Objective To evaluate the application of high-performance liquid chromatography (HPLC) in diagnosis and screening of thalassemia. Methods Automated HPLC was used to measure HbF and HbA2 in 100 genetically diagnosed thalas-semic patients and 35 normal children. The results were compared with those from traditional tests including alkali denaturation test and cellulose acetate electrophoresis. The diagnose accordance rates, sensitivity and specificity were compared. Results Seventy-fourβthalassemia, 64 were heterozygous with single mutations and 10 were compound heterozygous with double muta-tions. Twenty-sixαthalassemia, 25 were compound mutations and one was heterozygous with single mutation. The HbF percent-age from HPLC was higher than that from alkali denaturation tests in either thalassemia or normal children (P<0.01). HbF level from HPLC inα-thalassemia was signiifcantly different from that in the normal children (P=0.011). The percentage of HbA2 from HPLC was higher than that from cellulose acetate electrophoresis (P=0.010). HbA2 in the single heterozygousβ-thalassemia were twice higher than that in the double heterozygous mutatedβ-thalassemia (P<0.01). The combination of HbF-HbA2 (≥4.0%) from HPLC with MCV (<80 lf) and MCH (<27 pg) had high accordance rates (99.3%), sensitivity (99.0%) and speciifcity (100.0%) in diagnosis of thalassemia. Conclusions When the results of HPLC are combined with MCV and MCH, it can be applied to the diagnosis of thalassemia with high speciifcity, high sensitivity and has high diagnostic accordance rate with genetic results. HPLC can be an ideal approach to screenβthalassemia.

AIM:To investigate whether the protective effect of adiponectin on glucose and lipid metabolism is achieved through down-regulating major histocompatibility complex classⅡ( MHCⅡ) in the adipose tissue.METHODS:Adiponectin knockout ( KO) mice and C57BL/6 ( WT) mice were fed with high-fat diet and standard diet for 24 weeks, re-spectively.The body weight, fasting blood glucose (FBG), fasting insulin (FINS), homeostasis model assessment of insu-lin resistance (HOMA-IR), triglyceride (TG), total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol ( HDL-C) , hepatic histology, and classⅡtrans-activator ( CIITA) , histocompatibility 2 class II antigen E beta (H2-Eb1) and cluster of differentiation 74 (CD74) mRNA and MHC II protein levels in adipose tissue were measured at sacrifice.siRNA targeting MHC II and overexpression vector was used in 3T3-L1 cells to explore the effect of adiponectin on the protein level of MHCⅡ.RESULTS:The levels of body weight, FBG, FINS, HOMA-IR, TC, TG, LDL-C, hepatic steatosis, CIITA, H2-Eb1 and CD74 mRNA expression, and MHCⅡ protein expression in the KO mice were higher than those in the WT mice that fed with high-fat diet or standard diet.In 3T3-L1 cells, inhibition of adiponectin reversed MHC II protein level induced by specific siRNA.The expression of MHC II in adipocytes decreased after adiponectin was overexpressed.CONCLUSION: Adiponectin improves glucose and lipid metabolism through sup-pressing the expression of MHCⅡin the adipose tissue.

Objective To optimize extraction technology of Fufang-Fuqing-Xiji.Methods Orthogonal test combined with Plackett-Burman experimental design was adopted to study the influence factors of water extraction in Fufang-Fuqing-Xiji. The contents of chlorogenic acid, paeoniflorin and caffeic acid in Fufang-Fuqing-Xiji were determined by HPLC gradient elution method at the same time. A multi-index comprehensive score method was applied to analyze the data and to screen the optimum extraction technology. Results The best extraction conditions of Fufang-Fuqing-Xiji were as following: threaded 80 mesh sieve, soaked 2 h, decocted 3 times with 12 times the amount of water for 1h each time.Conclusions This optimized extraction technology was rational and stable, which could be used for the extraction ofFufang-Fuqing-Xiji.

OBJECTIVETo compare the therapeutic effect of Astragalus and Angelica Mixture (AAM) on treating CKD patients according to different CKD primary diseases, staging and TCM syndromes.

METHODSA multicentre, open-label, and self control clinical design was used, and thirty-two patients in line with inclusive criteria were recruited. Based on maintaining their previous basic CKD treatment, patients additionally took AAM (Astragalus and Angelica each 30 g), once a day, three months consisted of one therapeutic course. Serum creatinine (SCr), estimated glomerular filtration rate (eG- FR), 24 h urinary total protein (UTP), plasma albumin (ALB), hemoglobin (Hb), and changes of TCM syndrome factor integrals were compared before treatment, at the end of month 1, 2, and 3. The differences in the aforesaid indices were compared between CKD patients with different CKD primary diseases (chronic glomerulonephritis, chronic renal tubulointerstitial disease, hypertensive renal damage), different CKD stages (CKD 3 and CKD 4), and patients of qi-blood deficiency syndrome (QBDS) and non-QBDS.

RESULTSAAM could improve 78.12% (25/32) patients' renal function. Compared with before treatment, SCr decreased (12.08% +/- 10.11%), eGFR increased (21.14% +/- 18.55%), and ALB increased (2.76% +/- 1.97%) at the end of 3-month treatment (all P < 0.05). As for TCM syndrome factor integrals, compared with before treatment, the integrals for qi deficiency syndrome, blood deficiency syndrome, and yin deficiency syndrome decreased, while the integrals for dampness heat syndrome and turbid-toxin syndrome increased (all P < 0.05). There was no obvious difference in all indices except the integral for hypertensive renal damage patients of yin deficiency syndrome (P > 0.05). The SCr decreasing percent was 19.82% +/- 8.30% for patients of non-QBDS and 5.24% +/- 10.75% for patients of QBDS. The latter was higher with statistical difference (P < 0.05). As for TCM syndrome factor integrals, the integral differences of qi deficiency and blood deficiency were obviously higher in patients of QBDS, when compared with patients of non-QBDS (P < 0.05).

CONCLUSIONAAM could improve the renal function of CKD patients, elevate their ALB levels, and ameliorate associated qi deficiency syndrome, blood deficiency syndrome, and yin deficiency syndrome, especially for CKD patients of QBDS.

Adolescent ; Adult ; Aged ; Angelica ; Astragalus Plant ; Drugs, Chinese Herbal ; therapeutic use ; Female ; Humans ; Male ; Middle Aged ; Phytotherapy ; Renal Insufficiency, Chronic ; drug therapy ; Treatment Outcome ; Yin Deficiency ; drug therapy ; Young Adult

Abstract: Objective　To explore the carrying status of four common deafness genes and mutations on 10 loci in newborns in Hainan, and to analyze the molecular epidemiological characteristics of deafness genes and their loci, so as to provide scientific basis for formulating neonatal deafness gene screening strategy and promoting children's hearing health in Hainan. Methods　Newborns born in Hainan from January 2020 to December 2021 were selected as the research objects. The demographic characteristics of the research objects were collected. At the same time, the plantar blood of newborns was collected, and multiplex PCR amplification and directed hybridization combined with high-throughput sequencing technology were applied to detect 10 mutation loci on 4 common deafness genes. T-test or chi square test was used to process the data. Results　A total of 7 124 newborns were included in the study through informed consent, 219 cases of deafness gene mutation were detected with the detection rate of deafness gene of 3.07%. The detection rates of GJB2, SLC26A4, MT-RNR1 and GJB3 were 1.56% (111/7 124), 1.18% (84/7 124), 0.21% (15/7 124) and 0.11% (8/7 124) respectively. Among the 10 loci of the four genes, the positive detection rate of c.235delC locus of GJB2 was the highest, which was 1.38% (98/7 124), followed by c.919-2A>G of SLC26A4 (0.87%, 62/7 124); 2.63% (113/4 289) of the newborns who passed the preliminary hearing screening still carried the deafness gene; in terms of gene type, the detection rate of GJB2 gene in newborns who failed the hearing screening was higher than that in newborns who passed the hearing screening [2.23% (63/7 124) vs 1.12% (48/7 124)，P<0.01]; in terms of gene loci, the detection rate of c.235delC locus in newborns who failed hearing screening was higher than that in newborns who passed hearing screening [2.09% (59/7 124) vs 0.91% (39/7 124)，P<0.01]. Conclusion　The most common deafness genes types in Hainan were GJB2 and SLC26A4; The most common gene mutation sites were c.235delC and c.919-2A>G; 2.63% of the newborns who passed the preliminary hearing screening still carried the deafness gene, among which the high-risk newborns with MT-RNR1 and GJB3 genes were found. Therefore, hearing screening should be combined with deafness gene screening to improve the detection rate of children at high risk of hearing loss.

Objective To estimate the current quality of the reporting of randomized controlled trials (RCTs) related to digestive diseases in China. Methods All the papers related to RCTs published in Chinese Journal of Digestion from 1999 to 2008 were hand-searched by professional staff then evaluated and analyzed them according to the international reference standard. ResultsIn the 3298 issues of the recent ten years, there were 92 research papers of RCTs which was accounting for 2.8%. The sample size ranged from 18 to 5241. Sixty-one (66.0%) trials included the exact standard of internalize and exclusion. Sixteen (17.4%) trails told the specific method of random allocation and 22(23.9%) were double-blinded. Fifty-eight (63.0%) trials compared the baseline condition of each groups. Seventy-three(79.3%) trails showed the specific approach of statistic. In the end, only 7(5.7%) trails were identified as the strictly-designed RCTs. Conclusions The quantity and quality of the clinical RCTs can not satisfy the demand of clinical practice. Strictly-scientific designed, multicentered, large sample prospective clinical RCT should be advocated.

AIM: To investigate the CPU86017 and its enantiomers inhibit abnormal gene expression of calcineurin and NFκB in rat cardiomyopathy induced by L-thyroxin and compare the effect of CPU86017 (racemate) with its 4 enantiomers: (7S, 13R), (7S, 13S), (7R,13S), and (7R,13R)-CPU86017 in this model. METHODS: The animals were randomly divided into 7 groups. The rat hypertrophied model was produced by treatment with L-thyroxin 0.2 mg·kg-1·d-1, sc for 10 d and treated with CPU86017 or its enantiomers 4 mg·kg-1·d-1, sc from d 6 to d 10. The changes in left ventricular (LV) weight index, redox system, and the NO and iNOS activity in the myocardium were investigated. The expression of mRNA of calcineurin、NF-κB in the left ventricle was measured. RESULTS: There were significant cardiac hypertrophy and oxidative stress in rats treated by L-thyroxin. The expression of calcineurin, NFκB mRNA were upregulated (P<0.05, compared with that of control). After treatment with CPU86017 (racemate and enantiomers), LV remodeling and the redox system were improved. CPU86017 and (7S,13R)-CPU86017 showed a better improvement on LV remodeling and the redox than the other isomers and restored the normal expression of calcineurin, NF-κB (P<0.05, P<0.01), respectively. CONCLUSION: It suggested that an up-regulation of calcineurin and NFκB possibly related to the altered intracellular calcium handling system plays a role in the progression of L-thyroxin induced cardiomyopathy and CPU-86017 and its 7S,13R-CPU86017 enantiomer effectively inhibit the abnormal expression of calcineurin and NFκB genes, the NOS enzyme and oxidant stress in the cardiomyopathy.

Objective To study histone acetylation patterns in peripheral boiood mononuclear cells (PBMC) from type 2 diabetic patients PBMC form 12type 2 diabetic patients and 12 healthy contrcl subiects were collecyed The global histone H3/H4 acetylation was determined by assay kit The differetial expression of the histone acetyltransferases and histone deacetylases were measured by real time PCR.Results increased H3/H4 acetylation was observed in PBMC of type 2 diabetic patients compared with control subjects (H3:0.134±0.035vs0.181±0.032,p<0.05;H4:0.266±0.070VS0.324±0.062,P<0.01).the mRNA levels of p300,CREBBP,HDAC2,HDAC7,and STRTL in patients PBMC compared with healthy control subieects were 3.11±0.38,2.78±0.45,3.55±0.30,0.77±0.37 and 0.40±0.25 folds respectively conclusion histone acetylation appears abnormal in PBMC of type 2 diabetic patients.

Objective To examine the expression and analyze the significance of autophagy-related gene microtubule-associated protein 1 light chain 3 (MAP1-LC3,LC3),p62 and lysosorne-associated membrane protein 2 (LAMP-2) in pancreatic tissues of mice with acute pancreatitis (AP).Methods Twenty mice were randomized into AP group and control group,and the number of mice was equal between two groups.AP group was intra-peritoneally injected by 20％ L-arginine solution (two injections of 4 g/kg body weight,every 1 h) in the dosage of 4 g/l kg twice every 1 hour to establish AP model,while control group was administered with equal volume of normal saline by intra-peritoneal injection.All the mice were euthanized at 24 hour after the last injection.Pancreatic histopathological changes were measured.In addition,the protein expressions of LC3,p62 and LAMP-2 were detected by Western blot.Results No obvious pathological changes were observed in control group.Pancreatic acinar edema,structure destruction,missing,the obvious widening of interlobular septum,small interlobular septum and acinar septum,and the necrosis of acinar cells at different degrees were observed in AP group.The pathological score for tissue edema,hemorrhage,necrosis and inflammation in AP group was 3.13 ± 0.50,2.83 ± 0.32,3.25 ± 0.46 and 3.16 ± 0.47,respectively,which was all 0 in control group.The differences were statistically significant between AP group and control group (P ＜ 0.01).In AP group,the ratio of LC3-Ⅱ/LC3-Ⅰ,p62 and LAMP-2 protein in pancreatic tissue were 1.16 ± 0.08,0.94 ± 0.04 and 0.35 ± 0.04,respectively,which were 0.24 ± 0.02,0.34 ± 0.03 and 0.95 ± 0.03 in control group.The ratio of LC3-Ⅱ/LC3-Ⅰ and p62 protein in pancreatic tissue in AP group were much higher than those in control group,while LAMP-2 in AP group was lower than that in control group,and there was statistically significant difference between two groups (all P ＜0.01).Conclusions Intraperitoneal injection of L-arginine could induce acute pancreatitis,and autophagy is impaired,which was associated with decreased LAMP-2 protein expression.

Objective The purpose of this study was to develop a high-throughput micro-plate radiobinding assay (RBA) of glutamic acid decarboxylase antibody (GAD-Ab) and to evaluate its clinical application. Methods 35labeled GAD65 antigen was incubated with sera for 24 h on a 96-well plate, and then transferred to the Millipore plate coated with protein A, which was washed with 4℃ PBS buffer, and then counted by a liquid scintillation counter. The GAD-Ab results were expressed by WHO standard unit (U/ml). A total of 224 healthy controls, 162 patients with type 1 diabetes mellitus(T1DM) and 210 patients with newly diagnosed type 2 diabetes (T2DM) were recruited. A total of 119 TI DM and healthy cases with gradually changing GAD-Ab levels were selected to compare the consistency of micro-plate RBA with conventional radioligand assay (RLA). Blood samples were obtained from the peripheral vein and finger tip in 32 healthy controls, 35 T1DM and 24 T2DM patients, and tested with micro-plate RBA and then compared with the conventional RLA to investigate the reliability of finger tip sampling. Linear correlation,student's t-test, variance analysis and receiver operating characteristic (ROC) curve were performed using SPSS 11.5. Results (1) The optimized conditions of micro-plate RBA included 2 μl serum incubated with3 ×104 counts/min 35S-GAD for 24 h under slow vibration, antigen-antibody compounds washed 10 times by 4℃ PBS buffer, and radioactivity counted with Optiphase Supermix scintillation liquid. (2)The intra-batch CV of the micro-plate RBA was 3.8%- 10.2%, and the inter-batch CV was 5.6%- 11.9%. The linearity analysis showed a good correlation when the GAD-Ab in serum samples ranged from 40.3 to 664 U/ml and the detection limit of measurement was 3.6 U/ml. The results from Diabetes Autoantibody Standardization Program (DASP) 2005 showed that the sensitivity and specificity for GAD-Ab were 78% (39 positive among 50 new-onset T1DM) and 98% (2 positive among 100 healthy controls). The results of GAD-Ab obtained with micro-plate RBA and RLA were closely correlated (r=0.915,P<0.001) with a high concordance level of 97.5% and a Kappa value of 0.95. (3)TI DM and T2DM patients showed higher positive rates for GAD-Ab than the healthy controls(46.9% and 5.2% vs 0.89% ,X2=123.5 and 10. 1 ,P <0.001 and <0.01, respectively). (4)The consistency of GAD-Ab measurement with RBA using finger tip blood and RLA measurement using venous blood was 96.7% (r =0.946,P <0.001, Kappa value: 0.905). Conclusions The micro-plate RBA of GAD-Ab has high sensitivity, specificity and reproducibility, and can be measured with finger tip blood sampling. It might be a better alternative for clinical practice.