Determination of Serum Unbound Bilirubin and Its Clinical Application Value in Neonates / 中山大学学报(医学科学版)

ObjectiveTo investigate the clinical appropriateness and application value of the peroxidase （POD） method for the detection of unbound bilirubin （UB） in neonatal serum. MethodsHydrogen peroxide （0.33 mol/L） and three different final concentrations （0.019， 0.038， 0.075 μg/mL） of horseradish peroxidase （HRP） were added to standard bilirubin solution （1， 2， 3 μmol/L） to obtain a standardized HRP primary rate constant Kp. Then 25 μL of neonatal serum was diluted by 41.6 fold， and measured with 2.4 and 4.8 μg/mL HRP at 37 ℃ under the dark， to determine the UB concentration. The accuracy， precision， and stability of the methodology were validated. The clinical characteristics of 33 jaundiced neonates were collected， including total serum bilirubin （TSB）， indirect bilirubin （IDB）， albumin （ALB）， bilirubin to albumin molar ratio （BAMR）， etc. The experimental data were analyzed by Graphpad Prism 8.0. ResultsA standardized Kp of （7.20±1.08） mL·μg-1·min-1 was determined at pH 7.4±0.2， 37 ℃ in the dark. The HRP activity and UB concentrations remained stable at -20 ℃ for 3 weeks and a week， respectively. The mean intra-day and inter-day coefficients of variation of the serum samples with different UB concentrations were less than 10%. In this study， the UB concentrations in 33 jaundiced neonates （gestational age ≥35 weeks） were measured by the POD method in the range of （0.32～1.20） μg/dL， which was positively correlated with TSB， IDB and BAMR. Of the five infants whose UB concentrations measured more than 1 μg/dL， three received intensive phototherapy （60%）. ConclusionsThe POD method combined with a standard equipment spectrophotometer to detect serum UB concentrations in neonates is easy to operate， rapid to detect， and low cost. This method has good accuracy and precision， which is convenient for clinical implementation. Moreover， the measurement of serum UB may assist us in better management of neonatal jaundice in clinical practice.

Influence of premature rupture of membranes on the early prognosis of extremely premature infants / 中国当代儿科杂志

OBJECTIVE@#To study the influence of premature rupture of membranes (PROM) on the early prognosis of extremely premature infants, and to provide a basis for the management of extremely premature infants and prenatal consultation.@*METHODS@#A total of 179 extremely premature singleton infants who were born from 2017 to 2019 were enrolled. According to the presence or absence of PROM, they were divided into two groups: PROM group (@*RESULTS@#Compared with the non-PROM group, the PROM group had significantly higher incidence rates of earlyonset sepsis and necrotizing enterocolitis (NEC) (@*CONCLUSIONS@#PROM increases the incidence rates of early-onset sepsis and NEC in extremely premature infants and does not increase the incidence rates of other adverse outcomes. For pregnant women with PROM at the risk of extremely preterm delivery, prevention of miscarriage and chorioamnionitis is recommended to prolong gestational weeks, reduce the incidence rate of infection, and thus improve the outcome of extremely premature infants.

Association of Visit-to-Visit Variability of Blood Pressure with Cardiovascular Disease among Type 2 Diabetes Mellitus Patients: A Cohort Study

BACKGROUND: Increasing evidence has shown that visit-to-visit variability (VVV) of blood pressure (BP) is associated with an increased risk of cardiovascular disease (CVD). The objective of this study was to evaluate the impact of VVV of systolic blood pressure (SBP) and diastolic blood pressure (DBP) on the risk of CVD among patients with type 2 diabetes mellitus (T2DM) in China. METHODS: We conducted a retrospective cohort study of 10,163 T2DM patients who were not previously diagnosed with CVD from January 2008 to December 2012 in Ningbo, China. The VVV of BP was calculated using five metrics, including standard deviation (SD), coefficient of variation (CV), variation independent of mean, average real variability, and successive variability (SV) of measurements, obtained over a 24-month measurement period. Hazard ratios and 95% confidence intervals (CIs) were estimated by Cox proportional hazards regression models for the associations of variability in BP with risk of CVD. RESULTS: A total of 894 CVD events were observed during a median follow-up of 49.5 months. The hazard ratio in the highest quintile of SD of SBP was 1.24 (95% CI, 1.01 to 1.52) compared with patients in the lowest quintile. The association between higher VVV of DBP and risk of CVD was not consistent across different metrics and sensitivity analyses. CONCLUSION: Higher VVV of SBP was associated with an increased risk of CVD, irrespective of the mean SBP level. Future studies are needed to confirm these findings.

Etiologyical Analysis of 133 Patients with Elderly Macrocytic Anemia and Diagnostic Significance of Laboratory Tests / 中国实验血液学杂志

OBJECTIVE@#To study the etiology of macrocytic anemia in elderly patients and to evaluate the diagnostic significance of laborotory tests.@*METHODS@#133 elderly macrocytic anemia patients, whose age>60 years old, hemoglobin100 fL, and bone marrow cell test was performed, and these patients were grouped according to diseases, and the bilirubin, lactate dehydrogenase, folic acid, vit B12 and serum ferritin were tested, then the results of tests were compared and analyzed.@*RESULTS@#The majority of the cases were diagnosed as megaloblastic anemia (MA), myelodysplasia syndrome (MDS), acute leukemia/multiple myeloma (AL/MM) and hemolytic anemia (HA). Usually HA was a simple anemia, while others were accompanied by decrease of other 1 or 2 series. HA patients were often with significant high level of well volume (MCV), red cell distribution width(RDW), reticulocytes (RC) and indirect bilirubin (IBIL) (P<0.01). However, MA patients were often with high level of LDH. Serum ferritin (SF) level was significantly higher in both MDS and AL/MM groups (P<0.01).@*CONCLUSION@#Common causes of macrocytic anemia in elderly patients are MA, MDS, AL/MM and HA. The combination detection of MCV, RDW, RC, LDH, IBIL and SF contributes to enhancing the accuracy of diagnosis.

Intellectual Property Provisions Relating to Drug Regulation in the TPP Agreement / 中国药学杂志

OBJECTIVE: To study the intellectual property provisions relating to drug regulation in the TPP agreement, analysize the interaction mechanism between drug regulation and intellectual property rights on the basis of information sharing, study the possible impact, and then discusses the references to China. METHODS: This article analysizes the TPP final text and relevant research literatures, study the interaction mechanism between drug regulation and intellectual property rights and the possible impact of, and then discuss the relevantlaw of China. RESULTS: The TPP agreement provides several intellectual property provisions relating to drug regulation, demanding the concordant relation between the registration of pharmaceutical and patent protection period, drug data protection and patent protection, which will establish the interaction mechanism based on information sharing. The intellectual property provisions relating to drug regulation are a summary of the experience of the TPP parties forming a coordinated and interactive mechanism between encouraging innovation and ensuring drug safety and an attempt to a wide range practice, which will have abroad impact on the biopharmaceutical industry. CONCLUSION: Refer to the relevant experience in TPP agreement, China can amend the Pharmaceutical Administration Law and the Patent Law in the process of revision by improving the information sharing mechanism to achieve a higher level of institutional interaction.

Screening drugs for regulating tissue factor gene expression / 中国实验血液学杂志

This study was purposed to screen the drugs for regulating tissue factor (TF) gene expression through establishing stable cell line with luciferase gene having TF promoter transcription activity, so as to provide the basis for further studying the molecular mechanism of screened drugs. A series of luciferase reporter gene plasmids under control of 5'-truncated TF promoter (including -2174 bp - +128 bp, -684 bp - +128 bp, -247 bp - +128 bp and -201 bp - +128 bp) were constructed. The above plasmids were separately electroporated into U937 cells to establish stably transfected sublines. The function of stable cell line was testified by treatment with ATRA, the luciferase gene activity was analyzed by treating established cell line with bortezomib (BTZ) and CDA-II, and drugs for regulating TF gene expression were screened. The results indicated that the BTZ of 5 nmol/L could activate TF gene transcription activity, up-regulate the expression level of TF transcripts; CDA-II of 1 mg/ml could suppress TF gene transcription activity, down-regulate the expression level of TF transcripts. The functional analysis of TF promoter transcription revealed that the region of regulating TF promoter transcription activity by BTZ and CDA-II was between -201 to 0 bp. It is concluded that stable cell line U937 expressing luciferase activity of TF promoters is established, the novel drugs regulating TF gene expression are screened out by means of this established cell line. This study provides basis for screening the new drugs and further studying their molecular mechanisms.

Establishment of stable subline of K562 cells overexpressing high mobility group B1 protein / 中国实验血液学杂志

This study was aimed to establish a stable subline of K562 cells (K562-HMGB1) overexpressing HMGB1 protein and K562-HMGB1 sublines served as control, so as to provide a basis for exploring the role of hmgb1 gene in occurrence and development of leukemia and their mechanism. Protein-coding gene of hmgb1 was amplified by PCR with cDNA as template, which was synthesized by reverse transcription from total RNA extracted from U937 cells. The PCR-amplified hmgb1 gene was ligated into PMD18-T vector (PMD18-T-HMGB1 vector), and then transformed into E. coli strain DH5α. DH5α containing PMD18-T-HMGB1 vector were grown on LB agar plate supplemented with 100 µg/ml ampicillin overnight. The single ampicillin-selected DH5α clone was picked for culturing overnight and then harvested for plasmid extraction. The extracted plasmid was characterized to contain hmgb1 gene digested with the desired restriction enzymes of KpnI/XhoI. The correctness of hmgb1 sequence was confirmed with DNA sequencing. The insert of hmgb1 gene contained in PMD18-T-HMGB1 vector was cut out with restriction enzymes of KpnI/XhoI and then ligated into eukaryotic expression vector pcDNA3.1 to form pcDNA3.1-HMGB1 vector. 10µg of pcDNA3.1-HMGB1 or pcDNA3.1 plasmid was separately electroporated into K562 cells. At 48 hours after electroporation the cells were cultured with G418 at a final concentration of 800 µg/ml for over 2 weeks. Finally stably transfected sublines of K562 cells containing hmgb1 gene (K562-HMGB1), and of K562 containing pcDNA3.1 vector (K562-pcDNA3.1) served as a control, were obtained. The transcriptional or translational expression of hmgb1 gene was detected with RT-PCR or Western blot, respectively, to testify transfected efficiency and validity of stable subline of K562-HMGB1. The results indicated that the eukaryotic expression vector pcDNA3.1-HMGB1 plasmid was successfully constructed and was electroporated into K562 cells. The transcriptional or translational expression of hmgb1 gene in the stable subline of K562 cells containing hmgb1 gene was overexpressed. It indicated that stable subline of K562-HMGB1 cells was successfully established. It is concluded that the stable sublines of K562-HMGB1 cells or K562-pcDNA3.1 cells are successfully established, which provides a basis for exploring the roles and mechanisms of hmgb1 gene in leukemogenesis and development of leukemia.

Clinical prognostic multivariate analysis of incidental gallbladder cancer: a case controlled retrospective study / 中华外科杂志

<p><b>OBJECTIVE</b>To investigate the prognostic risk factors in incidental gallbladder cancer (IGBC) and evaluate the effect of laparoscopic cholecystectomy (LC) applied in treating IGBC.</p><p><b>METHODS</b>The retrospective study enrolled 55 patients with incidental gallbladder adenocarcinoma treated between January 2001 and December 2008. The patients were divided into three groups according to different surgical approaches: laparoscope group (n = 23), conversion group (n = 6) and laparotomy group (n = 26). Survival analysis and Cox regression model were applied to comparing the difference of survival rate between three groups and to analyzing the related prognostic risk factors of IGBC.</p><p><b>RESULTS</b>The overall 1-, 3- and 5-year survival rates were 74.3%, 47.7% and 35.8% respectively. And the median survival time was 36 months. The outcome of survival rate between three groups was not different statistically. Cox regression analysis indicated that pathologic T stage was an independent risk factor influencing IGBC (OR = 2.75, P = 0.00). The prognosis was getting worse according to the rising depth of tumor invasion. However, the other factors, such as surgical approach, tumor incisional implantation, ect.were not related to the prognosis (P > 0.05).</p><p><b>CONCLUSION</b>The factor of pathologic T stage is related to the prognosis of IGBC for which LC, compared with open cholecystectomy, should not be regarded as a negative factor in treatment.</p>