BACKGROUND:In vitro construction of tissue-engineered intestinal models plays an important role in intestinal regeneration and intestinal disease research.The interaction of intestinal nervous system and intestinal epithelial barrier to maintain body homeostasis is a hot topic in the bionic construction of tissue-engineered intestinal tract.OBJECTIVE:To construct a bionic model that can mimic the enteric nervous system in vivo.METHODS:Using fibroin protein with villus structure as scaffold,human induced neural stem cells solidified with collagen were added to intestinal epithelial cells(Caco-2 and HT29-MTX-E12)for 3-day culture to construct a co-culture system of intestinal epithelial cells and nerve cells(co-culture group).Human induced neural stem cells or intestinal epithelial cells cultured alone that were inoculated with fibroin scaffolds were set as controls.Cell morphology was observed by scanning electron microscopy and hematoxylin-eosin staining.Cell activity was detected by Live/Dead cell staining.Human induced neural stem cell differentiation was detected by β-microtubulin immunofluorescence staining.Intestinal epithelial histological properties and barrier function were detected by microvillin,sucrase-isomaltase,tight junction protein 1,E-calmodulin,and mucin-2 immunofluorescence staining.The function of mucus secretion from intestinal epithelial cells was detected by Alcian blue staining.Alkaline phosphatase staining was performed to detect differentiation of intestinal epithelial cells,at the same time,sucrase-isomaltase,tight junction protein 1,and alkaline phosphatase mRNAs were detected by RT-qRCR.RESULTS AND CONCLUSION:The neuralized intestinal mucosal co-culture model with villi structure was successfully constructed,and neural stem cells and intestinal epithelial cells on the fibroin scaffold showed good cellular activities.After neuralization,the activity of alkaline phosphatase and sucrase-isomaltase in intestinal epithelial cells was enhanced,while the expression level of tight junction protein 1 was up-regulated.To conclude,the neuralized bionic intestinal epithelial model is beneficial to the maturation of intestinal mucosal epithelial cells and the formation of barrier function.

BACKGROUND:In vitro construction of tissue-engineered intestinal models plays an important role in intestinal regeneration and intestinal disease research.The interaction of intestinal nervous system and intestinal epithelial barrier to maintain body homeostasis is a hot topic in the bionic construction of tissue-engineered intestinal tract.OBJECTIVE:To construct a bionic model that can mimic the enteric nervous system in vivo.METHODS:Using fibroin protein with villus structure as scaffold,human induced neural stem cells solidified with collagen were added to intestinal epithelial cells(Caco-2 and HT29-MTX-E12)for 3-day culture to construct a co-culture system of intestinal epithelial cells and nerve cells(co-culture group).Human induced neural stem cells or intestinal epithelial cells cultured alone that were inoculated with fibroin scaffolds were set as controls.Cell morphology was observed by scanning electron microscopy and hematoxylin-eosin staining.Cell activity was detected by Live/Dead cell staining.Human induced neural stem cell differentiation was detected by β-microtubulin immunofluorescence staining.Intestinal epithelial histological properties and barrier function were detected by microvillin,sucrase-isomaltase,tight junction protein 1,E-calmodulin,and mucin-2 immunofluorescence staining.The function of mucus secretion from intestinal epithelial cells was detected by Alcian blue staining.Alkaline phosphatase staining was performed to detect differentiation of intestinal epithelial cells,at the same time,sucrase-isomaltase,tight junction protein 1,and alkaline phosphatase mRNAs were detected by RT-qRCR.RESULTS AND CONCLUSION:The neuralized intestinal mucosal co-culture model with villi structure was successfully constructed,and neural stem cells and intestinal epithelial cells on the fibroin scaffold showed good cellular activities.After neuralization,the activity of alkaline phosphatase and sucrase-isomaltase in intestinal epithelial cells was enhanced,while the expression level of tight junction protein 1 was up-regulated.To conclude,the neuralized bionic intestinal epithelial model is beneficial to the maturation of intestinal mucosal epithelial cells and the formation of barrier function.

ObjectiveTo investigate the therapeutic effect and mechanism of Xueshisanjia San against liver fibrosis by regulating PTEN-induced putative kinase （PINK1）/Parkin signaling pathway-mediated mitophagy. MethodsForty specific pathogen free （SPF）-grade male C57BL/6 mice were randomized into the control， model， silibinin （100 mg·kg^-1）， high-dose （15.16 g·kg^-1） Xueshisanjia San， and low-dose （7.58 g·kg^-1） Xueshisanjia San groups. The mouse model of liver fibrosis was constructed by intraperitoneal injection of 20% carbon tetrachloride solution. The treatment lasted for 6 weeks. Blood was collected from the abdominal aorta after intraperitoneal anesthesia， and the liver was separated. Liver pathology was examined by hematoxylin-eosin （HE） staining， Masson staining， and Sirius Red staining. Transmission electron microscopy （TEM） was employed to observe the mitochondrial morphology in the liver tissue. The levels of alanine aminotransferase （ALT）， aspartate aminotransferase （AST）， C-reactive protein （CRP）， total bilirubin （TBil）， transforming growth factor-β₁ （TGF-β₁）， tumor necrosis factor-α （TNF-α）， interleukin-6 （IL-6） in the serum of mice were measured by enzyme-linked immunosorbent assay. Immunohistochemistry， immunofluorescence assay， and Western blot were employed to determine the protein levels of liver fibrosis markers α-smooth muscle actin （α-SMA） and collagen Ⅰ， as well as mitophagy markers microtubule-associated protein 1 light chain 3 （LC3）， p62， Beclin-1， PINK1， Parkin， and translocase of outer mitochondrial membrane 20 （TOM20）. ResultsCompared with the control group， the model group exhibited elevated levels of ALT， AST， CRP， TBil， IL-6， TGF-β₁， and TNF-α in the serum （P<0.05）， pathological changes such destroyed structure of hepatic lobules， disarrangement of hepatic cells， and collagen accumulation， swollen， vacuolated， and fragment mitochondria， down-regulated expression of p62 and TOM20， and up-regulated expression of LC3， Beclin-1， PINK1， and Parkin （P<0.05）. Compared with the model group， all the treatment groups exhibited declined levels of ALT， AST， CRP， TBil， IL-6， TGF-β₁， and TNF-α in the serum （P<0.05）， alleviated pathological damage of liver tissue and mitochondrial damage， up-regulated expression of p62 and TOM20， and down-regulated expression of α-SMA， COL1A1， LC3， Beclin1， PINK1， and Parkin （P<0.05）. ConclusionXueshisanjia San may prevent excessive mitophagy and improve mitochondrial quality by inhibiting PINK1/Parkin signaling pathway， thereby alleviating liver fibrosis.

This article reports a rare case of autoinflammatory disease presenting initially with skin induration and swelling after trauma as the initial manifestation, followed by progressive limb weakness. The patient was a middle-aged female who developed skin induration and swelling after trauma, which gradually progressed to limb weakness, dysarthria and bilateral facial paralysis, accompanied by livedo reticularis of the lower extremities, diffuse skin induration of the limbs, and beaded subcutaneous nodules in the right upper limb. The patient had a susceptibility to infection since childhood and a history of chronic livedo reticularis. Skin pathological examination revealed panniculitis. A comprehensive etiological screening for special infections and autoimmune diseases was completed with an unremarkable results, and whole-exome sequencing showed no abnormal findings. Following a multidisciplinary discussion combined with RNA sequencing results, the patient was diagnosed with an autoinflammatory disease, with a suspected type Ⅰ interferonopathy. Treatment with tofacitinib resulted in gradual improvement of clinical symptoms. This case highlights the importance of detailed medical history collection, systematic physical examination and multidisciplinary collaborative diagnosis and treatment, and underscores the pivotal role of molecular diagnosis in the confirmation of rare diseases. It can provide a reference for the clinical diagnosis and management of similar rare cases.

OBJECTIVE To investigate the acute lung injury effects of pulmonary phospholipids and their phospholipase A2(PLA2)decomposition products-lysophospholipids and fatty acids-on mice.METHODS Mice were randomly assigned to the following groups:① solvent control(PBS)and PLA2;② solvent control and glycerol phospholipid groups:1,2-dioleoyl-sn-glycero-3-phosphoserine(DOPS),1,2-dipalmitoyl-sn-glycero-3-phosphoserine(DPPS),1,2-dioleoyl-sn-glycero-3-phosphoethanol-amine(DOPE),1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine(DPPE),1,2-dipalmitoyl-sn-glycero-3-phosphocholine(DPPC),and 1-stearoyl-2-oleoyl-sn-glycero-3-phosphocholine(SOPC);③ solvent con-trol and fatty acid groups:palmitic acid(PA),oleic acid;④ solvent control and lysophospholipid groups:1-oleoyl-2-hydroxy-sn-glycero-3-phosphoserine(18∶1 LysoPS),1-stearoyl-sn-glycero-3-phosphoserine(18∶0 LysoPS),1-palmitoyl-sn-glycero-3-phosphoserine(16∶0 LysoPS),1-palmitoyl-sn-glycero-3-phos-phoethanolamine(16∶0 LysoPE),1-palmitoyl-sn-glycero-3-phosphocholine(16∶0 LysoPC);⑤ solvent control,PLA2,DPPC,PA,16∶0 LysoPC,16∶0 LysoPS,and 18∶1 LysoPS.Following anesthesia,mice were administered nebulized PBS in the solvent control group,2.1 ug·kg-1 PLA2 in PBS in the PLA2 group and 2.5 mg·kg-1 of the corresponding substance in PBS in other experimental groups.For group①,survival times were recorded and survival curves were plotted.At 1 h post-treatment,lung tissues from groups ①②③④ were collected,photographed to obtain white light images,and subjected to HE staining to assess histopathological changes and pathological scoring.At 2 h post-treatment,pulmonary blood flow in group ⑤ was assessed using laser speckle contrast imaging,arterial blood gas was analyzed with a blood gas analyzer,and lung function was evaluated using whole-body pleth-ysmography.At 6 hours post-treatment,blood cells from group ⑤ were analyzed using an automated hematology analyzer.RESULTS Compared with the solvent control group,severe pathological changes were observed in lung tissues of the PLA2 group,accompanied by extensive inflammatory infiltration and interstitial thickening,with all mice succumbing within 240 min.In mice treated with glyc-erol phospholipids,alveolar structures remained clear,alveolar walls were intact and continuous,and alveolar spaces were translucent,with only occasional minor inflammatory cell infiltration in the septa.No significant pathological alterations were detected in the fatty acid groups.Minor inflammatory cell infiltration was seen in the 16∶0 LysoPE and 16∶0 LysoPC groups.However,such pathological changes as patchy hemorrhage,alveolar interstitial edema,increased alveolar wall thickness,and elevated neutrophil counts were observed in the 18∶1 LysoPS,18∶0 LysoPS,and 16∶0 LysoPS groups.Pathological scores based on HE staining were significantly increased in the 16∶0 LysoPS and 18∶1 LysoPS groups com-pared with the solvent control.The percentage of the lung tissue injury area was also markedly higher in the 16∶0 LysoPS group.A significant decrease in the mean fluorescence intensity of blood flow was observed in the 16∶0 LysoPS group.Arterial partial pressure of oxygen(pO2)was significantly reduced in the PLA2 group,while arterial partial pressure of carbon dioxide(pCO2)was markedly elevated in the 16∶0 LysoPS and 18∶1 LysoPS groups.Lung function tests revealed that the 16∶0 LysoPS group exhibited significant increases in expiratory time,end-expiratory pressure,and enhanced pause,in contrast to significant decreases in tidal volume,expired volume,and minute volume.The 18∶1 LysoPS group also exhibited a significant decline in minute volume.No significant changes in inflammatory cell concentrations were detected in blood,with the exception of neutrophils in the 16∶0 LysoPS group,which showed a significant but physiologically normal increase.CONCLUSION Pulmonary phospholipids and their PLA2-derived fatty acid metabolites do not induce severe lung injury in mice while the lyso-phospholipid metabolites,particularly lysophosphatidylserine,are found to cause significant lung injury.

OBJECTIVE To investigate the impact of vesicular stomatitis virus envelope glycopro-tein-G(VSV-G)modification on the mucosal immune efficacy of antigen-loaded engineered exosome vaccines.METHODS In vitro experiments:Dendritic cells(DCs)were divided into three groups:cell-control(treated with culture medium),receptor binding domain(RBD)(transfected with plasmid RBD),and RBD+VSV-G(co-transfected with plasmids RBD and VSV-G).Expression levels of RBD and VSV-G were assessed using Western blotting,flow cytometry,and immunofluorescence.Exosomes were extracted via ultracentrifugation,whose morphology,size distribution,and marker proteins were analyzed using transmission electron microscopy,nanoparticle tracking analysis,and Western blotting that confirmed the expressions of RBD and VSV-G in the exosomes.In vivo experiments:① Female BALB/c mice were divided into the control group Mock exosomes(Mock-Exo)(derived from the supernatant of cell-control),RBD decorated exosomes(RBD-Exo)(derived from the RBD cell supernatant),and RBD and VSV-G decorated exosomes(RBD+VSV-G-Exo)(derived from RBD+VSV-G cell supernatant).Follow-ing intranasal immunization with the respective vaccines,the nasal retention effects were evaluated using in vivo imaging.Flow cytometry was used to assess the ability to recruit immune cells to the nasal tissue.Serum RBD-specific immunoglobulin G(IgG)and mucosal immunoglobulin A(IgA)(bronchoal-veolar lavage fluid/nasal wash)were quantified at 7 and 21 d post-immunization by enzyme-linked immuno-sorbent assay.Body weight changes were monitored and key serum biochemical parameters along with histopathological damage to major organs were analyzed following immunization.② Female BALB/c mice were divided into the Mock-Exo group(intranasally inoculated with Mock-Exo),RBD+VSV-G-Exo group(intranasally inoculated with RBD+VSV-G-Exo),and RBD+VSV-G-Exo(im)group(intramus-cularly injected with RBD+VSV-G-Exo).RESULTS In vitro experiments:RBD and VSV-G were successfully expressed in cells,with positive rates of RBD+and VSV-G+cells at 64.4%and 31.2%,respectively.The extracted exosomes exhibited regular morphology and qualified purity,with a particle size of approximately 138 nm and successfully loaded RBD and VSV-G proteins.In vivo experiments:Compared to Mock-Exo and RBD-Exo,RBD+VSV-G-Exo prolonged nasal retention time to 96 h and markedly increased the numbers of CD49B+natural killer cells,CD11c+dendritic cells,and F4/80+macrophages in nasal tissues.RBD+VSV-G-Exo induced robust RBD-specific immune responses,with serum IgG titers,BALF IgA titers,and nasal wash IgA titers reaching 1∶5 215,1∶2 560,1∶1 114,respec-tively.In contrast,no RBD-specific IgA antibody titers were detected in the BALF and nasal wash of mice treated with RBD+VSV-G-Exo(im).Mice showed stable body weight gain during 30 d post-immu-nization.Major serum biochemical indices were within normal reference ranges,and no obvious patho-logical changes were observed in major organs or olfactory bulbs 7 d after immunization.CONCLU-SION VSV-G modification extends the retention time of engineered exosome vaccines in nasal tissues,enhance their ability to recruit immune cells,and induce a high-level antigen-specific respiratory mucosal immune response.

Objective:To investigate the effectiveness of multi-channel urodynamic testing(MUT)in preopera-tive evaluation of stress urinary incontinence(SUI),and its impact on postoperative complications and treatment efficacy.Methods:The clinical data of 120 patients were collected.They were hospitalized for SUI and received surgical treatment in the gynecology department of Peking University People's Hospital from May 2020 to June 2022.Among them,60 cases(no-MUT group)were evaluated only by the six-step preoperative evaluation of un-complicated SUI recommended by the American College of Obstetrics and Gynecology without routine MUT,and another 60 cases(MUT gruop)underwent routine MUT and clinical evaluation.Both groups of patients underwent tension-free vaginal tape(TVT-E).The differences in general conditions,intraoperative conditions,postoperative complications,and subjective and objective cure rates between the two groups were compared.Pelvic floor dis-tress inventory-20(PFDI-20)and incontinence quality of life(l-QOL)were used to evaluate the quality of life,and patient-initiated sexual questiornaire-12(PISQ-12)was used to evaluate patients' quality of sex life.Results:①The positive rate of urodynamic SUI diagnosis was 53.3％(32/60)in MUT group.②The subjective cure rates of both groups were 96.7％;the objective cure rate was 98.3％and 96.7％respectively.No significant difference was observed in the postoperative subjective and objective cure rate of patients between two groups(P＞0.05).③After the removal of the urinary catheter,the rate of voiding dysfunction was 5.0％in the no-MUT group and 8.3％in the MUT group.The rate of sling release was 1.7％and 3.3％respectively.The rate of newly developed overactive bladder(OAB)was 1.7％and 3.3％,respectively,with no statistically significant differences(P＞0.05).④No significant difference was observed in the postoperative complications,sexual activity,and quality of life scores between the two groups(P＞0.05).Conclusions:Patients with uncomplicated SUI assessed by the six step preoperative evaluation method can skip routine MUT without increasing surgical complications,and the treatment effect is comparable to that of routine MUT.

Objective:To investigate the effectiveness of multi-channel urodynamic testing(MUT)in preopera-tive evaluation of stress urinary incontinence(SUI),and its impact on postoperative complications and treatment efficacy.Methods:The clinical data of 120 patients were collected.They were hospitalized for SUI and received surgical treatment in the gynecology department of Peking University People's Hospital from May 2020 to June 2022.Among them,60 cases(no-MUT group)were evaluated only by the six-step preoperative evaluation of un-complicated SUI recommended by the American College of Obstetrics and Gynecology without routine MUT,and another 60 cases(MUT gruop)underwent routine MUT and clinical evaluation.Both groups of patients underwent tension-free vaginal tape(TVT-E).The differences in general conditions,intraoperative conditions,postoperative complications,and subjective and objective cure rates between the two groups were compared.Pelvic floor dis-tress inventory-20(PFDI-20)and incontinence quality of life(l-QOL)were used to evaluate the quality of life,and patient-initiated sexual questiornaire-12(PISQ-12)was used to evaluate patients' quality of sex life.Results:①The positive rate of urodynamic SUI diagnosis was 53.3％(32/60)in MUT group.②The subjective cure rates of both groups were 96.7％;the objective cure rate was 98.3％and 96.7％respectively.No significant difference was observed in the postoperative subjective and objective cure rate of patients between two groups(P＞0.05).③After the removal of the urinary catheter,the rate of voiding dysfunction was 5.0％in the no-MUT group and 8.3％in the MUT group.The rate of sling release was 1.7％and 3.3％respectively.The rate of newly developed overactive bladder(OAB)was 1.7％and 3.3％,respectively,with no statistically significant differences(P＞0.05).④No significant difference was observed in the postoperative complications,sexual activity,and quality of life scores between the two groups(P＞0.05).Conclusions:Patients with uncomplicated SUI assessed by the six step preoperative evaluation method can skip routine MUT without increasing surgical complications,and the treatment effect is comparable to that of routine MUT.

Glucagon-like peptide-1 receptor agonists (GLP-1RA) have been widely used in the treatment of type 2 diabetes mellitus (T2DM). However, the acceleration of heart rate caused by GLP-1RA should not be ignored. In the general population and patients with diabetes, increased heart rate has an independent correlation with the incidence and mortality of cardiovascular diseases. In general, the long-acting GLP-1RA seem to exert a greater effect in increasing heart rate, and the effect is dose-dependent and negatively correlated with baseline heart rate. The increase in heart rate caused by GLP-1RA may be related to enhanced sympathetic nervous activity, reflex tachycardia as a response to vasodilation, etc. It is advisable to closely monitor the increased heart rate induced by GLP-1RA in clinical practice, especially in patients with high-risk factors for cardiovascular disease. In case of elevated heart rate, the management begins with immediate discontinuation of the GLP-1RA and symptomatic intervention should be given if necessary.

Objective:To explore the clinical features of nivolumab-induced Stevens-Johnson syndrome/toxic epidermal necrolysis (SJS/TEN).Methods:Relevant databases at home and abroad (as of December 31, 2023) were searched to collect case reports of nivolumab-induced SJS/TEN, and the demographic characteristics, nivolumab application, combination drugs, clinical manifestations, intervention measures, and outcomes were extracted and analyzed descriptively and statistically.Results:A total of 27 case reports were included and 29 patients were enrolled in the study, including 18 males and 11 females. The age ranged from 45 to 86 years, with an average age of 67 years. The primary diseases were mainly melanoma, stomach cancer, and lung cancer. Twelve patients had records of nivolumab administration, and the dosage was within the recommended range in the labels; 13 patients had records of combination drugs, mainly other antineoplastic drugs, hypoglycemic drugs, antihypertensive drugs, lipid-regulating drugs, etc. The time from using nivolumab to the diagnosis of SJS/TEN was 7 d to 3 years, and 20 patients were <8 weeks. The clinical manifestations were mainly diffuse erythema, flaky skin peeling and erosion, mucosal involvement, etc. Sixteen patients had skin biopsy records, all of which met the histopathological characteristics of SJS/TEN. After the diagnosis of SJS/TEN, 17 patients discontinued nivolumab and received symptomatic treatments, of which 15 patients had improved skin symptoms, one patient had worsened skin symptoms, and one patient had no record of skin outcome; 12 patients had no record of whether or not discontinuing nivolumab, of which 8 patients had improved skin symptoms, 2 patients had worsened skin symptoms, one patient had no record of skin outcome, and one had no record of prognosis. One patient rechallenged nivolumab, severe SJS/TEN recurred. Thirteen of 29 patients died. Of them, 1 died due to cardiac arrest, 4 due to worsened skin rash, and 8 due to primary disease progression.Conclusions:SJS/TEN caused by nivolumab mostly occurs within 8 weeks of treatment, and the clinical manifestations were similar to those caused by other drugs. The mortality rate of nivolumab-induced SJS/TEN is high, and skin rash could be improved after withdrawal of nivolumab and symptomatic treatments.