Objective:To assess the value of cerebral small vessel disease (CSVD) burden in predicting prognosis in acute ischemic stroke (AIS) patients with anterior circulation large vessel occlusion (LVO) after endovascular therapy (EVT).Methods:The study was a cross-sectional study. A total of 242 patients with AIS due to anterior circulation LVO received EVT in the First Affiliated Hospital of Nanjing Medical University from February 2018 to September 2022. The clinical and imaging data of all patients were analyzed retrospectively. On follow-up MRI within 7 days after EVT, CSVD features [white matter hyperintensity (WMH), lacune, perivascular space, cerebral microbleed, cerebral atrophy] and CSVD burden score (0-5) was evaluated. Modified Rankin scale (mRS) score at 90 days after EVT was assessed. Patients were categorized into a mild burden group (0-1 points) and a moderate-severe burden group (2-5 points) based on CSVD burden score. Meanwhile, patients were categorized into a good prognosis group (0-2 points) and a bad prognosis group (3-6 points) based on mRS score at 90 days after EVT. Mann-Whitney U test and χ2 test were used to compare the difference of clinical and imaging indexes between the 2 groups, and variables with P<0.1 in the univariate analysis were included in the multifactorial logistic regression to screen for independent factors to predict the prognosis. Results:There were 169 patients in the good prognosis group and 73 patients in the bad prognosis group out of 242 patients. Compared with the good prognosis group, age, incidence of hyperlipidemia, baseline National Institutes of Health Stroke Scale (NIHSS) scores, incidence of hemorrhagic conversion, CSVD burden scores, incidence of periventricular WMH scores of 3 and/or deep WMH scores≥2, and incidence of moderate-severe cerebral atrophy of patients in the bad prognosis group were higher, and the incidence of complete recanalization was lower (all P<0.05). Multivariate analysis showed hyperlipemia ( OR=8.438, 95% CI 1.691-42.119, P=0.009), baseline NIHSS score ( OR=1.103, 95% CI 1.047-1.162, P<0.001), complete recanalization ( OR=0.131, 95% CI 0.038-0.454, P=0.001) and hemorrhage transformation ( OR=1.952, 95% CI 1.031-3.697, P=0.040) were independent factors for the prognosis of EVT in patients with LVO AIS. There were 157 cases in the mild burden group and 85 cases in the moderate-severe burden group. The 90-day mRS score was higher in the moderate-severe burden group compared with the mild burden group ( Z=-2.24, P=0.025). Conclusion:CSVD burden has some clinical implications in predicting the prognosis of EVT in patients with anterior circulation LVO AIS.

Objective:To investigate the value of multi-parametric analysis based on dual-layer detector spectral CT (DLCT) in predicting the initial recurrence risk for papillary thyroid carcinoma (PTC).Methods:From November 2021 to October 2022, 102 PTC patients confirmed by pathology were retrospectively collected at the First Affiliated Hospital of Nanjing Medical University in this cross-sectional study. There were 25 males and 77 females, with an age of (42±13) years old. The initial recurrence risk assessment for PTC patients was categorized into a low-risk group (75 cases) and an intermediate-high-risk group (27 cases). Clinical data, including age, gender, body mass index, history of nodular goiter, history of Hashimoto thyroiditis, and preoperative thyroid function, were collected. Tumor morphological features, including size, location, shape, aspect ratio, the degree of thyroid capsule contact, calcification, and cystic change, were evaluated. Quantitative DLCT parameters, including iodine concentration (IC), standardized iodine concentration (NIC), effective atomic number (Z eff), standardized effective atomic number (NZ eff), electronic density (ED), CT values under different energy levels (40-200 keV, 30 keV intervals) and slope of energy spectrum curve (λ HU) both in the arterial and venous phase were measured. The differences in clinical, morphological features, and spectral CT quantitative parameters between the two groups were compared using independent sample ttest, Mann-Whitney U test, or χ2 test. Multivariate logistic regression analyses were used to construct three models based on clinical and morphological features, quantitative DLCT parameters and their combination, respectively. The receiver operating characteristic curve was used to evaluate the predictive performance of these models for the initial recurrence risk of PTC patients, and the area under the curve (AUC) was compared using the DeLong test. Results:Significant differences were found in gender, lesion long diameter, lesion short diameter and calcification between the low-risk group and intermediate-high-risk groups ( P<0.05). The arterial phase IC, arterial phase Z eff, arterial phase λ HU, arterial phase CT 40 keV, venous phase NIC and venous phase NZ eff in intermediate-high-risk group were significantly lower than those in the low-risk group ( P<0.05). The logistic regression analysis revealed that the clinical model included gender ( OR=2.895, 95% CI 1.047-8.002, P=0.040) and lesion long diameter ( OR=1.142, 95% CI 1.042-1.251, P=0.004), with an AUC of 0.720, sensitivity of 63.0%, and specificity of 78.7% in predicting the initial recurrence risk of PTC patients. The DLCT quantitative parameter model included arterial phase IC ( OR=0.580, 95% CI 0.370-0.908, P=0.017), venous phase NIC ( OR=0.077, 95% CI 0.011-0.536, P=0.010), and venous phase NZ eff ( OR=0.002, 95% CI 0.001-0.103, P=0.009), with an AUC of 0.774, sensitivity of 71.9%, and specificity of 70.0%. The AUC of the combined model was 0.857, with a sensitivity of 74.1%, and specificity of 88.0%, outperforming the clinical model ( Z=2.92, P=0.004) and the DLCT quantitative parameter model ( Z=2.07, P=0.046). Conclusion:Multi-parametric analysis based on DLCT can help predict the initial recurrence risk for PTC, and combining it with clinical and morphological features, the predictive accuracy can be improved.

Objective To investigate the effect of microRNA-214-3p (miR-214-3p) expression in cancer-associated fibroblasts (CAFs) on the cisplatin sensitivity of ovarian cancer cells and its mechanism. Methods Sixty-four ovarian cancer patients were selected as study subjects and divided into platinum-partially sensitive group and platinum-sensitive group based on progression-free survival after chemotherapy. Real-time quantitative polymerase chain reaction (qRT-PCR) was used to detect the relative expression of miR-214-3p in ovarian cancer tissues from the two groups, and the 2-year survival rates of patients with different clinical characteristics were compared. CAFs and normal ovarian fibroblasts (NFs) were primarily cultured, and qRT-PCR and immunofluorescence experiments were used to detect the expression of miR-214-3p and p62 protein in CAFs and NFs. The expression levels of SQSTM1 gene in different types of ovarian cells were searched through the CSIOVDB database. CAFs were transiently transfected with miR-214-3p mimic (mimic group) and miR-214-3p mimic NC (NC group), and untransfected CAFs were selected as control group. Cell culture supernatants from each group were collected, and an indirect co-culture model of ovarian cancer cell line SKOV3 with CAFs from each group was established. The CCK-8 method, DCFH-DA method, qRT-PCR, and immunoblotting were used to detect the proliferation rate, half-maximal inhibitory concentration of cisplatin (IC50), reactive oxygen species (ROS) content, and relative expression levels of miR-214-3p, cisplatin resistance gene CCND1, and autophagy protein p62 in SKOV3 cells under different culture conditions. Results The relative expression of miR-214-3p was lower in the platinum-partially sensitive group than in the platinum-sensitive group (P < 0.01). There were statistically significant differences in the 2-year survival rates among patients with different International Federation of Gynecology and Obstetrics (FIGO) stages, platinum partial sensitivity, and low miR-214-3p expression (P < 0.01). The relative expression of miR-214-3p was lower in ovarian CAFs than in NFs, while the expression level of p62 protein was higher in CAFs than in NFs (P < 0.01). Online analysis of the CSIOVDB database showed that the expression level of SQSTM1 gene in ovarian CAFs was higher than that in ovarian cancer epithelial cells and NFs (P < 0.01). Compared with SKOV3 cells indirectly co-cultured with CAFs in the control group and CAFs in NC group, SKOV3 cells indirectly co-cultured with mimic CAFs showed reduced proliferation rate, cisplatin IC50, and ROS content, increased relative expression of miR-214-3p, and decreased relative expression of CCND1 mRNA and p62 protein (P < 0.01). Conclusion The expression of miR-214-3p in CAFs is correlated with the sensitivity of ovarian cancer cells to cisplatin. Low expression of miR-214-3p in CAFs can promote the proliferation of ovarian cancer cells and ROS-mediated autophagy, thereby reducing the sensitivity of ovarian cancer cells to cisplatin.

ObjectiveTo explore the clinical efficacy of compound Wufengcao liquid （CWL） on tuberculous ulcer and the influence on macrophage polarization. Method① Clinical experiment： A total of 145 patients with tuberculous ulcer who were treated in Nanjing Integrated Traditional Chinese and Western Medicine Hospital were randomized into observation group， control group Ⅰ， and control group Ⅱ according to the random number table method. In addition to the basic anti-tuberculosis chemotherapy， CWL， Kangfuxin liquid， and isoniazid solution （local external application） were respectively used in the observation group， control group Ⅰ， and control group Ⅱ. The treatment lasted 4 weeks for each group. The total effective rate in wound healing， traditional Chinese medicine（TCM） syndrome score， and histopathological morphology of wound were observed and the expression of inducible nitric oxide synthase （iNOS） and arginase-1 （Arg-1） in wound tissue was measured. ② Cell experiment： RAW264.7 cells were cultured in DMEM （10% fetal bovine serum， 1% double-antibody solution） in a cell incubator （37 °C， 5% CO₂）. Phorbol 12-myristate 13-acetate （PMA） was used to induce the differentiation of RAW264.7 cells into macrophages. Lipopolysaccharide （LPS） was employed to stimulate polarization of macrophages into M1 type and interleukin-4 （IL-4） to induce the polarization into M2 type. Kangfuxin solution， isoniazid solution， and CWL were respectively applied to the above cell model for 36 h. The cell supernatant was collected and centrifuged. Western blot was used to detect the protein expression of tumor necrosis factor-α （TNF-α）， transforming growth factor-β （TGF-β）， iNOS， and Arg-1， and flow cytometry （FCM） to detect the expression of CD86 and CD206. Result①Clinical experiment： The total effective rate in the CWL group ［98.0% （48/49）］ was higher than that in the control group Ⅰ ［87.5% （42/48）， χ²=3.962， P<0.05］ and control group Ⅱ ［83.3% （40/48）， χ²=6.162， P<0.05］. After 28 days of treatment， compared with control group Ⅰ and control group Ⅱ， CWL decreased the TCM syndrome score （P<0.05） and obviously improved the histopathological morphology of the wound. Immunohistochemistry results showed that the iNOS expression in local focus tissue was lower （P<0.05） and the expression of Arg-1 was higher （P<0.05， P<0.01） in the CWL group than in the control group Ⅰ and control group Ⅱ after 28 days of treatment. ② Cell experiment： Western blot assay showed that the expression of iNOS and TNF-α in LPS group increased compared with that in the M0 group （P<0.01） and the expression in the LPS+ isoniazid group， LPS+ Kangfuxin group， and LPS+CWL group was lower than that in the LPS group （P<0.05）. The expression of iNOS in LPS+Kangfuxin group and LPS+ CWL group was lower than that in the LPS+isoniazid group （P<0.05， P<0.01）， and the expression of TNF-α in LPS+ CWL group was lower than that in LPS+isoniazid group （P<0.01）. The expression of TNF-α in LPS+ CWL group decreased compared with that in the LPS+ Kangfuxin group （P<0.05）. The expression of Arg-1 and TGF-β in IL-4 group was higher than that in the M0 group （P<0.01）， and the expression in the IL-4+isoniazid group， IL-4+Kangfuxin group， and IL-4+ CWL group was higher than that in the IL-4 group （P<0.05）. The expression of Arg-1 and TGF-β in the IL-4+ Kangfuxin group and IL-4+CWL group was higher than that in the IL-4+isoniazid group （P<0.05， P<0.01）， and the expression was higher in the IL-4+CWL group than in the IL-4+Kangfuxin group （P<0.05， P<0.01）. The FCM result showed that the expression of CD86 and CD206 in LPS group and IL-4 group was higher than that in M0 group （P<0.01）. CD86 expression in LPS+isoniazid group， LPS+ Kangfuxin group， and LPS+CWL group was lower than that in the LPS group （P<0.01）. The expression of CD86 in LPS+Kangfuxin group and LPS+ CWL group increased compared with that in the LPS+isoniazid group （P<0.01）， and the expression was higher in the LPS+ CWL group than in the LPS+Kangfuxin group （P<0.01）. CD206 expression in IL-4+ isoniazid group， IL-4+Kangfuxin liquor group， and IL-4+ CWL group was increased compared with that in the IL-4 group （P<0.01）. CD206 expression in IL-4+Kangfuxin liquid group and IL-4+ CWL group was decreased compared with that in the IL-4+isoniazid group （P<0.01）. CD206 expression in IL-4+CWL group was lower than that in the IL-4+ Kangfuxin group （P<0.05）. ConclusionCWL can promote the healing of tuberculous ulcers， and the mechanism is that it inhibits the expression of iNOS， TNF-α， and CD86 and promotes the expression of Arg-1， TGF-β， and CD206， thereby regulating M1/M2 polarization balance.

Objective:To investigate the value of magnetization transfer imaging (MTI) and fat suppression T 2WI (FS-T 2WI) in predicting the clinical activity of Graves ophthalmopathy (GO). Methods:From October 2020 to July 2021, 64 GO patients were prospectively enrolled in the First Affiliated Hospital of Nanjing Medical University. According to the clinical activity score (CAS), the patients were divided into active group (CAS≥3, 39 patients and 78 eyes) and inactive group (CAS<3, 25 patients and 50 eyes). The coronal MTI and FS-T 2WI were scanned for pre-treatment assessment. Magnetization transfer ratio (MTR) of extraocular muscles, and signal intensity ratio (SIR) between extraocular muscles and temporalis were measured, respectively. The independent-sample t-test was used to compare the MTR and SIR between two groups. The correlations between MRI parameters and CAS were analyzed using Spearman correlation analysis. The receiver operating characteristic (ROC) curve was performed to evaluate the value of each and combined parameters for predicting the clinical activity of GO. The DeLong test was used to compare the area under the curve (AUC). Results:The MTR of active group and inactive group were 0.45±0.04 and 0.51±0.04, respectively, the difference was statistically significant ( t=7.62, P<0.001). The SIR were 3.4±0.6 and 2.6±0.5, respectively, and the difference was also statistically significant ( t=-8.20, P<0.001). MTR was negatively correlated with CAS ( r=-0.46, P<0.001), while SIR was positively correlated with CAS ( r=0.63, P<0.001). The AUC of MTR, SIR and the combination of MTR and SIR for predicting the clinical activity of GO were 0.840, 0.845 and 0.905, respectively. The combination of MTR and SIR showed higher performance than MTR or SIR alone, and the differences were statistically significant ( Z=2.61, P=0.009; Z=2.15, P=0.032). Conclusions:The quantitative parameters of MTI and FS-T 2WI, namely MTR and SIR, can be used to evaluate the clinical activity of GO. Integrating MTI and FS-T 2WI can improve the diagnostic efficiency.

Objective:To explore the diagnostic value of radiomics based on arterial-venous mixed images derived from dual-energy CT (DECT) data in diagnosis of cervical lymph nodes (LNs) metastasis of papillary thyroid cancer (PTC).Methods:From June 2017 to December 2018, eighty-four patients with preoperatively DECT scanning and pathologically confirmed PTC (129 non-metastatic LNs and 97 metastatic LNs) in the First Affiliated Hospital of Nanjing Medical University were included in this study. The clinical and imaging data of all patients were retrospectively analyzed. The training cohort consisted of 62 PTC cases with 156 LNs (91 non-metastatic LNs and 65 metastatic LNs). An independent validation cohort consisted of 22 PTC patients with 70 LNs (38 non-metastatic LNs and 32 metastatic LNs). Semi-automatic LNs segmentation was conducted on arterial-venous mixed images derived from DECT using Syngo.via Frontier Radiomics software. Totally 1 226 radiomics features were extracted from arterial-venous mixed images for each LN. The least absolute shrinkage and selection operator (LASSO) regression was applied for radiomics features selection and signature building. The logistic regression modeling was used to construct diagnostic models based on the CT image features of LNs (model 1), the radiomics signature (model 2) and the combination of the CT image features and radiomics signature (model 3). An intuitive nomogram was plotted for model 3. The ROC curve analyses and area under the curve (AUC) were performed to evaluate the diagnostic efficiency of the three models, with the performances compared using the Delong test.Results:Model 1 was developed with LNs shape, degree of enhancement, pattern of enhancement, calcification and extra nodal extension. Three arterial phase radiomics features were selected and used to establish radiomics signature using LASSO regression (model 2). Model 3 was developed with LNs size, shape, degree of enhancement and radiomics signature. In both the training and validation cohort, model 3 showed the best diagnostic performance (AUC=0.965, 0.933), followed by model 2 (AUC=0.947, 0.910), and both these two models significantly outperformed model 1 (AUC=0.850, 0.846) (training cohort, Z=4.066 and 3.758, P both<0.001; validation cohort, Z=2.871 and 1.998, P=0.017 and 0.042) respectively. Conclusion:The radiomics model based on arterial-venous mixed images derived from DECT data can realize effective diagnosis of LNs metastasis in patients with PTC; and the combination model of radiomics signature with CT image features can further improve the diagnostic accuracy.

HuR (human antigen R), an mRNA-binding protein responsible for poor prognosis in nearly all kinds of malignancies, is a potential anti-tumor target for drug development. While screening HuR inhibitors with a fluorescence polarization (FP) based high-throughput screening (HTS) system, the clinically used drug eltrombopag was identified. Activity of eltrombopag on molecular level was verified with FP, electrophoretic mobility shift assay (EMSA), simulation docking and surface plasmon resonance (SPR). Further, we showed that eltrombopag inhibited cell proliferation of multiple cancer cell lines and macrophages, and the anti-tumor activity was also demonstrated in a 4T1 tumor-bearing mouse model. The data showed that eltrombopag was efficient in reducing microvessels in tumor tissues. We then confirmed the HuR-dependent anti-angiogenesis effect of eltrombopag in 4T1 cells and RAW264.7 macrophages with qRT-PCR, HuR-overexpression and HuR-silencing assays, RNA stability assays, RNA immunoprecipitation and luciferase assays. Finally, we analyzed the anti-angiogenesis effect of eltrombopag on human umbilical vein endothelial cells (HUVECs) mediated by macrophages with cell scratch assay and Matrigel angiogenesis assay. With these data, we revealed the HuR-dependent anti-angiogenesis effect of eltrombopag in breast tumor, suggesting that the existing drug eltrombopag may be used as an anti-cancer drug.

1,3-1,4-β-glucanase (E.C.3.2.1.73) is an important industrial enzyme which cleave β-glucans into oligosaccharides through strictly cutting the β-1,4 glycosidic bonds in 3-O-substituted glucopyranose units. Microbial 1,3-1,4-β-glucanase belongs to retaining glycosyl hydrolases of family 16 with a jellyroll β-sandwich fold structure. The present paper reviews the industrial application and protein engineering of microbial β-glucanases in the last decades and forecasts the research prospects of microbial β-glucanases.
Amino Acid Sequence ; Glycoside Hydrolases ; Models, Molecular ; Protein Engineering ; Substrate Specificity

The autolysis of brewer's yeast seriously affects the quality of beer and the quality of yeast is considered as one of the key factors in beer brewing. Previous studies on brewer's yeast autolysis showed that RLM1 gene, an important transcription factor in cell integrity pathway, is closely related to the autolysis of yeast. In this study, RLM1 was knocked out and overexpressed in a haploid brewer's yeast. RLM1 disruption resulted in poor anti-autolysis performance of yeast, whereas overexpression of RLM1 contributed to the anti-autolytic ability of yeast. In addition, RLM1 gene knockout affected the osmotic stress resistance, cell wall damage resistance, nitrogen starvation resistance and temperature tolerance of yeast strain. The transcriptional level of GAS1 involved in cell wall assembly and DNA damage response was regulated along with the expression of RLM1, whereas other genes in CWI pathway did not show apparent regularity. RLM1 might mainly affect the expression of GAS1 so as to improve the stress resistance of lager yeast in harsh environment. The result from this study help further understand the mechanism of yeast autolysis and lay a foundation for breeding brewer's yeast strain with better anti-autolytic ability.
Autolysis ; Beer ; Cell Wall ; Humans ; MADS Domain Proteins ; Saccharomyces cerevisiae ; Saccharomyces cerevisiae Proteins

Objective: To introduce the method of galvanic vestibular stimulation-vestibular evoked myogenic potentials (GVS-VEMP) as well as to observe and analyze the parameters and elicited rate of GVS-cVEMP and GVS-oVEMP in healthy young people in China. Methods: Twenty six normal young subjects were recruited for conventional examinations of GVS-VEMP. The subjects were 21-37 years old, average age was (25.8±3.7) years old, including 13 males and 13 females. The galvanic stimulation intensity of 3 mA/1 ms was used to evoke cVEMP and oVEMP on the sternocleidomastoid and inferior extraocular muscles respectively, and the intensity of stimulus was decreased until the response disappeared, the threshold, latency, amplitude, interval phase and interaural amplitude ratio(IAR) were calculated. SPSS18.0 software was used for statistical analysis. Results: All subjects were elicited normal GVS-cVEMP and GVS-oVEMP under 3 mA/1 ms, the elicited rate was 100%. The threshold of GVS-cVEMP was (1.18±0.47) mA, p1 latency was (10.43±1.54) ms, n1 latency was (17.91±1.20) ms, the amplitude was (102.47±56.77) uV and IAR was (0.26±0.20). The threshold of GVS-oVEMP was (1.12±0.50) mA, n1 latency was (8.46±1.05) ms, p1 latency was (11.83±1.27) ms, the amplitude was (9.12±6.82) uV and IAR was (0.25±0.20). In terms of gender and lateral comparison, only the GVS-oVEMP amplitude was higher for male than for female, which had significant statistical difference (P<0.05), and there was no statistical difference in the other parameters between GVS-cVEMP and GVS-oVEMP. Conclusion GVS-cVEMP and GVS-oVEMP could be elicited in healthy youth population, and the parameters could provide reference for subsequent vestibular function evaluation.