The anti-inflammatory effect of simplified Zhiqin Decoction was observed by using lipopolysaccharide (LPS)-induced inflammation mouse model. The main chemical constituents and the main mechanism of action of simplified Zhiqin Decoction were predicted by network pharmacology. Animal experiments verified the anti-inflammatory mechanism of simplified Zhiqin Decoction (this experiment was approved by the Animal Experiment Ethics Committee of Southwest University, approval number: IACUC-20210825-02). Simplifying Zhiqin Decoction has a significant anti-inflammatory effect on inflammatory mice, can significantly improve the overall macro shape of mice, reduce body temperature, water intake, increase the number of autonomous activities; alleviate liver, lung, spleen, thymus inflammation and pathological damage; decrease tumor necrosis factor-α (TNF-α), interleukin (IL)-1β and IL-6, nitric oxide (NO), prostaglandin E₂ (PGE₂) content in serum and urine. The contents of serum immune factors IgG, IgA and IgM were increased. Network pharmacology predicted 66 potential anti-inflammatory active components of simplified Zhiqin Decoction involving 132 inflammatory targets, and the key anti-inflammatory signaling pathway involved PI3K/AKT, TNF, JAK-STAT, etc. Animal experiments show that simplified Zhiqin Decoction can significantly reduce the expression of JAK2/STAT-PI3K/AKT-NF-κB-TNF signaling pathway related proteins in lung tissue of inflammatory mice. The results of this study showed that simplified Zhiqin Decoction had significant anti-inflammatory effects, and its main anti-inflammatory components were quercetin, β-sitosterol, kaempferol, wogonin, stigmasterol, luteolin, neobaicalein, etc. The main mechanism of its anti-inflammatory action is that it significantly inhibits the expression of JAK2/STAT-PI3K/AKT-NF-κB-TNF signaling pathway protein.

Objective To investigate the incidence rate,clinical characteristics,and prognosis of neonatal stroke in Shenzhen,China.Methods Led by Shenzhen Children's Hospital,the Shenzhen Neonatal Data Collaboration Network organized 21 institutions to collect 36 cases of neonatal stroke from January 2020 to December 2022.The incidence,clinical characteristics,treatment,and prognosis of neonatal stroke in Shenzhen were analyzed.Results The incidence rate of neonatal stroke in 21 hospitals from 2020 to 2022 was 1/15 137,1/6 060,and 1/7 704,respectively.Ischemic stroke accounted for 75％(27/36);boys accounted for 64％(23/36).Among the 36 neonates,31(86％)had disease onset within 3 days after birth,and 19(53％)had convulsion as the initial presentation.Cerebral MRI showed that 22 neonates(61％)had left cerebral infarction and 13(36％)had basal ganglia infarction.Magnetic resonance angiography was performed for 12 neonates,among whom 9(75％)had involvement of the middle cerebral artery.Electroencephalography was performed for 29 neonates,with sharp waves in 21 neonates(72％)and seizures in 10 neonates(34％).Symptomatic/supportive treatment varied across different hospitals.Neonatal Behavioral Neurological Assessment was performed for 12 neonates(33％,12/36),with a mean score of(32±4)points.The prognosis of 27 neonates was followed up to around 12 months of age,with 44％(12/27)of the neonates having a good prognosis.Conclusions Ischemic stroke is the main type of neonatal stroke,often with convulsions as the initial presentation,involvement of the middle cerebral artery,sharp waves on electroencephalography,and a relatively low neurodevelopment score.Symptomatic/supportive treatment is the main treatment method,and some neonates tend to have a poor prognosis.

Objective To explore the efficiency of artificial intelligence and radiomics-assisted X-ray in diagnosis of lumbar osteoporotic vertebral compression fractures(OVCF).Methods The clinical data of 455 patients diagnosed as lumbar OVCF by MRI in our hospital were selected.The patients were divided into the training group(n=364)and the validation group(n=91),X-ray films were extracted,the image delineation,feature extraction and data analysis were carried out,and the artificial intelligence radiomics deep learning was applied to establish a diagnostic model for OVCF.After verifying the effectiveness of the model by receiver operating characteristic(ROC)curve,area under the curve(AUC),calibration curve,and decision curve analysis(DCA),the efficiencies of manual reading,model reading,and model-assisted manual reading of X-ray in the early diagnosis of OVCF were compared.Results The ROC curve,AUC and calibration curve proved that the model had good discrimination and calibration,and excellent diagnostic performance.DCA demonstrated that the model had a higher clinical net benefit.The diagnostic efficiency of the manual reading group:the accuracy rate was 0.89,the recall rate was 0.62.The diagnostic efficiency of the model reading group:the accuracy rate was 0.93,the recall rate was 0.86,the model diagnosis showed good predictive performance,which was significantly better than the manual reading group.The diagnostic efficiency of the model-assisted manual reading group:the accuracy rate was 0.92,the recall rate was 0.72,and the recall rate of the model-assisted manual reading group was higher than that of the manual reading group,but lower than that of the model reading group,indicating the superiority of the model diagnosis.Conclusion The diagnostic model established based on artificial intelligence and radiomics in this study has reached an ideal level of efficacy,with better diagnostic efficacy compared with manual reading,and can be used to assist X-ray in the early diagnosis of OVCF.

Objective To develop a clinical prediction model for predicting risk factors for prolonged hospital stay after anterior cervical discectomy and fusion(ACDF).Methods The clinical data of 914 patients underwent ACDF treatment for cervical spondylotic myelopathy(CSM)were retrospectively analyzed.According to the screening criteria,800 eligible patients were eventually included,and the patients were divided into the development cohort(n=560)and the validation cohort(n=240).LASSO regression was used to screen variables,and multivariate Logistic regression analysis was used to establish a prediction model.The prediction model was evaluated from three aspects:differentiation,calibration and clinical effectiveness.The performance of the model was evaluated by area under the curve(AUC)and Hosmer-Lemeshow test.Decision curve analysis(DCA)was used to evaluate the clinical effectiveness of the model.Results In this study,the five factors that were significantly associated with prolonged hospital stay were male,abnormal BMI,mild-to-moderate anemia,stage of surgery(morning,afternoon,evening),and alcohol consumption history.The AUC of the development cohort was 0.778(95%CI:0.740 to 0.816),with a cutoff value of 0.337,and that of the validation cohort was 0.748(95%CI:0.687 to 0.809),with a cutoff value of 0.169,indicating that the prediction model had good differentiation.At the same time,the Hosmer-Lemeshow test showed that the model had a good calibration degree,and the DCA proved that it was effective in clinical application.Conclusion The prediction model established in this study has excellent comprehensive performance,which can better predict the risk of prolonged hospital stay,and can guide clinical intervention as soon as possible,so as to minimize the postoperative hospital stay and reduce the cost of hospitalization.

Objective To explore the risk factors for surgical site infection(SSI)after transforaminal lumbar interbody fusion(TLIF)for the treatment of lumbar degenerative diseases.Methods A total of 1 000 patients who underwent TLIF for lumbar degenerative diseases in our hospital were included and divided into the infection group(n=23)and the non-infection group(n=977)according to whether the surgical incision was infected.General data,surgical and laboratory indicators of patients were collected,and potential risk factors of SSI were screened by univariate analysis and multivariate regression analysis,a nomogram model was established,and its predictive efficiency was validated by the receive operating characteristic(ROC)curve.Results The incidence of SSI in patients after TLIF was 2.3％.The results of univariate analysis showed that age,operative time,intraoperative blood loss,preoperative C-reactive protein(CRP),smoking,and diabetes mellitus were the significant risk factors for the occurrence of SSI.Multivariate regression analysis showed that older age,longer operation time,more intraoperative blood loss,smoking and diabetes mellitus were the independent risk factors for postoperative SSI.ROC curve showed that the nomogram model established in this study has good predictive efficiency.Conclusion Older age,longer operation time,more intraoperative blood loss,smoking,and diabetes mellitus were independent risk factors for postoperative SSI.For patients with these high risk factors,corresponding intervention measures should be taken before operation to reduce the incidence of SSI.

Aim To explore the improvement effect of Bazi Bushen capsule on thymic degeneration in SAMP6 mice and the possible mechanism.Methods Twenty 12 week old male SAMP6 mice were randomly divided into the model group(SAMP6)and the Bazi Busheng capsule treatment group(SAMP6+BZBS).Ten SAMR1 mice were assigned to a homologous control group(SAMR1).The SAMP6+BZBS group was oral-ly administered Bazi Bushen capsule suspension(2.8 g·kg-1)daily,while the other two groups were orally administered an equal amount of distilled water.After nine weeks of administration,the morphology of the thymus in each group was observed and the thymus in-dex was calculated;HE staining was used to observe the structural changes of thymus tissue;SA-β-gal stai-ning was used to detect thymic aging;flow cytometry was used to detect the proportion of thymic CD3+T cells in each group;Western blot was used to detect the levels of p16,Bax,Bcl-2,and cleaved caspase-3 proteins in thymus;immunofluorescence was applied to detect the proportion of cortical thymic epithelial cells in each group;ELISA was employed to detect IL-7 lev-els in thymus.Results Compared with the SAMP6 group,the thymic index of the SAMP6+BZBS group significantly increased(P＜0.05);the disordered thy-mic structure was significantly improved;the positive proportion of SA-β-gal staining significantly decreased(P＜0.01);the proportion of CD3+T cells apparently increased(P＜0.05);the level of p16 protein signifi-cantly decreased(P＜0.05);the level of Bcl-2 pro-tein significantly increased(P＜0.05),while the lev-el of cleaved caspase-3 protein markedly decreased(P＜0.05);the proportion of cortical thymic epithelial cells evidently increased;the level of IL-7 significantly increased(P＜0.01).Conclusions Bazi Bushen capsule can delay thymic degeneration,inhibit cell ap-optosis in thymus and promote thymic cell development in SAMP6 mice,which may be related to increasing the proportion of cortical thymic epithelial cells and promoting IL-7 secretion.

AIM To study the chemical constituents from the leaves of Citrus reticulata Blanco and their anti-inflammatory activities.METHODS The 85%ethanol extract from the leaves of C.reticulata was isolated and purified by silica gel,D101 macroporous resin,MCI,ODS and Sephadex LH-20,then the structures of obtained compounds were identified by physicochemical properties and spectral data.The Griess method was used to determine their inhibitory activities on lipopolysaccharide-induced NO production in macrophages RAW 264.7 cells.The mice foot swelling inflammation model induced by carrageenan was established,and the levels of IL-1β,TNF-α were detected.RESULTS Twelve compounds were isolated and identified as nobiletin(1),tangeretin(2),5-demethylinoblitin(3),5,4'-dihydroxy-6,7,8,3'-tetramethoxy flavone(4),5-hydroxy-7,8,3',4'-trimethoxyflavanone(5),3,5,6,7,8,3',4'-heptamethoxyflavanone(6),hesperetin(7),5-hydroxy-6,7,3',4'-tetramethoxyphenone(8),β-balsam alcohol(9),stigmaster-5-en-3β-alcohol(10),p-hydroxybenzaldehyde(11),vanillin(12).Compounds 1,4,6,7,10 and 12 had strong inhibitory activites on NO release in LPS-induced RAW 264.7 cells,and the IC50 values were(25.21±2.10),(37.77±0.50),(38.19±1.58),(21.89±1.73),(43.81±1.18),(47.98±2.55),(41.23±1.11),(43.80±1.43)μmol/mL,respectively.Compounds 2-3 reduced IL-1β and TNF-α levels(P＜0.05,P＜0.01).CONCLUSION Compounds 6-7,9 are isolated from this plant for the first time.Compounds 1-4,8 exhibit strong in vitro anti-inflammatory activities,and compounds 2-3 exhibit significant in vivo anti-inflammatory activities.

A comprehensive analytical method based on ultra-fast liquid chromatography coupled with triple quadrupole/linear ion trap tandem mass spectrometry(UFLC-QTRAP-MS/MS) was established for simultaneous determination of the content of 45 bioactive constituents including flavonoids, alkaloids, amino acids, phenolic acids, and nucleosides in Epimedium brevicornum. The multiple bioactive constituents in leaves, petioles, stems and rhizomes of E. brevicornum were analyzed. The gradient elution was performed at 30 ℃ in an XBridge~® C_(18) column(4.6 mm×100 mm, 3.5 μm) with 0.4% formic acid aqueous solution-acetonitrile as the mobile phase at a flow rate of 0.8 mL·min~(-1). Single factor experiment and response surface methodology were employed to optimize the extraction conditions. Multivariate statistical analyses including systematic cluster analysis(SCA), principal component analysis(PCA), partial least squares discriminant analysis(PLS-DA), and one-way analysis of variance(One-way ANOVA) were carried out to classify the samples from different parts and identify different constituents. Grey relation analysis(GRA) and entropy weight-TOPSIS analysis were performed to build a multi-index comprehensive evaluation model for different parts of E. brevicornum. The results showed that there was a good relationship between the mass concentrations of 45 constituents and the corresponding peak areas, with the correlation coefficients(r) not less than 0.999 0. The precision, repeatability, and stability of the established method were good for all the target constituents in this study, with the relative standard deviations(RSDs) less than 5.0%(0.62%-4.9%) and the average recovery of 94.51%-105.7%. The above results indicated that the bioactive constituents varied in different parts of E. brevicornum, and the overall quality followed the trend of leaves > petioles > rhizomes > stems. This study verified the rationality of the Chinese Pharmacopoeia(2020 edition) stipulating that the medicinal part of E. brevicornum is the leaf. Moreover, our study indicated that the rhizome had the potential for medicinal development. The established method was accurate and reliable, which can be used to comprehensive evaluate and control the quality of E. brevicornum. This study provides data reference for clarifying the medicinal parts and rationally utilizing the resources of E. brevicornum.
Chromatography, High Pressure Liquid ; Epimedium ; Tandem Mass Spectrometry ; Chromatography, Liquid ; Multivariate Analysis

OBJECTIVE: To establish an intestinal organoid model that mimic acute graft versus host disease (aGVHD) caused intestinal injuries by using aGVHD murine model serum and organoid culture system, and explore the changes of aGVHD intestine in vitro by advantage of organoid technology. METHODS: 20-22 g female C57BL/6 mice and 20-22 g female BALB/c mice were used as donors and recipients for bone marrow transplantation, respectively. Within 4-6 h after receiving a lethal dose (8.0 Gy) of γ ray total body irradiation, a total of 0.25 ml of murine derived bone marrow cells (1×10⁷/mice, n=20) and spleen nucleated cells (5×10⁶/mice, n=20) was infused to establish a mouse model of aGVHD (n=20). The aGVHD mice were anesthetized at the 7th day after transplantation, and the veinal blood was harvested by removing the eyeballs, and the serum was collected by centrifugation. The small intestinal crypts of healthy C57BL/6 mice were harvested and cultivated in 3D culture system that maintaining the growth and proliferation of intestinal stem cells in vitro. In our experiment, 5%, 10%, 20% proportions of aGVHD serum were respectively added into the organoid culture system for 3 days. The formation of small intestinal organoids were observed under an inverted microscope and the morphological characteristics of intestinal organoids in each groups were analyzed. For further evaluation, the aGVHD intestinal organoids were harvested and their pathological changes were observed. Combined with HE staining, intestinal organ morphology evaluation was performed. Combined with Alcian Blue staining, the secretion function of aGVHD intestinal organoids was observed. The distribution and changes of Lgr5⁺ and Clu⁺ intestinal stem cells in intestinal organoids were analyzed under the conditions of 5%, 10% and 20% serum concentrations by immunohistochemical stainings. RESULTS: The results of HE staining showed that the integrity of intestinal organoids in the 5% concentration serum group was better than that in the 10% and 20% groups. The 5% concentration serum group showed the highest number of organoids, the highest germination rate and the lowest pathological score among experimental groups, while the 20% group exhibited severe morphological destruction and almost no germination was observed, and the pathological score was the highest among all groups(t=3.668, 4.334,5.309,P<0.05). The results of Alican blue staining showed that the secretion function of intestinal organoids in serum culture of aGVHD in the 20% group was weaker than that of the 5% group and 10% of the organoids, and there was almost no goblet cells, and mucus was stainned in the 20% aGVHD serum group. The immunohistochemical results showed that the number of Lgr5⁺ cells of intestinal organoids in the 5% group was more than that of the intestinal organoids in the 10% aGVHD serum group and 20% aGVHD serum group. Almost no Clu⁺ cells were observed in the 5% group. The Lgr5⁺ cells in the 20% group were seriously injuried and can not be observed. The proportion of Clu⁺ cells in the 20% group significantly increased. CONCLUSION The concentration of aGVHD serum in the culture system can affect the number and secretion function of intestinal organoids as well as the number of intestinal stem cells in organoids. The higher the serum concentration, the greater the risk of organoid injury, which reveal the characteristics of the formation and functional change of aGVHD intestinal organoids, and provide a novel tool for the study of intestinal injury in aGVHD.
Mice ; Female ; Animals ; Mice, Inbred C57BL ; Bone Marrow Transplantation ; Graft vs Host Disease ; Stem Cells ; Organoids

Objective: To investigate the effects of human umbilical cord mesenchymal stem cells (hUCMSCs) combined with autologous Meek microskin transplantation on patients with extensive burns. Methods: The prospective self-controlled study was conducted. From May 2019 to June 2022, 16 patients with extensive burns admitted to the 990^th Hospital of PLA Joint Logistics Support Force met the inclusion criteria, while 3 patients were excluded according to the exclusion criteria, and 13 patients were finally selected, including 10 males and 3 females, aged 24-61 (42±13) years. A total of 20 trial areas (40 wounds, with area of 10 cm×10 cm in each wound) were selected. Two adjacent wounds in each trial area were divided into hUCMSC+gel group applied with hyaluronic acid gel containing hUCMSCs and gel only group applied with hyaluronic acid gel only according to the random number table, with 20 wounds in each group. Afterwards the wounds in two groups were transplanted with autologous Meek microskin grafts with an extension ratio of 1∶6. In 2, 3, and 4 weeks post operation, the wound healing was observed, the wound healing rate was calculated, and the wound healing time was recorded. The specimen of wound secretion was collected for microorganism culture if there was purulent secretion on the wound post operation. In 3, 6, and 12 months post operation, the scar hyperplasia in wound was assessed using the Vancouver scar scale (VSS). In 3 months post operation, the wound tissue was collected for hematoxylin-eosin (HE) staining to observe the morphological changes and for immunohistochemical staining to observe the positive expressions of Ki67 and vimentin and to count the number of positive cells. Data were statistically analyzed with paired samples t test and Bonferronni correction. Results: In 2, 3, and 4 weeks post operation, the wound healing rates in hUCMSC+gel group were (80±11)%, (84±12)%, and (92±9)%, respectively, which were significantly higher than (67±18)%, (74±21)%, and (84±16)% in gel only group (with t values of 4.01, 3.52, and 3.66, respectively, P<0.05). The wound healing time in hUCMSC+gel group was (31±11) d, which was significantly shorter than (36±13) d in gel only group (t=-3.68, P<0.05). The microbiological culture of the postoperative wound secretion specimens from the adjacent wounds in 2 groups was identical, with negative results in 4 trial areas and positive results in 16 trial areas. In 3, 6, and 12 months post operation, the VSS scores of wounds in gel only group were 7.8±1.9, 6.7±2.1, and 5.4±1.6, which were significantly higher than 6.8±1.8, 5.6±1.6, and 4.0±1.4 in hUCMSC+gel group, respectively (with t values of -4.79, -4.37, and -5.47, respectively, P<0.05). In 3 months post operation, HE staining showed an increase in epidermal layer thickness and epidermal crest in wound in hUCMSC+gel group compared with those in gel only group, and immunohistochemical staining showed a significant increase in the number of Ki67 positive cells in wound in hUCMSC+gel group compared with those in gel only group (t=4.39, P<0.05), with no statistically significant difference in the number of vimentin positive cells in wound between the 2 groups (P>0.05). Conclusions: The application of hyaluronic acid gel containing hUCMSCs to the wound is simple to perform and is therefore a preferable route. Topical application of hUCMSCs can promote healing of the autologous Meek microskin grafted area in patients with extensive burns, shorten wound healing time, and alleviate scar hyperplasia. The above effects may be related to the increased epidermal thickness and epidermal crest, and active cell proliferation.
Female ; Humans ; Male ; Burns/surgery* ; Cicatrix ; Eosine Yellowish-(YS) ; Hyaluronic Acid/therapeutic use* ; Hyperplasia ; Ki-67 Antigen ; Prospective Studies ; Umbilical Cord ; Vimentin ; Young Adult ; Adult ; Middle Aged