A review of studies on the influence of impurities on protein crystallization is given.The possible sources of impurities and its effect on the protein crystallization are presented.The effects of impurities on protein crystallization,including nucleation,macroscopic morphologies,microscopic surface morphologies,growth rates,kinetics,quality,and repartitioning of impurities are reviewed.

Abstract: Objective　To explore the antiviral effect of baricitinib in the SARS-CoV-2 infection and influence on cytokine levels. Methods　Calu-3 cells were infected with SARS-CoV-2 at MOI of 0.1, and the levels of inflammatory cytokines (IL-6, IL-8, TNF-α and IL-1β), interferon β (IFN-β) and interferon-stimulated gene, IFIT2 in the infected cells were analyzed by qRT-PCR methods. At the same time, Calu-3 cells were infected with SARS-CoV-2 (MOI=0.1) after being treated with baricitinib for 2 hours. Cells were collected at 0, 24, 36, and 48 hours, and analyzed for the mRNA of the above genes in the drug-treated and untreated groups. Results　The mRNA levels of IL-6, TNF-a, IL-1β, IFN-β and IFIT2 in Calu-3 infected by SARS-CoV-2 cells were increased significantly. These cytokines were increased by nearly 100-fold post-infection 48 h compared with the control (P<0.000 1), and continued to increase with the infection time (P<0.001 or P<0.000 1). The increase of IL-8 mRNA level was not as significant as IL-6, TNF-α, IL-8, IL-1β, but it also showed a 2-4 folds increase. Baricitinib does not affect the level of viral RNA in Calu-3 cells after SARS-CoV-2 infection (P>0.05). However, baricitinib can significantly inhibit the up-regulation of IL-6 and TNF-α levels induced by SARS-CoV-2 infection (5.25-fold and 3.90-fold down-regulation, respectively, P<0.01), and has little effect on the levels of IL-8 and IL-1β . In addition, the drug could also significantly down-regulate the increase in IFN-β and IFIT2 levels caused by viral infection (10.51-fold and 90.78-fold down-regulation, respectively, P<0.000 1). Conclusions　Baricitinib inhibits the release of inflammatory cytokines to some extent, but it drastically down-regulates the expression of interferons and interferon-stimulated genes (ISGs), and has limited antiviral effect on SARS-CoV-2. Considering that interferon signal pathways play important roles on viral infection, caution should be exercised when using baricitinib to treat COVID-19 patients.

According to the characteristics of sludge samples from full-scale wastewater treatment bioreac-tors, the essential total DNA extraction method for most environmental samples, lysozyme-SDS-phenol/ chloroform method, was modified to improve sample pretreatment, intensify cell lysis and enhance the effi-ciency of impurity removal. Obtain a general total DNA extraction method for industrial sludge samples. Such a method was applied for total DNA extraction of sludge samples from several running full-scale an- aerobic or aerobic bioreactors in Shijiazhuang, China. The results indicated that the modified method was suitable for all the sludge samples in this study, showing the satisfying generality. The extracted total DNA of all sludge samples were pure, with about 1.8 of A260/ A280 ratio. The method was also efficient; with average total DNA yield of over 0.7 mg/g and maximum yield of 0.85 mg/g. Moreover, all the extracted to- tal DNA samples could serve as templates directly to amplify 16S rDNA by PCR. The PCR products could be separated well by denaturing gradient gel electrophoresis (DGGE) and the DGGE band patterns were clear enough to be used for further analysis. All these facts indicated that the total DNA extraction method provided in this study could meet the requirements of sludge samples research, from full-scale wastewater treatment bioreactors, using molecular biology technologies.

The skin ulceration syndrome of sea cucumber is a kind of desease induced by bacterium.In order to investigate the bacterium of infected sea cucumber and detect the N-acyl-homoserine lactones(AHLs) se-cretion of the bacterium,7 bacterial strains were isolated from the infected sea cucumber.These strains were identified by physiological-biochemical characteristics and 16S rDNA sequence.Results show that strain C6 belongs to Tenacibaculum,strain 4 belongs to Shewanella putrefaciens group,strain TB belongs to Vibrio,strain BP2,BP3,BP4 and BP6 belong to Pseudoalteromonas,respectively.AHLs were detected with strain Agrobacterium tumefaciens KYC55.Among these bacterial strains,strain C6,4,TB,BP3 and BP4 can se-cret AHLs,while strain BP2 and BP6 can’t.And the AHLs activity differs,from the highest to the lowest are 4,TB,BP4,BP3 and C6.

Objective To design a new vacant drowning rescuer in self-help can be effective and free when swimming.Meth-ods Vacant drowning rescuer includes the vacant,life-saving device,the waist straps and the ropes.Results Vacant drowning res-cuer has light weight,high strength,and small size.It is secured to the swimmer′s waist through waist strap and easy to operate. Conclusion Vacant drowning rescuer provides the security for the water activity,which can realize self-help or save others.It can be universally applied for the majority of swimming enthusiasts.

Objective To evaluate the value of the physiological and operative severity score for the enumeration of mortality and morbidity (POSSUM) and P-POSSUM in predicting the risks of orthopedic surgeries for senile patients with femoral neck fracture.Methods A total of 108 patients with femoral neck fractures who underwent hip joint replacement were retrospectively studied using POSSUM and P-POSSUM scoring system to predict their mortality and complication rate.The difference between predictive value and observed value was analyzed by chi-square test.Meanwhile,the patients were divided into two groups based on their POSSUM scores.The differences between two groups were analyzed.Results According to POSSUM scores,47 patients were predicted to have complications(the mean rate was 43.52％),but only 37 did actually (the rate was 34.26％).There was no significant difference between predicted values and observed values (P =0.238).The predicted death toll was 11 cases (the mean rate was 10.19％),but actually only 2 patients died (the rate was 1.85％).Predicted value was higher than observed value.In terms of complications,death toll agreed well with the predicted values calculated by P-POSSUM (predicted death of 4 cases' the mean mortality being 3.70％ ; actual death of 2 cases' the mortality was 1.85％) without significant difference (P =0.625).We divided the patients into two groups with the POSSUM scores 40,and there was no significant difference between predicted values and observed values (P =0.527,P =0.285).Conclusions POSSUM has better predictive ability of morbidity,but overestimates mortality.P-POSSUM more accurately predicts mortality than POSSUM.The predicted results of POSSUM and P-POSSUM scoring systems are satisfactory in the high risk group.

Objective To discuss the significance of near infrared spectroscopy (NIRS) in evaluation of intrapartum hypoxic-ischemic cerebral injury, and to provide a method to evaluate neonatal brain damage objectively and quantitatively. Methods A total of 63 neonates with fetal distress were divided into hypoxic-ischemic encephalopathy(HIE) group and non-HIE group. Thirtyfive newborns with no fetal distress were chosen as controls. Using NIRS, the brain regional oxygen saturation(rSO2) in these neonates were measured. Evaluation of brain rSO2 in the diagnosis of HIE was analyzed with receiver operating characteristic (ROC) curve. Results At the time of fetal head visible on vulval gapping and 5 min after birth, the HIE group showed decreased brain rSO2[(36. 6±5.0)% and (52. 0±4. 2)%], comparing with control group[(45. 9±4. 6)% and (59. 6±4. 4)%]and non-HIE group[(44.1±3.1) % and (57. 6±3. 5) %](P<0. 01) . The brain rSO2 was positively correlated with the pH and oxygen saturation of umbilical artery blood in all groups (P<0. 01). When the cut-off value of brain rSO2 was <39. 5% at fetal head visible on vulval gapping, the sensitivity and specificity of assessing HIE were 67% and 93%, respectively, while 70% and 86% when the cut-off value was <53. 5% at 5 min after birth. Conclusions The brain rSO2 obtained by NIRS could be used to evaluate brain oxygenation, and may be useful in predicting HIE in neonates with fetal distress.

Objective To investigate the incidence of and clinical factors influencing neonatal birth defects from different assisted reproductive technology. Methods Between October 1998 and December 2006,1271 newborns from mothers treated by in vitro fertilization techniques [ including in vitro fertilization (IVF), intracytoplasmic sperm injection (1CSI) and thaw embryo transfer (Thaw-ET) ] matched with 269 newborns from mothers treated by artificial insemination were enrolled in Reproductive Medicine Center in First Hospital Affiliated to Zhengzhou University. Their medical information was analyzed retrospectively to compared neonatal characteristics, the incidence of birth defect and anomalous organs involved between in vitro fertilization group and artificial insemination group. Results In group of in vitro fertilization, those newborns with low birth weight from IVF, ICSI and Thaw-ET were 20. 0% ( 134/671 ), 22. 4% (92/410), 18.9% (36/190)respectively, which were more than 11.5% (31/269) cases in group of artifical semination with statistical significance (P < 0. 05 ). The rates of multiple pregnancy of 23.8% ( 160/671 ), 25.4% (104/410) ,21.1% (40/190) in subgroup of 1VF, ICSI and Thaw-ET were significantly higher than 10. 0% ( 27/269 ) in group of artifical insemination( P < 0. 05 ). The rate of macrosomia in group of in vitro fertilization was significantly lower than that of artificial insemination group (3.9% vs 8. 2%, P <0.05). However, the incidence of birth defect involved in various organs did not show significant difference between two groups ( P>0.05 ). Conclusions The incidence of multiple pregnancy demonstrated obviously increasing trends born with various In Vitro Fertilization techniques, which pave the way to high risk pregnancy. However, the incidence of newborn birth defect was not increased significantly. Thus, to lower occurrence of multiple pregnancy was the key approach to obtain neonates health.

BACKGROUND:Mouse pluripotent stem cel s are induced to differentiate into insulin-secreting cel s that can effectively improve blood glucose levels in diabetic mice. OBJECTIVE:To detect mRNA and protein levels of insulin-like cel clusters from induced pluripotent stem cel s and to investigate the function of insulin-secreting cel s in vitro and in vivo. METHODS:Mouse induced pluripotent stem cel s cultured in vitro were induced to differentiate into insulin-secreting cel s using combined inducers through three stages. The morphology of endodermal cel s, islet-derived progenitor cel s and mature islet cel s in each stage was observed and relative gene expression levels were detected by PCR. Mature insulin-like cel clusters underwent dithizone staining and functions of insulin released in vitro were observed by ELISA assay. Final y, the insulin-secreting cel s were transplanted into the subrenal capsule of diabetic mice, and then blood glucose levels were observed. RESULTS AND CONCLUSION:The mature spherical insulin-like cel clusters were successful y obtained in vitro, which were in iron red by dithizone staining, and expression of insulin mRNA was determined by PCR. The insulin-like cel clusters could secrete insulin in response to various blood glucose levels by ELISA assay. In addition, after the cel s clusters were transplanted into the subrenal capsule of mice with type 1 diabetes, the blood glucose levels were marbedly improved.

AIM: To look for harmfulless anti-leukemia drug with selective high performance, lethal effect of small hairpin RNA (shRNA) on VEGFR2 gene expression of tumor cell line HL60 in vitro.METHODS: The most effective VEGFR2 siRNA was designed and screened. The shRNA oligo was designed and pU6/VEGFR2 entry clone was constructed. HL60 was transfected transiently and vascular endothelial growth factor receptor 2(VEGFR2) expression was tested with MTT assay, RT-PCR and Western blotting. The expression clone was constructed and cotransfected with ViraPowerTM Packaging Mix into 293FTTM cells to produce Lentiviral vectors harboring Lenti6/shVEGFR2. The virion supernatant was added into HL60 cells and VEGFR2 gene inhibitory effect was determined. RESULTS: The inhibitory rates of VEGFR2 siRNA c were high. VEGFR2 expression in HL60 was inhibited by using pU6/VEGFR2 entry clone constructed with shRNA and pENTRTM/U6. For HL60 cells, the inhibitory rate was 84.9%. The expression of VEGFR2 mRNA and protein decreased significantly. 48 hours after transfection of pU6/shVEGFR2 entry clone and transduction of Lenti6/shVEGFR2 expression clone, the cell inhibitory rates were similar. Cell growth inhibitory rate of entry clone descended rapidly after this time point, the expression clone changed slowly, reaching the peak at 96 hours, dropped slightly, having no significance deviation. CONCLUSION: in vitro, VEGFR2 shRNA using lentiviral vector blocks VEGF/VEGFR2 self-secretion in HL60 cells, which inhibits leukemia development.