spe-cifically cleave the carboxyl terminus of arginine and lysine.However,it was found that Trypsin has some missed enzymatic efficiency for the cleavage of lysine residues.Therefore,in actual proteomics sample preparation,a combination of Trypsin and LysC will be used to ensure adequate cleavage of lysine resi-dues.Our study revealed that the commonly employed LysC-Trypsin tandem cleavage method exerts an impact on the enzymatic cleavage of protein samples by Trypsin due to the subsequent cleavage of Trypsin by initially added LysC.Consequently,we adjusted the order of LysC and Trypsin tandem digestion,with Trypsin cleavage being performed first followed by the addition of LysC to target any missed lysine resi-dues.We comprehensively compared and analyzed three distinct sequential digestion methods,namely Trypsin-Trypsin (T-T),LysC-Trypsin (L-T),and Trypsin-LysC (T-L),in terms of their effects on pro-tein sample preparation quality.The results demonstrated that the Trypsin-LysC sequential digestion ap-proach not only minimizes missed protein lysine/arginine cleavage sites without increasing experimental costs,at the same time yielding peptides with a moderate amino acid sequence length.The use of Tryp-sin-LysC digestion enhances the adsorption and separation of peptide samples in RP-HPLC,as well as improves the depth of protein detection and amino acid sequence coverage during tandem mass spectrome-try analysis.This research work offers a novel technical solution and serves as a valuable reference for proteome sample preparation.

OBJECTIVE: To investigate the protective mechanisms of Chinese medicine Shexiang Tongxin Dropping Pills (STDP) on heart failure (HF). METHODS: Isoproterenol (ISO)-induced HF rat model and angiotensin II (Ang II)-induced neonatal rat cardiac fibroblast (CFs) model were used in the present study. HF rats were treated with and without STDP (3 g/kg). RNA-seq was performed to identify differentially expressed genes (DEGs). Cardiac function was evaluated by echocardiography. Hematoxylin and eosin and Masson's stainings were taken to assess cardiac fibrosis. The levels of collagen I (Col I) and collagen III (Col III) were detected by immunohistochemical staining. CCK8 kit and transwell assay were implemented to test the CFs' proliferative and migratory activity, respectively. The protein expressions of α-smooth muscle actin (α-SMA), matrix metalloproteinase-2 (MMP-2), MMP-9, Col I, and Col III were detected by Western blotting. RESULTS: The results of RNA-seq analysis showed that STDP exerted its pharmacological effects on HF via multiple signaling pathways, such as the extracellular matrix (ECM)-receptor interaction, cell cycle, and B cell receptor interaction. Results from in vivo experiments demonstrated that STDP treatment reversed declines in cardiac function, inhibiting myocardial fibrosis, and reversing increases in Col I and Col III expression levels in the hearts of HF rats. Moreover, STDP (6, 9 mg/mL) inhibited the proliferation and migration of CFs exposed to Ang II in vitro (P<0.05). The activation of collagen synthesis and myofibroblast generation were markedly suppressed by STDP, also the synthesis of MMP-2 and MMP-9, as well as ECM components Col I, Col III, and α-SMA were decreased in Ang II-induced neonatal rats' CFs. CONCLUSIONS STDP had anti-fibrotic effects in HF, which might be caused by the modulation of ECM-receptor interaction pathways. Through the management of cardiac fibrosis, STDP may be a compelling candidate for improving prognosis of HF.
Rats ; Animals ; Matrix Metalloproteinase 2/metabolism* ; Matrix Metalloproteinase 9/metabolism* ; RNA-Seq ; Transcriptome/genetics* ; Heart Failure/drug therapy* ; Collagen ; Collagen Type I/metabolism* ; Fibrosis ; Myocardium/pathology*

To investigate the therapeutic effect of Jingfang Granules on carbon tetrachloride(CCl_4)-induced liver fibrosis in mice and its mechanism. Forty-nine 8-week-old male C57 BL/6 J mice were randomly divided into a blank group, a CCl_4 group, a silybin group(positive control, 100 mg·kg~(-1))+CCl_4, a Jingfang high-dose(16 g·kg~(-1)) group, a Jingfang high-dose(16 g·kg~(-1))+CCl_4 group, a Jingfang medium-dose(8 g·kg~(-1))+CCl_4 group, and a Jingfang low-dose(4 g·kg~(-1))+CCl_4 group, with 7 mice in each group. The mice in the blank group and Jingfang high-dose group were intraperitoneally injected olive oil solution, and mice in other groups were intraperitoneally injected with 10% CCl_4 olive oil solution(5 mL·kg~(-1)) to induce liver fibrosis, twice a week with an interval of 3 d, for 8 weeks. At the same time, except for the blank group and CCl_4 group, which were given deionized water, the mice in other groups were given the corresponding dose of drugs by gavage once daily for 8 weeks with the gavage volume of 10 mL·kg~(-1). All mice were fasted and freely drank for 12 h after the last administration, and then the eyeballs were removed for blood collection. The liver and spleen were collected, and the organ index was calculated. The levels of alanine aminotransferase(ALT), aspartate aminotransferase(AST), total bile acid(TBA), and triglyceride(TG) in the serum of mice were detected by an automated analyzer. Tumor necrosis factor-α(TNF-α), interleukin-6(IL-6) and interleukin-1β(IL-1β) levels were detected by enzyme-linked immunosorbent assay(ELISA). Kits were used to detect the contents of superoxide dismutase(SOD), malondialdehyde(MDA), and glutathione(GSH) in the liver tissue. Pathological changes in the liver tissue were observed by hematoxylin-eosin(HE), Masson, and Sirius red staining. Western blot was used to detect protein expressions of transforming growth factor-β(TGF-β), α-smooth muscle actin(α-SMA) and Smad4 in the liver tissue. The results indicated that Jingfang Granules significantly reduced the organ index, levels of ALT, AST, TBA,TG, TNF-α, IL-6, and IL-1β in the serum, and the content of MDA in the liver tissue of mice with CCl_4-induced liver fibrosis. Jingfang Granules also significantly increased the content of SOD and GSH in the liver tissue. Meanwhile, Jingfang Granules down-regulated the protein levels of TGF-β, α-SMA, and Smad4. Furthermore, Jingfang Granules had no significant effect on the liver tissue morphology and the above indexes in the normal mice. In conclusion, Jingfang Granules has obvious therapeutic effect on CCl_4-induced liver fibrosis, and its mechanism may be related to reducing the expression of pro-inflammatory factors, anti-oxidation, and regulating TGF-β/Smad4 signaling pathway.
Mice ; Male ; Animals ; Tumor Necrosis Factor-alpha/metabolism* ; Interleukin-6/metabolism* ; Olive Oil/therapeutic use* ; Carbon Tetrachloride/metabolism* ; Liver Cirrhosis/metabolism* ; Liver ; Superoxide Dismutase/metabolism* ; Transforming Growth Factor beta/metabolism*

To analyze the risk factors for urinary tract infection (UTI) among inpatients. The case data of 1 875 inpatients receiving urinary bacterial culture in Beijing Haidian Hospital from October 2019 to May 2021 were analyzed retrospectively. According to the etiological diagnostic criteria of UTI, they were divided into infection group and non-infection group. The species and distribution of pathogens in the infection group were analyzed, and the case data and laboratory indexes were subjected to univariate analysis. The variables with statistical significance were selected for binary logistic regression to analyze the risk factors of urinary tract infection and establish a prediction model. The receiver operating characteristic (ROC) curve was drawn for each parameter included in the model, and the area under the curve (AUC) was calculated. The diagnostic and predictive efficacy of each parameter alone and their combination for UTI were evaluated. So, a total of 1 162 patients with non-infection group and 713 patients with UTI were detected. Among the cultured pathogens, the constituent ratio of Gram-negative bacteria, Gram-positive bacteria and fungi was 57.2%(408/713), 35.9%(256/713) and 6.9%(49/713) respectively. Multivariate analysis showed that, Age, duration of urinary catheterization>7 d, stroke and orthopedic surgery were the risk factors of UTI among inpatients. The use of antibiotics is a protective factor for urinary tract infections. The prediction model of UTI was established by the risk factors, age, duration of urinary catheterization>7 d, stroke, orthopedic surgery, urinary leukocyte esterase, urinary nitrite and Coefficient of variability of red blood cell volume distribution width (RDW-CV). The AUC of the combination of the eight parameters in diagnosing and predicting UTI was 0.835 (95%CI: 0.816-0.855), with the sensitivity of 70.7% and the specificity of 82.8%. In conclusion,the combination of the eight parameters can better assist in the diagnosis and prediction of UTI, and provide an experimental basis for clinicians to judge UTI.
Humans ; Retrospective Studies ; Inpatients ; Urinary Tract Infections/microbiology* ; Urinalysis ; Stroke

OBJECTIVE: To study the effect of low-dose dopamine adjuvant therapy on inflammatory factors and prognosis in preterm infants with necrotizing enterocolitis (NEC). METHODS: A total of 100 preterm infants with NEC from June 2017 to June 2019 were enrolled and divided into a dopamine treatment group and a conventional treatment group using a random number table, with 50 infants in each group. The infants in the conventional treatment group were given symptomatic treatment, and those in the dopamine treatment group were given low-dose dopamine adjuvant therapy in addition to the conventional treatment. ELISA was used to measure the levels of C-reactive protein (CRP), tumor necrosis factor-α (TNF-α), and interleukin-8 (IL-8). The two groups were compared in terms of time to relief of clinical symptoms, fasting time, treatment outcome, prognosis, and adverse reactions. RESULTS: Both groups had significant reductions in the levels of CRP, TNF-α, and IL-8 after treatment, and the dopamine treatment group had significantly lower levels of these markers than the conventional treatment group after treatment (P<0.05). Compared with the conventional treatment group, the dopamine treatment group had significantly shorter time to defecation improvement, time to relief of abdominal distension and diarrhea, and fasting time (P<0.05), a significantly higher response rate (P<0.05), and a significantly lower surgery rate (P<0.05). There were no significant differences in the mortality rate and incidence of adverse events between the two groups (P>0.05). CONCLUSIONS Low-dose dopamine adjuvant therapy can effectively improve the levels of inflammatory factors and clinical symptoms in preterm infants with NEC and has good safety, and therefore, it holds promise for clinical application.

OBJECTIVE: To study the protective mechanism of Chinese medicine Suxiao Jiuxin Pills (, SXJ) on myocardial ischemia and reperfusion (I/R) injury. METHODS: Mouse myocardial I/R injury model was created by 30-min coronary artery occlusion followed by 24-h reperfusion, the mice were then divided into the sham group (n=7), the I/R group (n=13), the tirofiban group (TIR, positive drug treatment, n=9), and the SXJ group (n=11). Infarct size (IS), risk region (RR), and left ventricle (LV) were analyzed with double staining methods. In addition, H9C2 rat cardiomyocytes were cultured with NaSO to simulate I/R in vitro. The phosphorylation of extracellular regulated protein kinases1/2 (ERK1/2), protein kinase B (AKT), glycogen synthase kinase-3β (GSK3β), and protein expression of GATA4 in nucleus were detected with Western blot assay. RESULTS: The ratio of IS/RR in SXJ and TIR groups were lower than that in I/R group (SXJ, 22.4% ±6.6%; TIR, 20.8%±3.3%; vs. I/R, 35.4%±3.7%, P<0.05, respectively). In vitro experiments showed that SXJ increased the NaSO-enhanced phosphorylation of AKT/GSK3β and nuclear expression of GATA4. CONCLUSION SXJ prevents myocardial I/R injury in mice by activating AKT/GSK3β and GATA4 signaling pathways.

Objective:To explore the effect of task-based rehabilitative training on neural circuit plasticity and forelimb motor function after C₅ spinal cord injury in mice. Methods:A total of 21 healthy C57/BL mice were randomly and equally divided into sham group, model group and training group. The model was established by left C₅ spinal cord crush injury. The lamina was removed without damaging the spinal cord in the sham group. Four weeks after injury, the training group received task-based rehabilitative training for four weeks. The horizontal ladder and rearing tests were used to assess motor function for forelimb before injury, and three days, two weeks, four weeks, six weeks and eight weeks after injury. The axons of the corticospinal tract in all mice were observed six weeks after injury by using biotinylated dextran amin (BDA) anterograde tracing. Eight weeks after injury, motor-evoked potential was applied to measure nerve conduction velocities in forelimb, while the axon sprouting and syntagmatic relation of neuron in the anterior horn of gray matter above lesion were observed by immunofluorescence double-labeling of BDA/neuron-specific nuclei protein (NeuN); the expression of Synapsin in the anterior horn of gray matter at lesion was observed by immunofluorescence double-labeling of NeuN/Synapsin I. Results:Eight weeks after injury, the latency of P1 and N1 was longer in the model group than in the sham group (P < 0.05), and was shorter in the training group than in the model group (P < 0.05). Compared with the sham group, the error rate of left forelimb increased, and the usage rate decreased (P < 0.05) in the model group and the training group; compared with the model group, the error rate of left forelimb decreased six weeks and eight weeks after injury (P < 0.05), and the usage rate increased eight weeks after injury (P < 0.05) in the treatment group. Compared with the model group, more axon sprouting co-localized with neurons in the anterior horn of gray matter above lesion (P < 0.05), and the expression of Synapsin I increased in the training group (P < 0.05). Conclusion:Task-based rehabilitative training could promote the neural circuit plasticity and improve the motor function of forelimb after spinal cord injury in mice.

Objective To compare the differences in the hypertension prevalence among children aged 7-12 in Guangzhou based on different references of hypertension. Methods A total of 7698 children aged 7-12 years old in Guangzhou were recruited by stratified cluster random sampling method. Demographic information such as gender and age was collected by questionnaire. Children’s height, weight, and blood pressure were objectively measured. There were five references for diagnosing children’s hypertension: Mi 2010, Mi 2017, Ma 2017, America 2004, and America 2017 reference. Results Based on the references above, the prevalence of hypertension for children aged 7-12 in Guangzhou reached a very high level. The prevalence of hypertension, high systolic blood pressure (SBP) and high diastolic blood pressure (DBP) based on Mi 2017 reference were all higher than those based on Mi 2010 reference, but both of them were distinctly higher than Ma 2017 reference. Compared with Mi 2010 reference, the agreement of diagnosis of high SBP and high DBP were both higher for Mi 2017 than those for Ma 2017. The agreement of high SBP was higher between Mi 2017 and America 2017 than that between Ma 2017 and America 2017 (Kappa: 0.846 vs. 0.727). Conclusion The prevalence of hypertension in children aged 7-12 in Guangzhou reachs a very high level. The prevalence of hypertension based on Mi 2017 reference is the highest and follows with America 2017 reference, and the agreement between them is excellent. Compared with Mi 2010 or America 2017 reference, the agreement for Ma 2017 is lower than that for Mi 2017 reference, respectively.

Objective To evaluate sedentary lifestyles after school in children aged 7 to 12 year-old living in Guangzhou, and to explore the association between sedentary behaviors after school with cardiometabolic risk factors. Methods Using the method of stratified cluster random sampling, this study recruited 7 to 12 year-old primary students (n=4 294) in Guangzhou. The physical examination and questionnaire were used to collect the sedentary lifestyles after school and cardiometabolic risk factors, analyzing the impact of different aedentary behavoir time after school on cardiometabolic risk factors. Results The average sedentary time after school per day were 194.3 min (boys: 200.3 min; girls: 187.3 min). Inter-quartile ranges of sedentary time after school per day were ≤130.0, 131.0-180.0, 181.0-240.0, and ≥241.0 min/d. Controlling for confounding factors, the odd ratios (OR) of central obesity, overweight/obesity, high TC status, high TG status and high LDL-C status in the highest compared to the lowest quartile of sedentary time after school per day were 1.39 (95%CI:1.08-1.80), 1.44 (95%CI:1.16-1.80), 1.26(95%CI:1.05-1.51), 1.63(95%CI:1.34-1.98), 1.28(95%CI:1.06-1.55), respectively. Conclusions Sedentary lifestyles have a positive relationship with childhood central obesity, overweight/obesity and dyslipidemia in primary school children. Therefore, it is essential to strengthen the intervention to the lifestyles of teenagers and reduce the sedentary behavior time of children and teenagers.