[Objective] To compare the characteristics of platelet-rich plasma derived exosomes (PRP-Exos) under different activation conditions and their differential effects on the proliferation capacit of human microvascular endothelial cells (HMEC-1) and human skin fibroblasts (BJ). [Methods] Ten healthy volunteers were recruited, and 10 mL of venous blood anticoagulated with EDTA-K2 was collected. PRP-Exos were prepared and extracted by the secondary centrifugation method. Transmission electron microscopy (TEM), nanoparticle tracking analysis (NTA), and Western blotting (WB) detection techniques were used to compare the morphological characteristics, particle size distribution, concentration, and the expression of surface marker proteins of PRP-Exos. Through in vitro cell culture, the differences in morphological changes and proliferation abilities of HMEC-1 and BJ cells induced by PRP-Exos in different activator groups were compared. [Results] 1) TEM observation showed that the morphologies of PRP-Exos in different activator groups were different. 2) NTA analysis showed that compared with the particle size of (109.60±2.71) nm in the normal saline group, the particle sizes of the thrombin group [(104.93±1.55) nm] and the mixed agent group [(103.83±1.58) nm] were relatively smaller, while the particle size of the calcium gluconate group [(113.57±4.70) nm] was larger and its particle size distribution range was wider. There were statistically significant differences among different groups (F=7.019, P<0.05). The concentration of exosomes obtained in the group with the mixture of calcium gluconate and thrombin was the highest [(4.87±0.63)×10⁶ particles/mL, P<0.001]. Both the thrombin group [(2.75±0.13)×10⁶ particles/mL, P < 0.01] and the calcium gluconate group [(3.82±0.06)×10⁶ particles/mL, P<0.001] obtained higher concentrations of exosomes compared with the normal saline group. The concentration in the calcium gluconate group was slightly higher than that in the thrombin group, and there was a significant difference between the two groups (P<0.05). 3) WB analysis showed that CD41, CD81, and TSG101 were expressed on the surface of PRP-Exos, and the protein signal intensities of CD41 and TSG101 in the group with the mixture of thrombin and calcium gluconate were the strongest (P<0.001). 4) The results of in vitro cell culture showed that PRP-Exos in the group with the mixture of thrombin and calcium gluconate could significantly promote the proliferation of HMEC-1 and BJ cells (P<0.05). [Conclusion] The PRP treated with the thrombin and calcium gluconate mixture group yielded the highest concentration of exosomes; under in vitro culture conditions, PRP-Exos from this group significantly promoted the proliferation of HMEC-1 and BJ cells.

Objective To study the traditional Chinese medicine(TCM)constitution characteristics of overweight/obese patients in Shanghai region and to investigate the correlation of TCM constitution with body composition.Methods Relevant data were collected from the patients with complete information of TCM constitution and human body composition analysis who visited the specialized outpatient clinic of acup-moxibustion catgut embedding therapy in the Department of Endocrinology,Shanghai Tenth People's Hospital from August 2020 to December 2022.The patients were divided into a normal body mass group(BMI＜24 kg/m2),an overweight group(24 kg/m2≤BMI＜28 kg/m2)and obesity group(BMI≥28 kg/m2),and then the distribution of TCM constitution types in the three groups of patients were analyzed.After that,the correlation between TCM constitution and body composition were explored with multiple regression analysis.Results(1)A total of 315 patients were included,of which 43 patients had normal body mass,85 patients were overweight and 187 patients were obese.(2)The TCM constitution types in descending order of the composition ratio in the normal body mass group and in the overweight group were spleen deficiency constitution,liver stagnation constitution,damp-heat constitution,yang deficiency constitution,and yin deficiency constitution,in the obese group were spleen deficiency constitution,yang deficiency constitution,damp-heat constitution,liver stagnation constitution,and yin deficiency constitution,and in the overweight/obese group were spleen deficiency constitution,damp-heat constitution,yang deficiency constitution,liver stagnation constitution,and yin deficiency constitution.No statistically significant differences of the distribution of TCM constitution types were shown between normal body bass population and overweight/obese population(P＞0.05).In both normal body mass population and overweight/obese population,the single body constitution type was common,and biased constitution was rare,and there was no statistically significant difference when comparing between the two groups(P＞0.05).(3)The results of multiple regression analysis showed that the basal metabolism of all patients was positively correlated with yang deficiency constitution and was negatively correlated with damp-heat constitution,and the differences were statistically significant(P＜0.01).It is indicated that if the score of yang deficiency constitution rose by one point,the basal metabolism would increase by 0.54 kcal,and if the score of damp-heat constitution decreased by one point,the basal metabolism would decrease by 1.005 kcal.Conclusion In Shanghai region,obesity may be the main indication of the variation of the body constitution.In addition to spleen deficiency constitution,the proportions of yang deficiency constitution,damp-heat constitution and liver stagnation constitution are also higher in obese patients.In terms of the correlation between TCM constitution and body composition,basal metabolism is positively correlated with yang deficiency constitution and is negatively correlated with damp-heat constitution.Therefore,for the patients with yang deficiency constitution and damp-heat constitution,the influence of the basal metabolism level on the development of the disease should be taken into account.

OBJECTIVE: To investigate whether electroacupuncture (EA) at sensitized acupoints could reduce sympathetic-sensory coupling (SSC) and neurogenic inflammatory response by interfering with 5-hydroxytryptamine (5-HT)ergic neural pathways to relieve colitis and somatic referred pain, and explore the underlying mechanisms. METHODS: Rats were treated with 5% dextran sodium sulfate (DSS) solution for 7 days to establish a colitis model. Twelve rats were randomly divided into the control and model groups according to a random number table (n=6). According to the "Research on Rat Acupoint Atlas", sensitized acupoints and non-sensitized acupoints were determined. Rats were randomly divided into the control, model, Zusanli-EA (ST 36), Dachangshu-EA (BL 25), and Xinshu (BL 15) groups (n=6), as well as the control, model, EA, and EA + GR113808 (a 5-HT inhibitor) groups (n=6). The rats in the control group received no treatment. Acupuncture was administered on 2 days after modeling using the stimulation pavameters: 1 mA, 2 Hz, for 30 min, with sparse and dense waves, for 14 consecutive days. GR113808 was injected into the tail vein at 5 mg/kg before EA for 10 min for 7 consecutive days. Mechanical sensitivity was assessed with von Frey filaments. Body weight and disease activity index (DAI) scores of rats were determined. Hematoxylin and eosin staining was performed to observe colon histopathology. SSC was analyzed by immunofluorescence staining. Immunohistochemical staining was performed to detect 5-HT and substance P (SP) expressions. The calcitonin gene-related peptide (CGRP) in skin tissue and tyrosine hydroxylase (TH) protein levels in DRG were detected by Western blot. The levels of hyaluronic acid (HA), bradykinin (BK), prostaglandin I2 (PGI2) in skin tissue, 5-HT, tryptophan hydroxylase 1 (TPH1), serotonin transporters (SERT), 5-HT 3 receptor (5-HT3R), and 5-HT 4 receptor (5-HT4R) in colon tissue were measured by enzyme-linked immunosorbent assay (ELISA). RESULTS: BL 25 and ST 36 acupoints were determined as sensitized acupoints, and BL 15 acupoint was used as a non-sensitized acupoint. EA at sensitized acupoints improved the DAI score, increased mechanical withdrawal thresholds, and alleviated colonic pathological damage of rats. EA at sensitized acupoints reduced SSC structures and decreased TH and CGRP expression levels (P<0.05). Furthermore, EA at sensitized acupoints reduced BK, PGI2, 5-HT, 5-HT3R and TPH1 levels, and increased HA, 5-HT4R and SERT levels in colitis rats (P<0.05). GR113808 treatment diminished the protective effect of EA at sensitized acupoints in colitis rats (P<0.05). CONCLUSION EA at sensitized acupoints alleviated DSS-induced somatic referred pain in colitis rats by interfering with 5-HTergic neural pathway, and reducing SSC inflammatory response.
Rats ; Animals ; Electroacupuncture ; Rats, Sprague-Dawley ; Serotonin ; Acupuncture Points ; Pain, Referred ; Calcitonin Gene-Related Peptide ; Signal Transduction ; Colitis/therapy* ; Indoles ; Sulfonamides

Objective To investigate the effects of Huaier polysaccharides on proliferation,apoptosis,and angiogenesis in A549 cells by regulating the Sonic hedgehog-Gli family zinc finger 1(Shh-Gli1)signaling pathway.Methods A549 cells were divided into control group(normal culture)and low,medium,high dose experimental groups(2,5,and 10 μg·mL-1 Huaier polysaccharides).After 24 h of treatment,cell proliferation level was detected by methyl thiazolyl tetrazolium assay,cell apoptosis level was detected by flow cytometry,cysteine aspartic acid specific protease-3(caspase-3)enzyme activity in cells was detected by colorimetric assay,and the expression levels of vascular endothelial growth factor(VEGF),vascular endothelial growth factor receptor 2(VEGFR2),Shh and Gli1 proteins in cells were detected by Western blot.Results The cell survival rates of control group and low,medium,high dose experimental groups were 100.00％,(73.13±3.86)％,(47.97±6.12)％and(30.73±6.67)％,respectively;cell apoptosis rates were(0.72±0.04)％,(19.21±1.27)％,(29.48±0.92)％and(46.29±2.81)％,respectively;caspase-3 enzyme activity in cells were 1.00,1.33±0.08,1.71±0.09 and 2.28±0.14,respectively;the relative expression levels of VEGF protein in cells were 1.17±0.04,0.95±0.08,0.39±0.04 and 0.11±0.03,respectively;the relative expression levels of VEGFR2 protein in cells were 0.99±0.04,0.80±0.03,0.29±0.03 and 0.24±0.03,respectively;the relative expression levels of Gli1 protein in cells were 1.22±0.04,1.07±0.04,0.60±0.04 and 0.41±0.05,respectively.There were statistical significance of above indicators between control group and low,medium,high dose experimental groups(all P＜0.05).Conclusion Huaier polysaccharides can inhibit proliferation and angiogenesis of A549 cells and promote apoptosis by regulating the Shh-Gli1 signaling pathway.

AIM To establish an HPLC method for the simultaneous content determination of gallic acid,protocatechuic acid,morroniside,loganin,sweroside,paeoniflorin,hypericin,astragalin,salvianolic acid B,salvianolic acid A,epimedin C and icariin in Bushen Huoxue Sanjie Capsules.METHODS The analysis was performed on a 30℃thermostatic Agilent 5 TC-C18 column(250 mm×4.6 mm,5 μm),with the mobile phase comprising of acetonitrile-0.1%phosphoric acid flowing at 1.0 mL/min in a gradient elution manner,and the detection wavelength was set at 240 nm.RESULTS Twelve constituents showed good linear relationships within their own ranges(r≥0.999 8),whose average recoveries were 97.11%-101.14%with the RSDs of 0.60%-2.65%.CONCLUSION This simple,accurate and reproducible method can be used for the quality control of Bushen Huoxue Sanjie Capsules.

Background The compound 6:2 chlorinated polyfluorinated ether sulfonic acids (6:2 Cl-PFESA) has been demonstrated abilities of strong bioaccumulation and placental barrier penetration, and it can also cross the blood-brain barrier. However, the mechanism of its neurodevelopmental toxicity in offspring induced by early-life exposure is still unknown. Objective To explore effects of 6:2 Cl-PFESA on the growth and the α-amino-3-hydroxy-5-methylisoxazole-4-propionic (AMPA) receptor gene expression in the hippocampus of offspring mice by establishing a 6:2 Cl-PFESA exposure animal model. Methods Thirty Kunming pregnant mice were randomly divided into five groups: control group, and 2, 10, 50, and 250 μg·L⁻¹ 6:2 Cl-PFESA exposure groups. The treatment groups were exposed to designed doses of 6:2 Cl-PFESA through drinking water from the first day of gestation until the end of lactation. The pups were weaned on postnatal day (PND) 21, and continued to be exposed to 6:2 Cl-PFESA through drinking water. Birth weight and body length of the offspring were recorded. Offspring mice were anesthetized and sacrificed respectively on PND7, PND21, and PND35, then their hippocampus was peeled from harvested brain tissue. The ultrastructure of hippocampus was observed via transmission electron microscopy; and the expression of AMPA receptors GluR1, GluR2, and GluR3 in the hippocampus was evaluated by real-time reverse transcription polymerase chain reaction. The learning and memory ability of the PND35 mice was measured by Morris water maze test before they were sacrificed. Results The birth weights and the lengths of the pups in the 10, 50, and 250 μg·L⁻¹ 6:2 Cl-PFESA exposure groups were (2.23±0.36), (1.92±0.20), (1.88±0.31) g, and (33.73±0.98), (32.91±1.30), (32.52±2.07) mm, respectively, which were lower than those in the control group, (2.78±0.35) g and (36.46±2.34) mm (P<0.05), respectively. The results of Morris water maze showed that the escape latencies in the orientation navigation experiment on the 4th day in the 250 μg·L⁻¹ 6:2 Cl-PFESA exposure group and on the 5th day in the 10, 50, and 250 μg·L⁻¹ 6:2 Cl-PFESA exposure groups were longer than those in the control group (P<0.05). In the space exploration experiment, the times of crossing platform in the 50 and 250 μg·L⁻¹ 6:2 Cl-PFESA exposure groups were decreased when compared with the control group (P<0.05), and the time of staying in the target quadrant of the 250 μg·L⁻¹ 6:2 Cl-PFESA exposure groups were also decreased (P<0.05). Via transmission electron microscopy, compared with the control group, the postsynaptic density was decreased and the synaptic cleft width was widened on PND35 in the 250 μg·L⁻¹ 6:2 Cl-PFESA exposure group. The mRNA expression levels of GluR1, GluR2, and GluR3 in the hippocampus of pups exposed to 250 μg·L⁻¹ 6:2 Cl-PFESA during different developmental stages were significantly lower than those in the control group (P<0.05). Except for the 2 μg·L⁻¹ 6:2 Cl-PFESA exposure group on PND7, the 6:2 Cl-PFESA exposure inhibited the mRNA expression levels of GluR1, GluR2, and GluR3 in the hippocampus of pups at different developmental stages (P<0.05). Among them, the 6:2 Cl-PFESA exposure during early development resulted in the highest decrease in the expression levels of GluR1 and GluR2 mRNA in the hippocampus of pups on PND7; GluR3 mRNA expression level in the hippocampus of the exposed pups on PND21 showed the maximum inhibitory effect; the expression levels of GluR1, GluR2, and GluR3 mRNA all showed the least decrease in the hippocampus of the exposure groups on PND35. Conclusion Early-life exposure to 6:2 Cl-PFESA may affect the growth and development of offspring mice, alter the hippocampal synaptic structure, and influence the learning and memory abilities, which may be related to their inhibitory effects on the expression levels of AMPA receptor subunits GluR1, GluR2, and GluR3 genes in the hippocampus of offspring mice at various developmental stages.

Objective To study the changes in serum immunoglobulin levels in children with thalassemia who undergo repeated blood transfusions and explore their correlation with delayed hemolytic transfusion reactions(DHTR).Methods Serum samples from children with thalassemia who received blood transfusion treatment from June 2022 to April 2023(ob-servation group)and healthy children who underwent physical examination(control group)in our hospital were collected.The levels of serum immunoglobulins(IgG subtype,IgM,IgA,IgE and IgD)were detected using flow cytometry CBA multi-factor quantitative detection technology,and the differences between the two groups were compared.The children were divided into 4 groups according to different transfusion numbers:≤10 numbers,11-30 numbers,31-50 numbers and＞50 numbers,and the differences between different blood transfusion numbers and serum immunoglobulin levels in each group were compared using one-way analysis of variance(ANOVA).Children with thalassemia with DHTR were in the hemolysis group,and children with thalassemia who did not experience DHTR were in the non-hemolysis group.The changes in serum immunoglobulins(IgG subtypes,IgM,IgA,IgE and IgD)between the two groups were compared to explore the correlation between serum immunoglobulins in thalassemia children with repeated transfusion and DHTR.Results The levels of IgG1,IgG3,IgG4 and IgA in the observation group were significantly higher than those in the control group,with the increase of(2.07±2.12),(0.67±2.03),(0.30±0.37)and(6.04±11.40)mg/mL,respectively,while the level of IgD in observation group was significantly lower than that in the control group,with a decrease of(0.03±0.01)mg/mL,P＜0.05.No significant difference was noticed in IgG2,IgM and IgE between the groups(P＞0.05).IgG1 and IgG4 both significantly increased with the number of blood transfusions.The IgG1 in the 4 groups increased sequentially as(0.30±0.62),(0.41±0.51)and(3.60±3.48)mg/mL,and IgG4 increased sequentially as(0.12±0.13),(0.22±0.07)and(0.21±0.38)mg/mL.IgG2,IgM and IgD showed a significant decrease,with IgG 2,IgM,and IgD in four groups decreased as(0.91±1.50),(0.14±0.10)and(0.05±0.05)mg/mL,respectively,showing significant differences with the number of blood transfusions(P＜0.05).No sig-nificant difference was found in IgG3,IgA and IgE with different number of transfusions(P＞0.05).IgG1,IgG3 and IgG4 in the hemolysis group were significantly higher than those in the non-hemolysis group,with an increase of(4.44±3.41),(0.73±1.26)and(0.52±0.40),respectively(P＜0.05).IgD in the hemolysis group was significantly lower than that in the non-hemolysis group,with a decrease of(0.00±0.06)mg/mL,P＜0.05.No significance was noticed in IgG2,IgM,IgA and IgE between the hemolysis group and the non-hemolysis group(P＞0.05).Conclusion The serum immunoglobulin levels of children with thalassemia who undergo repeated blood transfusions are abnormal.There are differences in correlation between the number of blood transfusions and serum immunoglobulin levels among children with thalassemia who undergo repeated blood transfusions.The relevant serum immunoglobulins for DHTR in children with thalassemia who undergo repeated blood transfusions are IgG1,IgG3 and IgG4.

Objective:To investigate the value of transanal multipoint full-layer puncture biopsy (TMFP) in predicting pathological complete response (pCR) after neoadjuvant radiotherapy and chemotherapy (nCRT) in patients with locally advanced rectal cancer (LARC) and to establish a predictive model for providing clinical guidance regarding the treatment of LARC.Methods:In this multicenter, prospective, cohort study, we collected data on 110 LARC patients from four hospitals between April 2020 and March 2023: Beijing Chaoyang Hospital of Capital Medical University (50 patients), Beijing Friendship Hospital of Capital Medical University (41 patients), Qilu Hospital of Shandong University (16 patients), and Zhongnan Hospital of Wuhan University (three patients). The patients had all received TMFP after completing standard nCRT. The variables studied included (1) clinicopathological characteristics; (2) clinical complete remission (cCR) and efficacy of TMFP in determining pCR after NCRT in LARC patients; and (3) hospital attended, sex, age, clinical T- and N-stages, distance between the lower margin of the tumor and the anal verge, baseline and post-radiotherapy serum carcinoembryonic antigen (CEA) and carbohydrate antigen (CA)19-9 concentrations, chemotherapy regimen, use of immunosuppressants with or without radiotherapy, radiation therapy dosage, interval between surgery and radiotherapy, surgical procedure, clinical T/N stage after radiotherapy, cCR, pathological results of TMFP, puncture method (endoscopic or percutaneous), and number and timing of punctures. Single-factor and multifactorial logistic regression analysis were used to determine the factors affecting pCR after NCRT in LARC patients. A prediction model was constructed based on the results of multivariat analysis and the performance of this model evaluated by analyzing subject work characteristics (ROC), calibration, and clinical decision-making (DCA) curves. pCR was defined as complete absence of tumor cells on microscopic examination of the surgical specimens of rectal cancer (including lymph node dissection) after NCRT, that is, ypT0+N0. cCR was defined according to the Chinese Neoadjuvant Rectal Cancer Waiting Watch Database Study Collaborative Group criteria after treatment, which specify an absence of ulceration and nodules on endoscopy; negative rectal palpation; no tumor signals on rectal MRI T2 and DWI sequences; normal serum CEA concentrations, and no evidence of recurrence on pelvic computed tomography/magnetic resonance imaging.Results:Of the 110 patients, 45 (40.9%) achieved pCR after nCRT, which was combined with immune checkpoint inhibitors in 34 (30.9%). cCR was diagnosed before puncture in 38 (34.5%) patients, 43 (39.1%) of the punctures being endoscopic. There were no complications of puncture such as enterocutaneous fistulae, vaginal injury, prostatic injury, or presacral bleeding . Only one (2.3%) patient had a small amount of blood in the stools, which was relieved by anal pressure. cCR had a sensitivity of 57.8% (26/45) for determining pCR, specificity of 81.5% (53/65), accuracy of 71.8% (79/110), positive predictive value 68.4% (26/38), and negative predictive value of 73.6% (53/72). In contrast, the sensitivity of TMFP pathology in determining pCR was 100% (45/45), specificity 66.2% (43/65), accuracy 80.0% (88/110), positive predictive value 67.2% (45/67), and negative predictive value 100.0% (43/43). In this study, the sensitivity of TMFP for pCR (100.0% vs. 57.8%, χ 2=24.09, P<0.001) was significantly higher than that for cCR. However, the accuracy of pCR did not differ significantly (80.0% vs. 71.8%, χ 2=2.01, P=0.156). Univariate and multivariate logistic regression analyses showed that a ≥4 cm distance between the lower edge of the tumor and the anal verge (OR=7.84, 95%CI: 1.48-41.45, P=0.015), non-cCR (OR=4.81, 95%CI: 1.39-16.69, P=0.013), and pathological diagnosis by TMFP (OR=114.29, the 95%CI: 11.07-1180.28, P<0.001) were risk factors for pCR after NCRT in LARC patients. Additionally, endoscopic puncture (OR=0.02, 95%CI: 0.05-0.77, P=0.020) was a protective factor for pCR after NCRT in LARC patients. The area under the ROC curve of the established prediction model was 0.934 (95%CI: 0.892-0.977), suggesting that the model has good discrimination. The calibration curve was relatively close to the ideal 45° reference line, indicating that the predicted values of the model were in good agreement with the actual values. A decision-making curve showed that the model had a good net clinical benefit. Conclusion:Our predictive model, which incorporates TMFP, has considerable accuracy in predicting pCR after nCRT in patients with locally advanced rectal cancer. This may provide a basis for more precisely selecting individualized therapy.

Objective:To explore the characteristics of SMN1 gene variants and carry out functional verification for two children with Spinal muscular atrophy (SMA). Methods:Two male children with complicated SMA diagnosed at the Children′s Hospital Affiliated to Capital Institute of Pediatrics respectively in July 2021 and April 2022 due to delayed or retrograde motor development were selected as the study subjects. Clinical data of the children were collected. Primary culture of skin fibroblasts was carried out, and peripheral blood samples were collected from both children and their parents. Multiplex ligation-dependent probe amplification, combined long-range PCR and nested PCR, and Sanger sequencing were carried out to detect the copy number and variants of the SMN1 gene. Absolute quantitative real-time PCR, Western blotting and immunofluorescence were used to determine the transcriptional level of the SMN gene, expression of the SMN protein, and the number of functional SMN protein complexes (gems body), respectively. This study was approved by Medical Ethics Committee of the Children′s Hospital Affiliated to Capital Institute of Pediatrics (Ethics No. SHERLLM2021009). Results:Child 1, a 1-year-old boy, was clinically diagnosed with type 1 SMA. Child 2, a 2-and-a-half-year-old boy, was clinically diagnosed with type 3 SMA. Both children were found to harbor a paternally derived SMN1 deletion and a maternally derived SMN1 gene variant, namely c. 824G>T (p.Gly275Val) and c. 884A>T (p.*295Leu). Compared with the normal controls and carriers, the levels of full-length SMN1 transcripts in their peripheral blood and skin fibroblast cell lines were significantly decreased ( P<0.05), and the levels of SMN protein normalized to that of β-actin, and the numbers of gems bodies in the primary fibroblast cells were also significantly lower ( P<0.05). Based on the guidelines from the American College of Medical Genetics and Genomics, both variants were classified as likely pathogenic (PS3+ PM3+ PM5+ PP3; PS3+ PM3+ PM4+ PP3). Following the diagnosis, both children had received nusinersen treatment. Although their motor function was improved, child 1 still died at the age of 2 due to severe pulmonary infection. The walking ability of child 2 was significantly improved, and his prognosis appeared to be good. Conclusion:Two cases of clinically complicated SMA have been confirmed by genetic testing and experimental studies, which has provided a reference for their accurate treatment.

Objective To compare the differences in virulence-related factor aspartate protease,biofilm formation,and gene expression among clinical isolates of Candida parapsilosis(C.parapsilosis).Methods Gene sequencing and microsatellite typing(MT)method were adopted to identify C.parapsilosis isolated from patients with clinical fungal infection.The production of secreted aspartate protease and biofilm formation ability of each strain were de-tected,and the expression of biofilm formation related-genes BCR1,EFG1,and HWP1,as well as aspartate prote-ase virulence genes SAPP1,SAPP2,SAPP3 were compared among the strains.Results A total of 8 clinically iso-lated C.parapsilosis strains were collected,all of which were identified as genotype Ⅰ.Based on microsatellite ty-ping results,8 clinical strains were divided into 4 microsatellite types.G1,G2,and G3 strains isolated from the urine,peripherally inserted central catheters(PICC),and blood of patient A were of different subtypes.J1,J2,J3,J4,and J5 strains were of the same type,and isolated from blood specimens of patient B at different periods.All 8 clinical strains could form biofilm,and their biofilm formation ability was higher than that of the standard strain of C.parapsilosis(ATCC 22019).G1,G3 and J5 strains had strong biofilm formation ability,J1,J2,J3,and J4 strains had moderate biofilm formation ability,and G2 strain had weak biofilm formation ability.All of the eight clinical isolates secreted aspartate protease,and their in vitro expression levels of the enzyme were higher than that of the standard strain(ATCC 22019).G3,G1,and G2 strains showed low,moderate,and high in vitro enzyme expression respectively,with statistical differences(all P＜0.05).Enzyme expressed moderately in J1 and J5 strains,and highly in J2,J3,and J4 strains.Difference between moderate and high expressions was statistically significant(P＜0.05).The expression levels of biofilm formation genes BCR1,EFG1,and HWP1 in various strains isolated from patients A and B increased.In strains isolated from patient A,the expression level of EFG1 gene in G1 strain was higher than that in G2 strain(P＜0.05).There was no statistically significant difference in BCR1,EFG1,and HWP1 gene expression levels among strains isolated from patient B.The expression levels of as-partate protein genes(SAPP1,SAPP2,and SAPP3)in various strains isolated from patients A and B increased.The expression levels of SAPP1 and SAPP2 in strain G1 were higher than those in G2 and G3(both P＜0.05).There was no statistically significant difference in the expression levels of SAPP1,SAPP2,and SAPP3 genes in strains from patient B.Conclusion Clinical isolates of C.parapsilosis have higher biofilm formation and aspartate protease production abilities than standard strain.The expression of virulence factors varies among strains isolated from different specimens,while there is no significant difference in the expression of virulence factors among strains isolated at different periods.Patients may have been infected with different MT types of C.parapsilosis in multiple sites during the same period.