Objective To explore the clinical characteristics of patients with diffuse large B-cell lymphoma (DLBCL) accompanied with KMT2D gene mutation and the impact of its co-mutated genes on prognosis. Methods Clinical data of 155 newly diagnosed DLBCL patients were obtained. The second-generation sequencing method was used to detect 475 hotspot genes, including KMT2D mutation. Patients were divided into the KMT2D mutation group and KMT2D wild-type group based on the presence or absence of KMT2D gene mutation. Clinical characteristics, differences in co-mutated genes, and survival differences between the two groups were compared. Results The frequency of KMT2D mutation was 31%, which is predominantly observed in elderly patients (P=0.07) and less in the double-expressor phenotype (P=0.07). Compared with the KMT2D wild-type group, KMT2D gene mutation was associated with higher co-mutation rates of CDKN2A (OR=2.82, P=0.01) and BCL2 (OR=3.84, P=0.016), while being mutually exclusive with MYC gene mutation (OR=0.11, P=0.013). In univariate survival analysis, no statistically significant difference in overall survival (OS) was found between the KMT2D mutation group and the wild-type group (P=0.54). Further analysis of the prognostic significance of KMT2D with other gene mutations indicated that patients with KMT2D^mutBTG2^mut had poorer OS than those with KMT2D^wt BTG2^mut (P=0.07) and KMT2D^wt BTG2^wt (P=0.05). On the contrary, patients with KMT2D^mut CD79B^mut had better OS than those with KMT2D^mut CD79B^wt (P=0.09), with no prognostic impact observed for other co-mutated genes. Multivariate Cox regression analysis revealed that Ann Arbor stages Ⅲ and Ⅳ (HR=2.751, 95%CI: 1.169-6.472, P=0.02), elevated LDH levels (HR=2.461, 95%CI: 1.396-4.337, P=0.002), Ki-67 index>80% (HR=1.875, 95%CI: 1.066-3.299, P=0.029), and KMT2D^mutBTG2^mut(HR=4.566, 95%CI: 1.348-15.471, P=0.015) were independent risk factors for OS in patients with DLBCL (P<0.05). Conclusion DLBCL patients with KMT2D mutation often have multiple gene mutations, among which patients with a co-mutated BTG2 gene have poor prognosis.

Objective:To investigate the relationship between the polymorphism of endoplasmic reticulum aminopeptidase 1 (ERAP-1) gene and the occurrence of pre-eclampsia (PE).Methods:A case-control study was conducted in Beijing Obstetrics and Gynecology Hospital from October 2018 to October 2021. A total of 51 PE pregnant women with onset gestational age<34 weeks were selected as the PE group, and 48 normal pregnant women during the same period were selected as the control group. Venous blood samples were collected from the pregnant women before delivery and umbilical cord within 5 minutes after delivery. Single nucleotide polymorphisms (SNP) of ERAP-1 gene in the pregnant women and their fetus were detected by next-generation sequencing. Univariate analysis and multivariate logistic regression analysis were used to analyze all the SNP loci and alleles detected in the two groups, and the significant SNP were screened.Results:(1) A total of 13 target SNP loci of maternal ERAP-1 gene were selected by univariate analysis. Among them, the frequency distribution of genotypes at 96096828, 96121524, 96121715, 96122260 and 96122281 showed statistically significant differences between PE group and control group (all P<0.05). Multivariate logistic regression analysis showed that the risk of PE in pregnant women with TC genotype at locus 96121524 was 2.002 times higher than those with TT genotype (95% CI: 0.687-5.831, P=0.020). (2) A total of 4 target SNP loci of ERAP-1 gene in fetal were selected by univariate analysis, and there was no statistical significance in gene polymorphism of the 4 loci between PE group and control group (all P>0.05). Multivariate logistic regression analysis showed that the risk of PE in fetus with genotype AA at locus 96121406 was 0.236 times that of fetus with genotype GG (95% CI: 0.055-1.025, P=0.016). Conclusion:ERAP-1 gene with TC genotype at 96121524 in the mother and GG genotype at 96121406 in the fetus might be related to the incidence of PE.

Objective: To explore the resistance of CD133+ cells in HepG2 cell line to doxorubicin (DOX) and its mechanism. Methods: CD133+ cells were sorted by magnetic beads and CD133+ positive rate was detected by flow cytometry. MTT assay was used to detect the resistance to DOX-induced apoptosis of CD133+ cells. The expression of BCRP transporter mRNA was detected by RT-PCR. The expression of apoptosis-related proteins was detected by Western blotting. Immunofluorescence assay was used to detect the activation and transportation of P65 after DOX treatment. Results: Magnetic beads sorting could efficiently sort the CD133+ cells from HepG2 cells. MTT proliferation assay showed that CD133+ cells had stronger resistance to DOX than CD133- cells (P<0.05). Immunofluorescence showed that the activation rate and content of P65 in CD133+ cells were significantly higher than those in CD133- cells and HepG2 cells (P<0.05). The results of RT-PCR showed that the mRNA content of BCRP in CD133+ cells was significantly increased compared with CD133- cells and HepG2 cells (all P<0.05). Compared with HepG2 and CD133- groups, the expression of Bax and p53 in CD133+ cells was significantly decreased (P<0.05), while the expression of Bcl-2 and Survivin protein in CD133+ cells was significantly increased (P<0.05 or P<0.01). Conclusion: The molecular mechanism of high DOX-resistance of the CD133+ cell subsets in HepG2 cells is the high expression of the survival-related proteins NF-κB, Bcl-2, Survivin and the drug-resistance transporter BCRP, and low expression of apoptosis-promoting proteins p53 and Bax.

Objective: To discuss the application of Mohs microsurgery in nasal and facial basal cell carcinoma (BCC) and analyze the pathological and clinical features. Methods: The clinical data of 127 patients who were diagnosed by pathology as nasal and facial BCC in Qilu Hospital of Shandong University from January 2010 to January 2015 were retrospectively analysed. The value of Mohs microsurgery was discussed and the nasal & facial sites of BCC lesions, clinical and histopathology features were summarized. Results: The proportion of male and female was 1.27︰1 in 127 patients, the ages ranged from 27 to 91 years. The top three inflicted area in nasal and facial was followed by nasal dorsum, nasal root and upper lip.The most frequent clinical type was nodular ulcerative type.The most common pathological type was nodular and pigmented. Routine surgical resection was performed in 62 cases (48.8%) while Mohs micrographic surgery in 38 cases (29.9%). Follow-up duration was 37 months on average. Local recurrence occurred in 5 cases in routine surgical resection group while there was no recurrence in Mohs micrographic surgery group. There was no distant metastasis in all cases. Conclusions There are few specific clinical manifestation in nasal & facial BCC. Surgical treatment is prefered, especially by Mohs micrographic surgery, because it can strictly control the scope of surgical resection and obtain malformation repairment as well as beauty in nasal and facial region.

OBJECTIVETo investigate the effect of Metformin on the proliferation and collagen synthesis of the human keloids fibroblasts as well as the effect on phosphorylation of Akt/FoxO1 signal transduction pathway.

METHODSFibroblasts of keloid were divided into control group treated with medium solution and experimental groups treated with different concentrations of Metformin. 48 h later CCK-8 assay was adopted to evaluate cell survival; Western blot was performed to detect the Akt and FoxO1 phosphorylation; and Hydroxyproline reagent kit was used to detect the collagen synthesis.

RESULTSWith different concentrations (30, 60, 90, 120 mmol/L) of Metformin, the absorbance of cultured keloid fibroblasts detected by CCK8 assay decreased by (13.30 ± 2.04)%, (22.64 ± 4.70)%, (54.00 ± 5.34)% and (63.12 ± 3.48)%. The growth of fibroblasts was suppressed by Metformin in a dose-dependent manner. It showed that the level of phoshpo-akt and phoshpo-foxOl in keloids fibroblasts in experimental groups was lower than that in the control group and the collagen synthesis were also decreased in experimental groups, all in a dose-dependent manner (P < 0.05, P < 0.01).

CONCLUSIONSMetformin can effectively inhibit the proliferation and collagen synthesis of the human keloids fibroblasts in vitro, which may be associated with the suppression of phosphorylation of Akt/FoxO1 signaling pathway

Cell Proliferation ; drug effects ; Collagen ; biosynthesis ; Dose-Response Relationship, Drug ; Fibroblasts ; cytology ; drug effects ; metabolism ; Forkhead Box Protein O1 ; Forkhead Transcription Factors ; metabolism ; Humans ; Keloid ; pathology ; Metformin ; pharmacology ; Phosphorylation ; Proto-Oncogene Proteins c-akt ; metabolism ; Signal Transduction ; drug effects

Objective To evaluate the trueness of 13 routine measurements for serum uric acid and the role of reference method in improving harmonization and trueness among routine measurement systems. Methods The research is related to the reagent evaluation.Usingisotope dilution liquid chromatography tandem mass spectrometry ( ID-LC/MS/MS) method as the comparison method, Wako, Sekisui, DiaSys, Maker,Dirui,Leadman,BSBE,Biosina,Mindray,MedicalSystem,LongMarch,and Kehua 13 kinds of uric acid kits were chosen as the evaluation methods with Hitachi 7170A as the analyzer.serum uric acid in 40 fresh frozen serawere collected from clinical laboratory of Beijing hospital in 2014,coveringboth physiological and pathological status ( 80 -940 μmol/L ) .19 kinds of prepared materials and the 40 fronzen sera were measuredby comparison method and evaluation methods and linear regression analysis was made for the results.The performance of evaluation methods was revealed and recalibration was performed on every evaluation methodby the linear regression equation.The variation of percent bias(%) of the uric acid values in 19 preparation materials was compared.Results All test methods demonstrated good precision ( CV<1.75) and good correlation (R2 >0.998, P<0.01) with the comparison method when measuring uric acid values in 40 fresh frozen sera The meanpercent bias was 0.17% ( -3.06% -7.31%).After recalibration, 4 of 19 samples with no matrix effect values percent bias reduced and met the demands of quality ( <4.8%) induced from biological variation.Conclusion All test methods demonstrated good trueness and their calibration traceability was verified.Recalibration using reference method or standard reference materials contributes to harmonization among methods.

BACKGROUND:Bilateral-sagitta-split-ramus-osteotomy (BSSRO) has become a conventional method to correct facial deformities, and the finite element method is a significant way to study biomechanics of the mandible and temporomandibular joint (TMJ) after BSSRO. OBJECTIVE: To establish a precise and high simulation model of mandible containing TMJ after BSSRO with internal fixation, which is the base to study the biomechanics of the mandible and TMJ after BSSRO. METHODS: Spiral CT scan was used to get the data of DICOM that were input into MIMICS to establish the three-dimensional model of the mandible. The three-dimensional model was wrapped into a single closed shel for mesh generation and conversion in ANSYS. Then, the model was input into the ANSYS software for temporomandibular joint reconstruction and simulation of BSSRO and internal fixation. RESULTS AND CONCLUSION: The three-dimensional finite element model of mandible containing TMJ after BSSRO was established using MIMICS and ANSYS. This model had biological similarity and geometric similarity in comparison with the human tissues. The model could undergo various internal fixations through antedisplacement, retroposition and rotational movement of the distal end. Based on different experimental purposes, the established model can apply a load to al parts to study changes in stress and displacement of different tissues after BSSRO and internal fixation, and it also can be used to study the effect of different fixation materials on the rear stability after internal fixation.

Objective To evaluate the effect of metformin on the proliferation of keratinocytes,and to investigate its possible mechanism.Methods HaCaT human keratinocytes were divided into several groups to remain untreated (control group) or be treated with different concentrations (25,50,75,100 mmol/L) of mefformin for 24 hours (intervention groups).Subsequently,CCK8 assay was conducted to evaluate the proliferation of HaCaT cells,real-time quantitative PCR to measure the mRNA expressions of miR-21-5p and its downstream target gene PDCD4,and Western blot to detect the expression of PDCD4 protein in HaCaT cells.Statistical analysis was done by using one-way analysis of variance for multiple group comparisons and SNK-q test for paired comparisons.Results After 24-hour treatment,the proliferation of HaCaT cells was inhibited by (5.43 ± 3.67)％,(19.61 ± 6.95)％,(45.93 ± 9.56)％ and (61.91 ± 6.93)％ by metformin of 25,50,75 and 100 mmol/L,respectively,with significant differences observed in cell proliferation inhibition rates among these intervention groups (F =246.90,P ＜ 0.05).Cellular proliferative activity was similar between the control cells (0.00 ± 3.00％) and those treated with 25 mmol/L metformin,but significantly higher in the control cells than in the other 3 metformin-treated groups (all P ＜ 0.05),and significantly different between the 4 metformin-treated groups (all P ＜ 0.05).The relative mRNA expression level (2-△△Q) of miR-21-5p was 0.90 ± 0.11,0.33 ± 0.05,0.21 ± 0.07 and 0.14 ± 0.04 (F =36.99,P ＜ 0.01),while that of PDCD4 was 2.11 ± 0.64,7.22 ± 1.13,11.16 ± 1.23 and 19.12 ± 3.16 (F =96.26,P ＜ 0.05),and the expression level of PDCD4 protein was 1.22 ± 0.08,2.09 ± 0.20,2.26 ± 0.1 1 and 2.37 ± 0.07 (F=75.37,P＜ 0.05),respectively,in HaCaT cells treated with metformin of 25,50,75 and 100 mmol/L.Similarly,no significant difference was observed between the control cells and those treated with 25 mmol/L metformin in the expression level of miR-21-5p mRNA,PDCD4 mRNA or protein,but decreased expression of miR-21-5p mRNA and increased expression of PDCD4 mRNA and protein were noted in cells treated with the other 3 concentrations of metformin compared with the control cells (all P＜ 0.05),and significant differences were also found in the expression levels of miR-21-5p mRNA as well as PDCD4 mRNA and protein among the 4 intervention groups (all P ＜ 0.05).Conclusion Metformin can markedly inhibit the proliferation of HaCaT cells in vitro,likely by downregulating miR-21-5p expression and upregulating PDCD4 expression.

Objective:To study the relationship between the expression of heat shock protein 27 (HSP27) in triple negative breast cancer (TNBC) and the clinico-pathological indexes of breast cancer, investigate the correlation between HSP27 and the fatty acid syn-thetase (FAS)/fatty acid synthetase ligand (FASL) of the cell apoptosis system in the Fas/Fasl system, and study the role of HSP27 in the invasion and metastasis of TNBC. Methods:The immunohistochemical S-P method was used to detect the expression of HSP27 and (FAS)/(FASL) in 100 TNBS tissue sampres, 100 non-TNBS tissue sampres, and 50 paraneoplastic tissues. This method was also used to analyze the correlations between the expression of HSP27 and the clinical and pathological indexes of TNBC, as well as be-tween the HSP27 expression and FAS/FASL expression. Results: HSP27 expression was significantly higher in TNBC than in the non-TNBC and paraneoplastic tissues (P<0.05). Statistically significant differences were observed in the FAS/FASL expression in the TNBC, non-TNBC, and paraneoplastic tissues (P<0.05). HSP27 expression had a negative correlation with FAS expression (P<0.05). HSP27 expression was positively correlated with FASL expression (P<0.05). FAS expression had a negative correlation with FASL ex-pression (P<0.05). HSP27 expression in TNBC was not correlated with age, staging, and tumor size (P>0.05), whereas HSP27 expres-sion was correlated with lymph node metastasis, number of nodal metastases, and P53 and Ki67 expression (P<0.05). Conclusion:The overexpression of HSP27 and the expression dysregulation of the FAS/FASL system may play a role in promoting TNBC transfer and invasion, cell proliferation, and poor prognosis.

Objective To study the biocompatibility of carboxymethyl chitosan (CMCS) membrane with melanocytes from healthy human skin,and to investigate the feasibility to transport and carry melanocytes by using CMCS membrane.Methods CMCS membrane was prepared by a casting method combined with a glutaraldehydebased cross-linking method.Melanocytes were isolated from the foreskin of healthy men,and subjected to primary culture and subculture.The third-passage melanocytes were classified into two groups to be cultured on the CMCS membrane (test group) or traditional culture plates (control group).Methyl thiazolyl tetrazolium (MTT) assay was performed to evaluate the proliferative activity of melanocytes,and a sodium hydroxide-based lysis method to determine melanin content.HMB45 staining was conducted,and tyrosinase activity was estimated for melanocytes.Results Inverted microscopy showed that melanocytes were evenly distributed on the CMCS membrane with a normal shape.The melanocytes adherent to the CMCS membrane stained positive for anti-HMB45 monoclonal antibody.The growth curve of the melanocytes on the CMCS membrane,which was obtained from MTT assay,demonstrated that CMCS membrane could support the normal growth of melanocytes.No significant difference was observed between the test group and control group in melanin content (0.083 ± 0.015 vs.0.066 ± 0.008,t =2.38,P ＞ 0.01) or tyrosinase activity (0.234 ± 0.083 vs.0.241 ± 0.061,t =0.23,P ＞ 0.05).Conclusion CMCS membrane can maintain the normal biological activity of melanocytes and have good biocompatibility with skin melanocytes.