Identification of heat shock protein hsp70 family genes from Rana amurensis and its expression profiles upon infection / 生物工程学报

Heat shock proteins (HSPs) widely exist in all organisms, the structures of which are usually extraordinarily conservative. They are also well-known stress proteins that are involved in response to physical, chemical and biological stresses. HSP70 is an important member of the HSPs family. In order to study the roles of amphibians HSP70 during infection, the cDNA sequence of Rana amurensis hsp70 family genes were cloned by homologous cloning method. The sequence characteristics, three-dimensional structure and genetic relationship of Ra-hsp70s were analyzed by bioinformatics methods. The expression profiles under bacterial infection were also analyzed by real-time quantitative PCR (qRT-PCR). Expression and localization of HSP70 protein were tested by immunohistochemical techniques. The results showed that three conservative tag sequences of HSP70 family, HSPA5, HSPA8 and HSPA13, were found in HSP70. Phylogenetic tree analysis indicated four members are distributed in four different branches, and members with the same subcellular localization motif are distributed in the same branch. The relative expression levels of the mRNA of four members were all significantly upregulated (P < 0.01) upon infection, but the time for up-regulating the expression levels were diverse in different tissues. The immunohistochemical analysis showed that HSP70 was expressed to different degrees in the cytoplasm of liver, kidney, skin and stomach tissue. The four members of Ra-hsp70 family have ability to respond bacterial infection to varying degrees. Therefore, it was proposed that they are involved in biological processes against pathogen and play different biological functions. The study provides a theoretical basis for functional studies of HSP70 gene in amphibians.

Puerarin alleviates lipopolysaccharide-induced acute kidney injury in mice by modulating the SIRT1/NF-κB pathway / 南方医科大学学报

OBJECTIVE@#To investigate the role of the SIRT1/NF-κB pathway in mediating the effect of puerarin against lipopolysaccharide (LPS)-induced acute kidney injury (AKI).@*METHODS@#Fifteen BALB/C mice were randomized into control group, LPS group and puerarin treatment group, and in the latter two groups, the mice were given an intraperitoneal injection of LPS (5 mg/kg), followed by daily injection of normal saline for 3 days or injection of puerarin (25 mg/kg) given 1 h later and then on a daily basis for 3 days. On day 5 after modeling, the kidney tissues were taken for histological observation and detection of cell apoptosis. The renal function indexes including urea nitrogen (BUN), serum creatinine (Scr) and kidney injury molecule 1 (KIM-1) and the levels of tumor necrosis factor (TNF-α) and interleukin 1β (IL-1β) were measured, and the expressions of SIRT1 and NF-κB-p65(acetyl K310) in the renal tissues were detected.@*RESULTS@#Intraperitoneal injection of LPS caused obvious glomerular capillary dilatation, hyperemia, renal interstitial edema, and renal tubular epithelial cell swelling and deformation in the mice. The mouse models of LPS-induced AKI also showed significantly increased renal tubular injury score and renal cell apoptosis (P < 0.01) with increased serum levels of BUN, Scr, KIM-1, TNF-α and IL-1β (P < 0.01), enhanced renal expressions of TNF-α, IL-1β and NF-κB p65(acetyl K310) (P < 0.01) and lowered renal expression of SIRT1 (P < 0.05). Treatment with puerarin effectively alleviated LPS-induced renal interstitial edema and renal tubular epithelial cell shedding, lowered renal tubular injury score (P < 0.01) and renal cell apoptosis rate (P < 0.01), and decreased serum levels of BUN, Scr, KIM, TNF-α and IL-1β (P < 0.01). Puerarin treatment significantly reduced TNF-α, IL-1β and NF-κB p65 (acetyl K310) expression in the renal tissue (P < 0.05) and increased SIRT1 expression by 17% (P < 0.05) in the mouse models.@*CONCLUSION@#Puerarin can effectively alleviate LPS-induced AKI in mice possibly by modulating the SIRT1/NF-κB signaling pathway.

Study on the Biological Function of Abemecilib in Inhibiting the Proliferation, Invasion and Migration of Small Cell Lung Cancer with High c-Myc Expression / 中国肺癌杂志

BACKGROUND@#Small cell lung cancer (SCLC) with high c-Myc expression is prone to relapse and metastasis, leading to extremely low survival rate. Cyclin-dependent kinases 4 and 6 (CDK4/6) inhibitor Abemaciclib plays a key role in the treatment of tumors, but the effects and mechanisms on SCLC remain unclear. This study was to analyze the effect and molecular mechanism of Abemaciclib in inhibiting proliferation, migration and invasion of SCLC with high c-Myc expression, with a view to expanding a new direction for reducing the recurrence and metastasis.@*METHODS@#Proteins interacting with CDK4/6 were predicted using the STRING database. The expressions of CDK4/6 and c-Myc in 31 cases of SCLC cancer tissues and paired adjacent normal tissues were analyzed by immunohistochemistry. The effects of Abemaciclib on the proliferation, invasion and migration of SCLC were detected by CCK-8, colony formation assay, Transwell and migration assay. Western blot was used to detect the expressions of CDK4/6 and related transcription factors. Flow cytometry was used to analyze the effects of Abemaciclib on the cell cycle and checkpoint of SCLC.@*RESULTS@#The expression of CDK4/6 was associated with c-Myc by STRING protein interaction network. c-Myc can directly modalize achaete-scute complex homolog 1 (ASCL1), neuronal differentiation 1 (NEUROD1) and Yes-associated protein 1 (YAP1). Moreover, CDK4 and c-Myc regulate the expression of programmed cell death ligand 1 (PD-L1). Immunohistochemistry showed that the expressions of CDK4/6 and c-Myc in cancer tissues were higher than those in adjacent tissues(P<0.0001). CCK-8, colony formation assay, Transwell and migration assay verified that Abemaciclib could effectively inhibit the proliferation, invasion and migration of SBC-2 and H446OE(P<0.0001). Western blot analysis further showed that Abemaciclib not only inhibited CDK4 (P<0.05) and CDK6 (P<0.05), but also affected c-Myc (P<0.05), ASCL1 (P<0.05), NEUROD1 (P<0.05) and YAP1 (P<0.05), which are related to SCLC invasion and metastasis. Flow cytometry showed that Abemaciclib not only inhibited the cell cycle progression of SCLC cells (P<0.0001), but also significantly increased PD-L1 expression on SBC-2 (P<0.01) and H446OE (P<0.001).@*CONCLUSIONS@#Abemaciclib significantly inhibits the proliferation, invasion, migration and cell cycle progression of SCLC by inhibiting the expressions of CDK4/6, c-Myc, ASCL1, YAP1 and NEUROD1. Abemaciclib can also increase the expression of PD-L1 in SCLC.

Research Progress on the Mechanism and Clinical Data of Cereblon in Reversing the Resistance of Lung Cancer to PD-1 Antibody by T cells / 中国肺癌杂志

Programmed cell death receptor 1 (PD-1) is a membrance-spanning protein mostly expressed in the T cell, and combines with programmed cell death ligand 1 (PD-L1) in the targeting cell. When binding to the ligand on tumor cells, PD-1 as an immunosuppressive molecule, can inhibit the immune function of T cells, thus tumor immune escape. For example, depletion of peripheral effector T cell and accelerate the transformation of effector T cells into regulator T cells. To solve this problem, PD-1 antibody is used to bind to PD-1 on T cells to inhibit the interaction between PD-1 on the T cells and PD-L1 on the tumor cells so that it can restore the function of T cells to kill tumor cell. PD-1 antibodies, such as Nivolumab and Pembrolizumb, are approved as a first-line treatment for advanced non-small cell lung cell cancer. However, due to the interaction of tumor cells, T cells and cytokines, some patients developed drug resistance which reduces the efficacy of immunotherapy. Hence, how to overcome resistance has become a urgent problem. Cereblon (CRBN), a substrate receptor of the DDB1-cullin-RING E3 ubiquitin ligase complex and the only known molecular receptor of immunoregulatory drugs, has been found to reverse PD-1 antibody resistance by binding to CRBN regulatory agents (CMS), exert T cell immune function by regulating proliferation, activation and metabolism of T cell. In this paper, the mechanism of down-regulation of T cells leading to resistance of PD-1 antibody in lung cancer, the mechanism of CRBN regulating T cells, and research progress of CRBN regulator in the treatment of lung cancer were reviewed.
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Clinical observation of Dahuang Zhechong pill combined with tenofovir disoproxil fumarate in treating liver fibrosis of chronic hepatitis B / 中国医师进修杂志

Objective:To investigate the efficacy of Dahuang Zhechong pill combined with tenofovir disoproxil fumarate in treating liver fibrosis of chronic hepatitis B and its effect of intestinal flora.Methods:One hundred and twenty chronic hepatitis B cases with liver fibrosis treating in Rugao Hospital Affiliated to Nantong University from March 2018 to November 2019 were randomly divided into the control group and the observation group with 60 cases in each group according to the random number table. The control group was orally given with tenofovir disoproxil fumarate (1 tablet/time and quaque die).The observation group was treated with Dahuang Zhechong pill(3 g/time and bis in die) at basis of control group. The course was for 48 weeks for two groups. Liver function, liver fibrosis, hepatitis B virus e antigen(HBeAg) negative conversion rate, hepatitis b virus (HBV)-DNA negative conversion rate, the efficacy and intestinal flora changes were compared between the two groups before and after treatment. The adverse reactions were recorded in two groups.Results:After treatment, the levels of aspartic transaminase(AST), alanine aminotransferase (ALT), gamma-glutamyltransferase(GGT), total bilirubin (TBIL) in observation group were lower than those in control group [(41.62 ± 5.93) U/L vs. (50.41 ± 7.12) U/L, (41.99 ± 5.76) U/L vs. (52.17 ± 7.09) U/L, (34.46 ± 5.31) U/L vs. (49.41 ± 6.23) U/L, (20.35 ± 3.04) μmol/L vs. (25.60 ± 3.95) μmol/L], and the differences were statistically significant ( P<0.05). After treatment, the levels of hyaluronic acid (HA), procollagen Ⅲ (PCⅢ), type Ⅳ collagen (Ⅳ-C), Laminin(LN) in observation group were lower than those in control group [(167.45 ± 18.75) μg/L vs. (209.44 ± 22.96) μg/L, (103.44 ± 12.75) μg/L vs. (140.08 ± 16.33) μg/L, (81.41 ± 10.06) μg/L vs. (126.36 ± 14.94) μg/L, (108.41 ± 12.72) μg/L vs. (169.41 ± 19.22) μg/L], and the differences were statistically significant ( P<0.05). The total effective rate in the observation group was higher than that in the control group [91.67%(55/60) vs. 76.27%(45/59)], and the difference was statistically significant ( P<0.05). The negative conversion ratio of HBeAg, HBV-DNA in the observation group were higher than those in the control group [28.33%(17/60) vs. 11.86%(7/59), 83.33%(50/60) vs. 66.10%(39/59)], and the differences were statistically significant ( P<0.05). Compared with the control group, the levels of bifidobacteria and lactobacillus in the observation group were higher and the levels of escherichia coli and enterococcus in observation group were lower, and the improvement of intestinal flora in the observation group were better than those in the control group ( P<0.01). Conclusions:Dahuang Zhechong pill combined with tenofovir disoproxil fumarate in treating liver fibrosis of chronic hepatitis B can improve the liver function and liver fibrosis, and can improve the serological conversion rate of HBeAg, inhibit HBV DNA replication, increase the therapeutic effect, and is safety. The effect of Dahuang Zhechong pill on intestinal flora may be related to the curative effect.

Ovarian thecoma: Ultrasonic findings and causes of missed and misdiagnosis / 中国医学影像技术

Objective: To observe the ultrasonic findings of ovarian thecoma (OT), and to analyze causes of missed and misdiagnosis. Methods: Data of 17 OT patients diagnosed by pathology were retrospectively analyzed, and the shape, border, internal echo, blood signal, posterior echo and ultrasonic diagnosis of OT lesions were observed. Results: There were 18 OT lesions in 17 patients, 16 case with unilateral single lesion and 1 case with bilateral single lesions, 2 (2/18, 11.11%) lesions grew outward of ovary and 16 (16/18, 88.89%) in the ovary. Fifteen lesions were found with preoperative ultrasound and 3 were missed. Among 15 OT lesions detected with ultrasound, the shape of 8 were regular and 7 were lobed, the border of 14 lesions were clear and of 1 obscure, 12 were solid, 2 were cystic and solid and 1 was cystic,11 were homogeneous hypoecho while 4 were uneven hypoecho, when CDFI found few blood signal in 9 and no blood signal in rest 6 lesions. Liquefaction was found in 3 lesions, while 12 had no liquefaction. All 15 OT lesions had no calcification, while 2 lesions had posterior echo attenuation and 13 had no attenuation. Among these 15 lesions, preoperative ultrasound only correctly diagnosed 1 lesion as OT. In the rest 14 lesions, preoperative ultrasound could not locate and quantitate 1 lesion, while located 6 lesions as ovarian source tumors, the nature of the tumor was not clear, miss diagnosed 5 as ovarian malignancy or ovarian teratoma, 2 as subserous myoma of uterus/broad ligament myoma. Conclusion: Ultrasonic findings of OT are lack of specificity, easy to be missed when relative small OT coexisting with other large ovarian lesions, or misdiagnosed as subserous myoma of uterus, broad ligament myoma and other ovarian tumors.

Result of carrier screening for spinal muscular atrophy among 3049 reproductive-age individuals from Yunnan region / 中华医学遗传学杂志

OBJECTIVE@#To perform carrier screening for spinal muscular atrophy (SMA) among 3049 reproductive-age individuals from Yunnan region and determine the copy number of survival motor neuron (SMN) gene and carrier frequencies.@*METHODS@#Multiplex ligation-dependent probe amplification (MLPA) was used to determine the copy number of exon 7 of SMN1 and SMN2 genes and identify those with a single copy of SMN1 gene. Prenatal diagnosis was performed for couples whom were both found to be SMA carriers.@*RESULTS@#In total 62 SMA carriers were identified among the 3049 subjects, which yielded a carrier frequency of 1 in 49 (2.03%). No statistical difference was found in the carrier frequency between males and females (1.91% vs. 2.30%, P>0.05). Respectively, 1.3% (41/3049) and 0.69% (21/3049) of the carriers were caused by heterozygous deletion and conversion of the SMN1 gene. The average copy number for SMN1 alleles was 1.99. Two couples were found to be both as SMA carriers, for whom the birth of an affected fetus was avoided by prenatal diagnosis.@*CONCLUSION@#No difference was found in the carrier frequency of SMA-related mutations between the two genders in Yunnan region, which was in keeping to an autosomal recessive inheritance pattern. Determination of the carrier frequency for SMA and SMN gene variants may provide a basis for genetic counseling and prenatal diagnosis for the disease.

Regulatory effects of sphingosine kinase-2 on astrocyte function and EAE progression in mouse model / 中华微生物学和免疫学杂志

Objective:To investigate the regulatory effects of sphingosine kinase-2 (SphK2) on the function of activated astrocytes and the progression of experimental autoimmune encephalomyelitis (EAE) in mice.Methods:Primary mouse astrocytes were isolated from wild-type (WT) C57BL/6 mice and sphk2 gene knock-out ( sphk2 -/-) mice and stimulated in vitro with interleukin 17 (IL-17). Real-time PCR was used to measure the expression of inflammatory cytokines and chemokines at mRNA levels. Western blot and immunofluorescence were used to detect the expression of glial fibrillary acidic protein (GFAP) and the phosphorylation of signal transducer and activator of transcription 3 (p-STAT3). An EAE mouse model was constructed using myelin oligodendrocyte glycoprotein 35-55 (MOG 35-55) polypeptide. Western blot was used to detect the expression of GFAP and p-STAT3 at protein level and real-time PCR was used to detect the expression of inflammatory cytokines and chemokines at mRNA level in spinal cords. Hematoxylin-Eosin (HE) and Luxol fast blue (LFB) staining were used to observe the changes in inflammatory cell infiltration and demyelination in spinal cords. Results:Compared with the WT group, the phosphorylation of STAT3 was obviously reduced in in vitro activated mouse astrocytes of sphk2 -/- mice, and the expression of inflammatory cytokines and chemokines including monocyte chemotactic protein 1 (MCP-1), TNF-α and IL-6 at mRNA level was also significantly decreased. Compared with the WT EAE group, changes in the above-mentioned cytokines and relative proteins in sphk2 -/- EAE mice in vivo were similar to those in vitro. Moreover, inflammatory cell infiltration and demyelination were significantly reduced in spinal cords of sphk2 -/- EAE mice. However, no significant difference in in vitro or in vivo GFAP expression was observed between WT and sphk2 -/- mice. Conclusions:SphK2 might regulate the function of reactive astrocytes through STAT3 molecular pathway, thereby regulating the production of inflammatory cytokines and chemokines and participating in the pathological process of EAE.

The diagnostic value of multimodal transvaginal contrast-enhanced ultrasonography combined with fallopian tube patency / 中华超声影像学杂志

Objective:To investigate the diagnostic value of multimodal transvaginal contrast-enhanced ultrasound combined with fallopian tubal patency.Methods:From November 2017 to November 2019, 212 patients with infertility were treated in the outpatient clinic of the Union Hospital of Fujian Medical University. Multimodal contrast-enhanced ultrasonography was used to analyze the diagnostic efficiency differences of four-dimensional transvaginal contrast-enhanced hysterosalpinx (TVS 4D-HyCoSy, 4D), three-dimensional contrast-enhanced hysterosalpingography (TVS 3D-HyCoSy 3D), two-dimensional contrast-enhanced hysterosalpingography (TVS 2D-HyCoSy 2D), and transvaginal harmonic imaging (TVS harmonic imaging, HI) individually and in different combinations. The diagnostic efficiency of tubal patency was compared between the high seniority group and the low seniority group.Results:Two patients gave up the examination because of pain. A total of 420 salpingography in 210 patients were successful, of which 375 were unobstructed and 45 were blocked (13 on the right, 18 on the left, and 7 on both sides). In high seniority group and low seniority group, the diagnostic efficiencies of different mode combinations on fallopian tubal patency were significantly different ( P<0.01). There were significant differences( P<0.05) between 4D+ 3D+ 2D+ HI group and 4D+ 3D+ 2D group, 4D+ 3D+ 2D group and 4D+ 3D group, 4D+ 3D group and 4D group. There was no significant difference ( P>0.05) in the diagnostic efficiency of fallopian tubal patency between high seniority group and low seniority group. Conclusions:The diagnostic efficiency of multimodal transvaginal contrast-enhanced ultrasound combined is higher than that of single mode, and ultrasound doctors in both high and low seniority groups can effectively diagnose fallopian tubal patency, which has important clinical value in the diagnosis of fallopian tube patency.

Analysis of the diagnosis and missed diagnosis of intrauterine adhesions by three-dimensional transvaginal sonography Render and tomographic ultrasound imaging / 中华超声影像学杂志

Objective:To investigate the value of three-dimensional transvaginal sonography(3D-TVS) in the diagnosis of intrauterine adhesions(IUA) and to analyze the causes of the missed diagnosis.Methods:Forty-seven patients with IUA were examined by three-dimensional transvaginal sonography(3D-TVS), 3D volume imaging (Render imaging) and tomographic ultrasound imaging(TUI imaging) in the Union Hospital of Fujian Medical University from January 2017 to July 2019. The abnormal echo data of the endometrium were recorded and analyzed, and the ultrasound diagnosis and hysteroscopic diagnosis were compared.Results:3D-TVS correctly diagnosed IUA was accurate in the 39 cases whose ultrasound imaging showed an uneven thickness of endometrial echo with the uterine cavity line having different degrees of echo continuity interruption. The diagnostic accuracy rate was 83.0%(39/47). In the Render imaging, 7 cases showed endometrial echo with honeycomb change, 28 cases showed partial echo loss with irregular low echo zone or low echo, and 4 cases showed corneal disappearance of one side. In TUI imaging, the endometrium was partly thinned in varying degrees where echo continuity was interrupted with hypoechoic band-like changes in all 39 cases. Three of the 8 missed IUA cases showed slender endometrium with filiform or membranous adhesions, and the other 5 were patients with uterine endometrial polyps.Conclusions:3D-TVS, Render imaging and TUI imaging technology can display stereo images, which contributes to the better preoperative diagnosis and postoperative follow-up. Care should be taken to avoid missed diagnosis and to improve the diagnostic accuracy for IUA by the techniques.

Construction and application of the multi-module training program of core competence for junior midwives / 中国实用护理杂志

Objective Constructing and applying the multi-module training program for junior midwives to improve the training quality. Methods The training program was constructed according to different modules of core competence. 11 junior midwives were selected from a hospital in Zhengzhou as trainees. The training effect was evaluated. Results Their core competence has been enhanced in theory knowledge, operating skills and training satisfaction (P<0.05). After training, the rate of episiotomy and the rate of postpartum hemorrhage were lower than before (P<0.05). Conclusion The multi-module training program can improve the core competence of junior midwives, which provides effective training method and promotes the quality of training.

Construction and application of the multi-module training program of core competence for junior midwives / 中国实用护理杂志

Objective@#Constructing and applying the multi-module training program for junior midwives to improve the training quality.@*Methods@#The training program was constructed according to different modules of core competence. 11 junior midwives were selected from a hospital in Zhengzhou as trainees. The training effect was evaluated.@*Results@#Their core competence has been enhanced in theory knowledge, operating skills and training satisfaction (P < 0.05). After training, the rate of episiotomy and the rate of postpartum hemorrhage were lower than before (P<0.05).@*Conclusion@#The multi-module training program can improve the core competence of junior midwives, which provides effective training method and promotes the quality of training.

2-Chloroethanol enhanced the expression of AQP4 protein in astrocytes via NMDAR/cAMP/PKA signaling pathway / 中国职业医学

OBJECTIVE: To investigate the role of N-methyl-D-aspartate receptor(NMDAR)/cyclic adenosine monophosphate(cAMP)/protein kinase A(PKA) signaling pathways in regulating 2-chloroethanol-induced aquaporin-4(AQP4) expression in astrocytes(AS). METHODS: i) AS in logarithmic growth phase were treated with 2-chloroethanol at the doses of 0.0, 7.5, 15.0 and 30.0 mmol/L for 12 hours, and the cells were collected for detection. ii) The AS in logarithmic growth phase were divided into blank control group, inhibitor control group, 2-chloroethanol group, and inhibitor intervention group. The inhibitor included dizocilpine(MK-801) and N-(2-[p-bromocinnamylamino-]ethyl)-5-isoquinolinesulfonamide(H89). The blank control group did not receive any treatment. The inhibitor control group was treated with a concentration of 10.0 μmmol/L MK-801 or 15.0 μmmol/L H89. The MK-801 intervention group was pretreated with MK-801 at a concentration of 10.0 μmmol/L for 30 minutes. The H89 intervention group was pretreated with H89 at a concentration of 15.0 μmmol/L for 1 hour. After the intervention, the AS in 2-chloroethanol group and MK-801, H89 intervention group were stimulated with 2-chloroethanol at a dose of 30.0 mmol/L for 12 hours. iii) The AS in each group were collected and used for Western blotting and real-time fluorescence quantitative polymerase chain reaction analysis to detect the protein and mRNA expression of AQP4, NMDAR receptor main subunit(NR1), NMDAR receptor 2 B subunit(NR2 B) and calmodulin dependent protein kinaseⅡ(CaMKⅡ). The Western blotting was adopted to detect the expression of phosphorylase-CaMKⅡ(p-CaMKⅡ) and PKA. Colorimetric method was used to detect the concentration of calcium(Ca~(2+)) in AS. The enzyme-linked adsorption test was used to measure adenylate cyclase(AC) activity and cAMP levels. RESULTS: i) The relative expression of protein and mRNA of AQP4, NR1 and NR2 B, PKA at protein level and CaMKⅡ at mRNA level, and the ratio of p-CaMKⅡ/CaMKⅡ protein, the concentration of Ca~(2+), AC activity and cAMP level in 30.0 mmol/L group were higher then those of 0.0 mmol/L group in AS(P<0.05). The relative protein expression of PKA and the concentration of Ca~(2+) increased with the increase of 2-chloroethanol(P<0.05). ii) The relative protein expression of AQP4 and the concentration of Ca~(2+) in the 2-chloroethanol group were higher than that of the blank control group and MK-801 control group(P<0.05). The relative protein expression of AQP4 and the concentration of Ca~(2+) in MK-801 intervention group were lower than that in 2-chloroethanol group(P<0.05). The relative protein expression of AQP4 and PKA in 2-chloroethanol group were higher than that of the blank control group and H89 control group(P<0.05). The relative protein expression of AQP4 and PKA in H89 intervention group was lower than that in 2-chloroethanol group(P<0.05). CONCLUSION: The 2-chloroethanol timulation induces the expression of AQP4 by activating NMDAR/cAMP/PKA signaling pathway in AS.

DNA recognition patterns of the multi-zinc-finger protein CTCF: a mutagenesis study

CCCTC-binding factor (CTCF) is a zinc-finger protein, serving an important part in the genome architecture as well as some biochemical processes. Over 70,000 CTCF binding DNA sites have been detected genome-wide, and most anchors of chromatin loops are demarcated with the CTCF binding. Various protein or RNA molecules interact with DNA-bound CTCF to conduct different biological functions, and potentially the interfaces between CTCF and its cofactors can be targets for drug development. Here we identify the effective region of CTCF in DNA recognition, which defines the exposed CTCF surface feature for the interaction of cofactors. While the zinc-finger region contributes the most in DNA association, its binding affinity varies based on different DNA sequences. To investigate the effectiveness of individual zinc-fingers, the key residues are mutated to inactivate the DNA binding ability, while the finger configuration and the spacing between fingers are preserved. The strategy is proved to be successful, while clear differences are observed in the DNA binding affinities among the 11 finger mutants and the result is consistent to previous studies in general. With the help of inactivated finger mutants, we identify the ineffective fingers and the dominant effective fingers, which form distinctive patterns on different DNA targets.

Clinical analysis of 251 cases of Pseudomonas aeruginosa bloodstream infection / 中国感染与化疗杂志

Objective To review the clinical characteristics and risk factors of Pseudomonas aeruginosa bloodstream infections. Methods The clinical data of P. aeruginosa bloodstream infections in the First Affiliated Hospital of Soochow University from January 2007 to December 2016 were analyzed retrospectively. Results Of the 251 patients identified, APACHE Ⅱ score on admission was 11.5±5.2. Majority (98.4％, 247/251) of the patients had fever. Leukopenia was found in 125 patients, leukocytosis in 87 patients, neutropenia in 122 patients, agranulocytosis in 113 patients, anemia in 193 patients, and hypoalbuminemia in 120 patients. Overall, 219 patients had at least one underlying disease, primarily hematological malignancy, or malignant solid tumor. Most (229) patients received invasive procedures such as deep venous catheter, urinary catheter, mechanical ventilation before blood sampling. P. aeruginosa was isolated from 108 of the 173 deep venous catheters. In addition, 130 patients received radiation or chemotherapy. Immunosuppressive agents were used in 124 patients. Among the 251 strains of P. aeruginosa, 87.3％ were susceptible to amikacin, followed by ciprofloxacin (85.7％) and cefepime (81.6％). Multidrug-resistant P. aeruginosa was isolated from 36 (14.3％) patients, and extensively drug resistant strain was isolated from 7 patients. All the 251 patients were treated withantimicrobial agents, mainly β-lactam/β-lactamase inhibitor combinations, carbapenems or fluoroquinolones. Overall, 20 (8.0％) of the 251 patients died, 37 (14.7％) refused further therapy due to worsening condition, and 194 (77.3％) improved. Binary logistic regression analysis showed that high APACHE Ⅱ score on admission, anemia and hypoalbuminemia were risk factors for poor outcome of bloodstream infectionscaused by P. aeruginosa. Conclusions P. aeruginosa bloodstream infection occurs more readily in immunocompromised patients. High APACHE Ⅱ score on admission, anemia and hypoalbuminemia are associated with poor prognosis. Appropriate empiric antimicrobial treatment as early as possible can improve the prognosis of P. aeruginosa bloodstream infection.

Relationship of aspect transverse ratio and capsule invasion of papillary thyroid microcarcinoma with different size and location / 中国医学影像技术

Objective To analyze the ultrasonic features of papillary thyroid microcarcinoma (PTMC) with different size and location,and to investigate the relationship of aspect transverse ratio (A/T) and capsule invasion in PTMC.Methods Totally 407 patients of PTMC with 495 nodules confirmed by pathology were enrolled.The nodules were divided into largest diameter≤0.5 cm group and largest diameter＞0.5 cm group.The ultrasonic signs of nodules were observed,and the relationship between A/T and thyroid capsule invasion was analyzed.Results The differences of blood type,relationship with capsule,calcification,morphology and A/T were statistically significant between the two groups (all P＜0.05).In nodules closed to capsule and A/T≥1,the rate of capsule invasion in diameter＞0.5 cm group (117/185,63.24％) was higher than that in diameter≤0.5 cm group (25/61,40.98％,P＜0.01).Taking A/T≥1 as the standard,the sensitivity of A/T in estimating capsule invasion of nodules closed to capsule in diameter≤0.5 cm group and diameter＞ 0.5 cm group was 89.29％ and 73.58％,the specificity was 29.41％ and 37.61％,respectively.In nodules adjacent to capsule and broken through capsule,the difference of capsule invasion rate was not significant between nodules with A/T≥1 and A/T＜1 (both P＞0.05).In nodules that contact capsule,the capsule invasion rate of A/T≥1 nodules (46/67,68.66％) was higher than that of A/T＜1 (10/27,37.04％).Taking A/T≥1 as the standard,the sensitivity of A/T in estimating capsule invasion of nodules touched capsule was 82.14％,and the specificity was 44.74％.Conclusion Ultrasonography can show the size,A/T and relationship with capsule in PTMC,which can provide diagnostic evidences in judging capsule invasion of PTMC.

Epidemiological features of brucellosis from 2010 to 2016 in Weinan, Shaanxi Province / 中华地方病学杂志

Objective To analyze the trend and epidemiological features of human brucellosis from 2010 to 2016 in Weinan,Shaanxi Province,and to provide scientific data for effective control of human brucellosis.Methods The data were obtained from the China Disease Surveillance Information Management System and the brucellosis monitoring and epidemic survey of Weinan City from 2010 to 2016.The "three distribution" of brucellosis,the positive rate of serological test of the key group and the infecting route of the epidemic were analyzed using descriptive epidemiology method.Results A total of 1 624 brucellosis cases were reported in Weinan City from 2010 to 2016 and none death case occurred.The morbidity rose from 1.96/100 000 in 2010 to 11.16/100 000 in 2014.The morbidity fallen from 5.46/100 000 in 2015 to 3.33/100 000 in 2016.The reported cases distributed in 10 counties of Weinan and mainly concentrated in Dali County (682 cases),Chengcheng County (269 cases) and Pucheng County (225 cases),accounting for 72.41％ (1 176/1 624).There were brucellosis occurring in all months of the year.The age group distributed mainly from 40 to 64 years old which accounted for 63.67％ (1 034/1 624) of the total reported cases.The peasants accounted for 91.75％ (1 490/1 624) in all cases.Men were significantly higher than women and the morbidity ratio of male to female was 3.28 ∶ 1.00 (1 245 ∶ 379).A total of 15 126 key occupational persons were tested and the positive rate of serology was 2.21％ (335/15 126) annually.The feeding and delivery process were the main ways of infection.Conclusions The brucellosis epidemic in Weinan shows a trend from rise to decline.The middle-aged male peasants account for major population in all cases and the ways of infection are diversified.

Identification of compound heterozygous mutations p.Gly400Val and p.Arg532Ter of the F11 gene in a Chinese patient with hereditary factor XI deficiency / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To investigate the phenotype and genotype defect characteristics of a Chinese patient with hereditary factor XI deficiency.</p><p><b>METHODS</b>The activated partial thromboplastin time (APTT), prothrombin time (PT), FXI activity (FXI:C) of the proband and his relatives were measured by a clotting method using automatic coagulation analyzer. FXI antigen (FXI:Ag) was assayed by enzyme-linked immunosorbent assay (ELISA). Fifteen exons of the F11 gene were amplified by PCR and sequenced. Pymol software was used to analyze the novel mutations.</p><p><b>RESULTS</b>The APTT of the proband was significantly prolonged (70.3 s, reference 34.5 s) with decreased FXI activity (6%, reference 50%-150%) and FXI antigen (1.9%, reference 50%-150%). The FXI activity and FXI antigen of his son was 31% and 39%, respectively. Two heterozygous F11 mutations were identified in the proband, which included a G to T substitution at nucleotide 1296 in exon 11 resulting in substitution of glycine by valine at codon 400 (p.Gly400Val) and a A to T substitution at nucleotide 1691 in exon 14 resulting in substitution of arginine (AGA) by a termination codon (TGA) at codon 532 (p.Arg532Ter). Analysis using Pymol indicated that the number of hydrogen bonds has changed, which led to a transformation of the structure of the FXI protein. The son of the proband was found to be heterozygous for the c.1296G to T (p.Gly400Val) mutation. NM_13142 c.1691A to T (p.Arg532Ter) is a novel mutation based on HGMD professional 2016.4. Based on 2015 Guidelines of ACMG, it is PVS1 (very strong pathogenicity).</p><p><b>CONCLUSION</b>The compound heterozygous mutations of F11 NM_13142 c.1296G to T (p.Gly400Val) and F11 NM_13142 c.1691A to T(p.Arg532Ter) probably underlies the FXI deficiency in the proband.</p>

DNA Vaccines Encoding Toxoplasma gondii Cathepsin C 1 Induce Protection against Toxoplasmosis in Mice

Toxoplasma gondii cathepsin C proteases (TgCPC1, 2, and 3) are important for the growth and survival of T. gondii. In the present study, B-cell and T-cell epitopes of TgCPC1 were predicted using DNAstar and the Immune Epitope Database. A TgCPC1 DNA vaccine was constructed, and its ability to induce protective immune responses against toxoplasmosis in BALB/c mice was evaluated in the presence or absence of the adjuvant α-GalCer. As results, TgCPC1 DNA vaccine with or without adjuvant α-GalCer showed higher levels of IgG and IgG2a in the serum, as well as IL-2 and IFN-γ in the spleen compared to controls (PBS, pEGFP-C1, and α-Galcer). Upon challenge infection with tachyzoites of T. gondii (RH), pCPC1/α-Galcer immunized mice showed the longest survival among all the groups. Mice vaccinated with DNA vaccine without adjuvant (pCPC1) showed better protective immunity compared to other controls (PBS, pEGFP-C1, and α-Galcer). These results indicate that a DNA vaccine encoding TgCPC1 is a potential vaccine candidate against toxoplasmosis.

Clinical significance of seven autoantibodies determination in the diagnosis of lung cancer / 临床检验杂志

Objective To investigate the diagnostic value of seven tumour-associated autoantibodies,including p53,PGP9.5,SOX2,GAGE7,GBU4-5,MAGE A1 and CAGE autoantibodies,in the newly diagnosed patients with lung cancer.Methods A total of 108 patients with newly diagnosed lung cancer,120 with benign lung diseases,227 with other cancers and 96 healthy controls were enrolled in the study.Their serum levels of seven autoantibodies were detected by enzyme linked immunosorbent assay (ELISA).The ROC curve was drawn and used to analyze their diagnostic efficiency for lung cancer.The diagnostic value of the combination of seven autoantibodies in different groups was also compared.Results The serum levels of seven autoantibodies in lung cancer patients were significantly higher than those in the patients with benign lung diseases or other cancers and healthy controls (P ＜ 0.05).The sensitivity,specificity and AUCROc of the combination of seven autoantibodies in the preliminary diagnosis of lung cancer were 62.00％,89.80％ and 0.769,respectively,and its sensitivity and AUCROC were higher than those of single autoantibody.The positive rate of the combination of seven autoantibodies in lung cancer patients was significantly higher than those in healthy controls (x2 =50.885,P ＜ 0.01) and the patients with benign lung diseases (x2 =56.341,P ＜ 0.01) or other cancers (x2 =46.812,P ＜ 0.01).Conclusion The combination detection of seven autoantibodies,including p53,PGP9.5,SOX2,GAGE7,GBU4-5,MAGE A1 and CAGE,may serve as potential markers for the diagnosis of lung cancer.