Objective:To explore the cytogenetic characteristics of hypospadias in children by karyotype analysis.Methods:From June 2008 to May 2018, 45 children with hypospadias in Tianjin Children's Hospital had cytogenetic abnormalities. Their median age was 10 months(range 3 hours to 5 years old). Of the 45 cases, 20 were proximal hypospadias, 1 was middle hypospadias. All 24 cases had varying degrees of genitourinary malformations. Among them, 15 cases had unilateral or bilateral cryptorchidism, 5 cases had scrotal division, 3 cases had penile scrotal transposition, 3 cases had small penis, 3 cases had indirect inguinal hernia, 1 case had repeated urethra, 1 case had hydrocele and 1 case had concealed penis. To the other systemic malformations, there was 1 with cleft lip and palate and 1 with congenital heart disease. G-banding karyotype analysis of peripheral blood lymphocytes was performed in all 45 cases.Results:Among the 45 cases of hypospadias with abnormal karyotypes, with an abnormal rate of 14.0%, 28 cases (62.22%) had sex chromosome abnormalities, including (47, XXY), (46, XX/47, XXY), (45, X0/47, XYY), etc. Sexual inversion occurred in 8 cases (17.78%), all of which were 46, XX. There were 4 autosomal abnormalities (8.89%), including (46, XY, 9p+ ), (46, XY, 10p+ ) and (46, XY, 1q+ ). Chromosome polymorphism was found in 4 cases (8.89%), including [46, XY, inv(9)] and [46, XY, 16qh+ ], and the equilibrium translocation of 1 case (2.22%) was [45, XY, -21, -22, + t(21; 22)]. Among the 45 cases, 8 sex reversal children with (46, XX) chromosome karyotype were all proximal hypospadias.Conclusions:Children with hypospadias may be associated with chromosomal karyotype abnormalities, including sex chromosomal abnormalities, autosomal abnormalities, chromosome polymorphism and balanced translocation. Among them, sex chromosome abnormality was the most common and balanced translocation was the least.

Objective:The purpose of this study is to evaluate the frequency of MDSCs in peripheral blood of hepatocellular carcinoma patients and to investigate the clinical significance of change of MDSCs in the peripheral blood and provide new ways for e-valuating immune state and prognosis of hepatocellular carcinoma patients.Methods: Blood samples were obtained from 62 patients with HCC and 20 healthy donors.The phenotype of CD3,CD4,CD33,HLA-DR and Th1,Th2 immune subsets in peripheral blood of each group were observed by FCM methods.Results:There were statically different frequencies in the peripheral blood between hepato-cellular carcinoma and healthy control group,which the proportion of total CD3+T lymphocytes and CD3+CD4+T cells were lower and the proportion of CD33+HLA-DR-MDSCs was higher in hepatocellular carcinoma patients.( P<0.05 ).The increase of percentage of MDSCs was greater in patients at Stage C and D than in patients at stage A and B.Conclusion:The Th1/Th2 ratio in the PBMC were of imbalance and MDSCs was significantly increased in peripheral blood of hepatocellular carcinoma patients.The increase of MDSCs was significantly correlated with clinical stage.CD33+HLA-DR-MDSCs may play an important role in prediction in prognosis and tumor immune status of hepatocellular carcinoma.

AIM: To investigate the effect of enhanced green fluorescence protein (EGFP) gene transfection on the cell cycle distribution of primary cultured human chondrocytes in order to establish a tracking method of cultured human nasoseptal chondrocytes. METHODS: pEGFP-N1 plasmid was amplified in E.coli, and purified by high purity kit. Primary cultured human chondrocytes,which were initially obtained from the nasoseptal cartilage, were cultured in vitro and transferred with pEGFP-N1 by means of electroporation with Amaxa nucleofector device. Transfering process and transient expression were evaluated by laser scanning confocal microscope (LSCM), the transfer efficiency and the cell cycle distribution were evaluated by flow cytometry. RESULTS: There was significant expression of EGFP at 24 h after transferring. The transfection efficiency of pEGFP-N1 into primary cultured human chondrocytes reached 35 37% at 48 h. It didn't affect the process of cell adherance and had no effect on the cell cycle distribution. CONCLUSION: Primary cultured human chondrocytes, which were transfected with pEGFP, are alive in vitro, and the transferring process doesn't affect the cell cycle distribution. These results suggest that pEGFP-N1 is an ideal transient expression vector for primary cultured human chondrocytes and it might be a well tracer in construction tissue engineered cartilage.

Objective To observe the preventive effect of Hirudoid (MPS) ointment on psoriasislike dermatitis in mice and explore its mechanism.Metbods 21 Balb/C mice were randomly divided into three groups:blank control group,positive control group (imiquimod group) and treatment group (imiquimod + Hirudoid group).The blank control group was treated with imiquimod matrix.The positive control group was treated with 5％ imiquimod cream to induce psoriatic dermatitis in mice's back.The treatment group was treated with 5％ imiquimod cream in the morning and Hirudoid cream in the afternoon.The changes of skin erythema,scales,infiltration severity and skin histopathology in the experimental area of mice back and serum levels of interleukin (IL)-17A and protein expression of tumor necrosis factor-α (TNF-α)in the three groups of mice were observed.Results The clinical and pathological findings of skin lesions in the positive control group showed that 5％ imiquimod cream successfully induced psoriasis-like dermatitis in mice.The appearance and pathological feature of skin lesions in the blank control group were almost close to normal skin.The degree of erythema,infiltration and scales in the back of the treatment group was significantly less than that in the positive control group,and the degree of microscopic inflammation in the treatment group was also lighter than that in the positive control group;The serum levels of IL-17A and TNF-α in the treatment group and the blank control group were significantly lower than that of the positive control group,with statistically significant difference (P ＜ 0.05).Conclusions Hirudoid can significantly reduce imiquimod-induced psoriasis-like dermatitis by inhibiting the expression of IL-17A and TNF-α.

Objective : To study the epidemiological characteristics of coronavirus disease 2019 （COVID-19） in Zhoushan, so as to provide reference for improving the prevention and control capability of COVID-19. Methods : All the confirmed cases of COVID-19 in Zhoushan, diagnosed according to China’s “COVID-19 diagnosis and treatment plan （fifth version） ” and reported from January 19 to February 17, 2020, were extracted from the infectious disease surveillance system. Data of general information, clinical characteristics, laboratory Results, transmission and detection routes were collected and analyzed. Results : By February 17, Ten confirmed cases of COVID-19 in Zhoushan had been reported, including 7 males and 3 females. They were all adults, with an average age of （50.90±15.00） years. Five cases were discharged. The incubation period ranged from 2 to 12 days, with a median of 5 days. The first symptoms were fever （8 cases）, cough （7 cases） and muscle pain （2 cases）. Chest computed tomographic （CT） scans showed ground glass opacities in the lungs of all the cases. Six cases had Wuhan related exposure. SARS-CoV-2 was tested positive in the sputum specimens or throat swabs of all the cases and the faeces of 4 cases. The last suspicious exposure of 5 cases occurred before, and that of another 5 cases occurred after the onset of clinical symptoms in the confirmed maternal cases. Three cases were found in active consultation after clinical symptoms, and another 7 cases were found in the monitoring of close contacts. Conclusions The 10 confirmed cases of COVID-19 reported in Zhoushan are all adults, and most have Wuhan related exposure. SARS-CoV-2 can be detected in the fecal samples, but the existence of fecal-oral transmission needs more research. SARS-CoV-2 has strong infectivity, and may also have infectivity before the onset of symptoms （at the end of incubation period）. Isolation and early detection of close contacts are conducive for early case-finding.

Objective To investigate the value of peripheral blood diamine oxidase(DAO)in evaluating the severity of ulcerative colitis(UC).Methods A total of 76 UC patients admitted to Daxing Hospital,Capital Medical University from June 2020 to June 2022 were included in UC group,and were divided into severe ulcerative colitis(SUC)group and non-SUC group according to the severity of the disease.A total of 76 healthy subjects were included as control group.The general data and serum DAO of all subjects were collected for statistical analysis.Results The serum DAO level in UC group was significantly higher than that in control group(P<0.05).The serum DAO level in SUC group was significantly higher than that in non-SUC group(P<0.05).Receiver operating characteristic curve analysis showed that DAO predicted area under the curve(AUC)of UC was 0.751(95%CI:0.672-0.829,P<0.001),and DAO predicted AUC of SUC was 0.866(95%CI:0.807-0.925,P<0.001).Conclusion Serum DAO is an effective test index for evaluating the severity of UC,which is worthy of clinical promotion.

Objective To construct a native promoter-regulated Aspergillus fumigatus strain containing red fluorescent protein-labeled calmodulin (CaM-RFP),and to observe the dynamic distribution of calmodulin during the growth of Aspergillus fumigatus.Methods Bilateral flanking sequences of Aspergillus fumigatus calmodulin gene were designed,and plasmids containing the two flanking sequences and mRFP-Aspergillus fumigatus pyrG gene (mRFP-AfpyrG) were amplified separately.The final linear PCR product for transformation was generated from the above three PCR products by fusion PCR.Then,the above linear fragment was transferred into the Aspergillus fumigatus strain by protoplast transformation,so as to construct the CaM-RFP Aspergillus fumigatus strain.The monoclonal colony was picked from the screening medium and subjected to culture.Then,the stablest fluorescent monoxenic strain of Aspergillus fumigatus was selected,and the transformant was verified by PCR.The recombinant strain and wild-type stain were cultured on solid nutrient media separately,and the morphology of these strains was observed by fluorescence microscopy at different time points.Additionally,the above 2 strains were cultured in liquid media separately,and XTT assay was performed to evaluate the growth activity of strains.Microscopy was also conducted to dynamically observe the CaM-RFP Aspergillusfumigatus strain,and analyze the spatial and temporal distribution of calmodulin during the growth and development of Aspergillus fumigatus.Results The fluorescent phenotype and PCR identification results both indicated the successful construction of the CaM-RFP Aspergillus fumigatus strain.The growth activity at 24 hours did not differ between the recombinant strain and wild-type stain (A490:0.689 ± 0.081 vs.0.678 ± 0.054,t =1.32,P ＞0.05),so did the morphology.During the polarized growth of Aspergillus fumigatus,calmodulin was always at the top of the hyphae,germination site of the hyphal branch and the top of new branches.Conclusion Calmodulin may be involved in the regulation of spore germination and polar hyphal growth of Aspergillus fumigatus.

Severe fever with Thrombocytopenia Syndrome (SFTS) is an emerging hemorrhagic fever disease in the rural areas of east-central China,which is caused by SFTSV-a newly discovered bunyavirus.SFTSV is most likely transmitted by tick bites but can also be transmitted within human beings.The onset of SFTS is sudden and with rapid progress,with main clinical manifestations as fever,thrombocytopenia,leucopenia and gastrointestinal and hepatorenal dysfunctions.Some patients may die from multiple organ failure,and the case fatality rate is approximately 10％.In this paper,we use the method of literature review to summarize the recent research progress of SFTS which includes the epidemic distribution characteristics,medium of transmission,host animals,transmission routes and susceptibility in the general population.

Objective To determine the expression of interleukin-6 (IL-6) in cystic lesions of patients with acne vulgaris,and to evaluate the in vitro effect of Propionibacterium acnes (P.acnes) on the production of IL-6 and activation of p38 mitogen-activated protein kinase (p38MAPK) in the human acute monocytic leukemia cell line THP-1.Methods Real-time fluorescence-based quantitative PCR was performed to determine the mRNA expression of IL-6 in cystic lesions of 6 patients with acne vulgaris,as well as in skin tissues of 6 healthy persons.Some cultured THP-1 cells were divided into 5 groups to be treated with 2 × 106 CFU/ml,2 × 107 CFU/ml and 2 × 108 CFU/ml heat-killed P.acnes suspensions (P.acnes groups),100 μμtg/L lipopolysaccharide (LPS group) and RPMI 1640 medium (control group) respectively.After 1-,3-and 6-hour treatment,real-time fluorescence-based quantitative PCR was conducted to determine the mRNA expression of IL-6 in the above groups.Enzyme-linked immunosorbent assay (ELISA) was performed to detect the level of IL-6 in the culture supernatant of cells in the 2 × 108-CFU/ml P.acnes group,LPS group and control group at 24 hours after the treatment.Western blot analysis was conducted to determine the protein expression of p38MAPK and phosphorylated p38MAPK in the 2 × 108-CFU/ml P.acnes group after 15-,30-and 60-minute treatment,as well as in the LPS group after 30-minute treatment and in the control group.Some other THP-1 cells were divided into 3 groups:2 × 108-CFU/ml P.acnes group treated with 2 × 108 CFU/ml P.acnes suspensions,SB203580 (an inhibitor of p38MAPK) group treated with 20 μmol/L SB203580 for 30 minutes followed by the treatment with 2 × 108 CFU/ml P.acnes suspensions,and control group treated with RPMI 1640 medium alone.After 6-hour treatment,the mRNA expression of IL-6 in the above 3 groups was measured by real-time fluorescencebased quantitative PCR.Results The mRNA expression of IL-6 was significantly higher in the cystic lesions of acne vulgaris than in the normal skin tissues (3.680:±:0.790 vs.1.155 ± 0.250,t =3.047,P ＜0.05).Two-way analysis of variance showed that there were significant difference in the mRNA expression of IL-6 among the 2 × 106-CFU/ml,2 × 107-CFU/ml and 2 × 108-CFU/ml p.acnes groups,LPS group and control group (F =532.3,P ＜ 0.001,v =4),and the mRNA expression of IL-6 significantly differed among different time points (F =526.6,P ＜ 0.001,v =2).There were also significant differences in the IL-6 level in the culture supernatant of cells among the 2 × 108-CFU/ml p.acnes group ([1 618.22 ± 32.23] ng/L),LPS group ([3 212.06 ± 353.00] ng/L) and control group ([147.10 ± 0.53] ng/L;v =2,F =102.35,P ＜0.01).After 15-,30-and 60-minute treatment with 2 × 108 CFU/ml P.acnes suspensions,the protein expression of phosphorylated p38MAPK obviously increased.The mRNA expression of IL-6 in THP-1 cells was significantly lower in the SB203580 group than in the 2 × 108-CFU/ml p.acnes group (t =15.91,P =0.004).Conclusions The mRNA expression of IL-6 evidently increases in the cystic lesions of patients with acne vulgaris.P.acnes can activate the signaling molecule p38MAPK in THP-1 cells,and promote the production of IL-6 by THP-1 cells.

Objective To evaluate the effect of Aspergillus fumigatus on the expression of tumor necrosis factor-α (TNF-oα) and activation of intracellular signaling molecule p38 mitogen-activated protein kinase (p38MAPK) in a human acute monocytic leukemia cell line THP-1.Methods Cultured THP-1 cells (2 x 105/ml) were divided into 4 groups to be treated with Aspergillus fumigatus suspensions at concentrations of 106 and 107 colony-forming units (CFU)/ml (106-and 107-CFU/ml Aspergillusfumigatus groups),100 mg/L β-glucan (a positive stimulus,β-glucan group),culture medium (blank control group) respectively for 1,3 and 6 hours.Real-time fluorescence-based quantitative PCR (qPCR) was conducted to determine the mRNA expression of TNF-α in the THP-1 cells in the above groups.Some other THP-1 cells were treated with 107 CFU/ml Aspergillusfumigatus suspensions (107-CFU/ml Aspergillusfumigatus group),β-glucan (β-glucan group) and culture medium (blank control group) separately for 24 hours,and enzymelinked immunosorbent assay (ELISA) was performed to detect the level of TNF-α in the culture supernatant of THP-1 cells.Western blot analysis was conducted to detect the levels of p38MAPK and phosphorylated p38MAPK in THP-1 cells after 15-,30-and 60-minute treatment with 107 CFU/ml Aspergillusfumigatus suspensions.After 2-hour incubation with the p38MAPK inhibitor SB203580 (20 μmol/L),some THP-1 cells were additionally treated with 107 CFU/ml Aspergillus fumigatus suspensions,β-glucan and culture medium separately for 6 hours,and those without SB203580 treatment served as the control group.Then,qPCR was performed to measure the mRNA expression of TNF-α in the THP-1 cells in the above groups.Results The mRNA expression of TNF-α significantly differed among the 106-and 107-CFU/ml Aspergillus fumigatus groups,β-glucan group and blank control group (F =110.983,P ＜ 0.001),and significantly increased over time (F =701.680,P ＜ 0.001).After 24-hour treatment with 107 CFU/ml Aspergillus fumigatus suspensions,the TNF-α level(6 236.30 ± 437.12 ng/L)significantly increased compared with the blank control group (132.10 ± 0.61 ng/L,P ＜ 0.01).Thirty minutes after the treatment with 107 CFU/ml Aspergillusfumigatus suspensions,the phosphorylated p38MAPK level significantly increased,but started to decrease at 60 minutes.The mRNA expression of TNF-α was significantly lower in the SB203580-treated Aspergillusfumigatus groups (3.83 ± 0.62) than in the SB203580-untreated Aspergillus fumigatus groups (187.23 ± 21.62).Conclusion After the treatment with Aspergillus fumigatus,human THP-1 cells can activate the signal molecule p38MAPK and secrete TNF-α,suggesting that monocytes may participate in the innate immune response to Aspergillusfumigatus infection.