Child, Preschool ; Female ; Humans ; Infant ; Male ; Scurvy

OBJECTIVESince glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common hereditary hemolytic erythrocyte enzyme deficiency, most cases have single nucleotide mutations in the coding region, and current test methods for gene mutation have some missed detections, this study aimed to investigate the feasibility of RT-PCR sequencing in the detection of gene mutation in G6PD deficiency.

METHODSAccording to the G6PD/6GPD ratio, 195 children with anemia of unknown cause or who underwent physical examination between August 2013 and July 2014 were classified into G6PD-deficiency group with 130 children (G6PD/6GPD ratio <1.00) and control group with 65 children (G6PD/6GPD ratio≥1.00). The primer design and PCR amplification conditions were optimized, and RT-PCR sequencing was used to analyze the complete coding sequence and verify the genomic DNA sequence in the two groups.

RESULTSIn the G6PD-deficiency group, the detection rate of gene mutation was 100% and 13 missense mutations were detected, including one new mutation. In the control group, no missense mutation was detected in 28 boys; 13 heterozygous missense mutations, 1 homozygous same-sense mutation (C1191T) which had not been reported in China and abroad, and 14 single nucleotide polymorphisms of C1311T were detected in 37 girls. The control group showed a high rate of missed detection of G6PD deficiency (carriers) in the specimens from girls (35%, 13/37).

CONCLUSIONSRT-PCR sequencing has a high detection rate of G6PD gene mutation and a certain value in clinical diagnosis of G6PD deficiency.

Adolescent ; Child ; Child, Preschool ; Female ; Glucosephosphate Dehydrogenase ; genetics ; Glucosephosphate Dehydrogenase Deficiency ; diagnosis ; genetics ; Humans ; Infant ; Male ; Mutation ; Reverse Transcriptase Polymerase Chain Reaction ; methods ; Sequence Analysis, DNA

Air Pollutants ; analysis ; Chromatography, High Pressure Liquid ; methods ; Urea ; analysis ; Workplace

Adult ; Breast Neoplasms ; secondary ; Diagnosis, Differential ; Female ; Humans ; Melanoma ; metabolism ; pathology ; secondary ; Melanoma-Specific Antigens ; metabolism ; Middle Aged ; S100 Proteins ; metabolism ; Sarcoma, Clear Cell ; pathology ; Skin Neoplasms ; pathology

Objective To investigate the regulation expression of leukemia inhibitory factor(LIF)in hu- man and animal osteoarthritic(OA)tissues and its clinical relevance.Methods 1)Thirty-five Japanese rabbits, aged eight months,were used to make models of experimental osteoarthritis.Operations were performed at the right knee and the sham ones at the left knee in each rabbit.Rabbits were sacrificed on the 3,7,14,28,42,56 and 84 days after operation respectively.Cartilage and synovium of the knee were collected to observe histological changes of osteoarthritis at different times;immunohistochemistry analysis was conducted to observe the LIF expression and distribution in the cartilage and synovium of the animals.2)From April 2003 to October 2003,32 samples of human articular tissues(cartilage,subchondral bone and synovium)were obtained in the operational procedures and a good quantity of RNA was isolated using Magnetic Beads.The patients who underwent articular operations donated the samples.In the reverse transcription-polymerase chain reaction(RT-PCR),the mRNA expression of LIF was mea- sured by semi-quantity analysis and the location of LIF protein was determined by enzyme-linked immunosorbent assay (ELISA).Results A slight expression of LIF was seen in normal cartilage but less in synovium.However,the expression of LIF was remarkable in synovial lining cells,superficial and middle layers of cartilage in animal os- teoarthritis.There was a significant difference in expression between the animal osteoarthritis and the control group (P＜0.05 ).In human tissue study,LIF mRNA was expressed to a very low level in normal articular tissues and there was no significant difference(P＞0.05)between different anatomical locations.In moderate degrading sub- chondral bone,LIF mRNA was expressed to its highest level.LIF was expressed to the highest level in seriously degrading cartilage tissues.The results were similar to ELISA testing results.LIF extents varied in different articular tissue sections.Conclusions LIF is an important mediator that can contribute to tbe pathogenesis of OA.The different temporal and spatial distributions of LIF in normal and OA tissues imply that LIF may play some important roles in pathogenesis of OA.

Survivin is a protein that inhibits apoptosis and regulates cell division.The cDNA sequence of survivin was amplified by RT-PCR and sub-cloned into the prokaryotic expression vector pET-21b(+),followed by transformation into E.coli strain BL21(DE3) and induction with IPTG.The recombinant survivin protein fusing with 6?His tag was expressed in E.coli in the form of inclusion body at the expression level over 60% of the total cell protein.Results of Western blotting showed that recombinant survivin reacted specifically with anti-human survivin antibody.After gel filtration,the recombinant protein reached the purity over 95%,which facilitate the study of diagnosing and inhibitor agents targeting survivin.

The common problems for non-special pediatric medical practitioners in the depart-ment of hematology include short-time of rotation, weak foundation of pediatric hematology and low enthusiasm of learning. The entrance education was highlighted in order to make refreshers familiar-ize with the severity of illness and eliminate the medical error. Basic theoretical study was strength-ened aiming at improving clinical skill and enhancing the learning interesting. Method of doctor-patient communication was reformed to reduce medical disputes.

Objective To investigate the clinical value of the probe melting curve analysis‐based PCR assay for the detection of three types of αT .Methods A total of 149 blood and prenatal archival DNA samples (6 of which were amniotic fluid samples)with three knownαT genes ,which included 63 carriers with Hb CS ,22 cases with Hb QS ,43 individuals with Hb WS and 1 double heter‐ozygote with Hb CS and Hb WS) as well as 20 samples with normalα‐globin gene sequence that had been confirmed by RBD com‐bined with DNA sequencing were selected to test the specificity of probe melting curve analysis by blind analysis .Results The probe melting curve analysis accurately detected 100 of the DNA samples previously characterized by S RBD combined with DNA sequencing .Conclusion Probe melting curve analysis‐based PCR assay for the detection ofαT is featured with rapidity and accuracy and can be applied to clinical and prenatal diagnosis .

BACKGROUND:The rabbits were widely used as experimental animal models in the research on etiology and pathological mechanism of steroid-induced osteonecrosis of the femoral head. It is stil a valuable and realistic research topic to improve and to innovate the modeling technology nowadays. OBJECTIVE:To improve the modeling technology on osteonecrosis of the femoral head in rabbits induced by glucocorticoid combined with lipopolysaccharide, with the focus on its reduced mortality and the guaranteed successful rate of modeling. METHODS:A total of 28 New Zealand white rabbits were randomly divided into the control group (n=10) and improvement group (n=18). Models of steroid-induced osteonecrosis of the femoral head were established according to different methods. In the improvement group, rabbits were injected with sodium penicilin (5.0 mg/kg) and amikacin sulfate (1.63×104 U/kg) in the left gluteus muscle. Twenty-four hours later, al rabbits were injected with prednisolone acetate (20 mg/kg) in the right gluteus muscle. Forty-eighthours later, 5.0 μg/kg of lipopolysaccharide was intravenously injectedvia the ear. From then on, two injections of prednisolone acetate (20 mg/kg) were respectively performed alternately in the left and right gluteal muscle at an interval of each 24 hours. Sodium penicilin (5.0 mg/kg) and amikacin sulfate (1.63×104 U/kg) were intraperitonealy injected for 2 consecutiveweeks. In the control group, 10 μg/kg lipopolysaccharide was injectedvia the ear vein of rabbit. From then on, prednisolone acetate (20 mg/kg) was intramuscularly injected at an interval of each 24 hours, totaly three times. Benzylpenicilin sodium 20×104 U/rabbit was intramuscularly injected once a week. RESULTS AND CONCLUSION:Rabbit models of steroid-induced osteonecrosis of the femoral head were successfuly established in both groups. Compared with the control group, the mortality was significantly reduced after model establishment in the improvement group, and the bone lacuna and osteonecrosis of the femoral head were apparent. These findings indicated that the improved technology of model establishment of steroid-induced osteonecrosis of the femoral head could be used to aleviate the damage degree on the gluteal muscles, to guarantee the successful rate of modeling, and to noticeably reduce the mortality of rabbits.