Establishment of a PCR-SSP method for the simultaneous amplification and identification of the presence of KIR genes / 中华医学遗传学杂志

OBJECTIVE@#To develop a polymerase chain reaction-sequence specific primer (PCR-SSP) method for simultaneous amplification and identification of the KIR genes among Chinese population.@*METHODS@#Peripheral blood samples from 132 healthy donors who had given blood at Shenzhen Blood Center from January 2015 to November 2015 were selected as the study subjects. Based on the polymorphism and single nucleotide polymorphism (SNP) information of high-resolution KIR alleles in the Chinese population and the IPD-KIR database, specific primers were designed to amplify all the 16 KIR genes and the 2DS4-Normal and 2DS4-Deleted subtypes. The specificity of each pair of PCR primers was verified by using samples with known KIR genotypes. During PCR amplification of the KIR gene, co-amplification the fragment of human growth hormone (HGH) gene by multiplex PCR was used as the internal control to prevent false negative results. A total of 132 samples with known KIR genotypes were randomly selected for blind inspection to verify the reliability of the developed method.@*RESULTS@#The designed primers can specifically amplify the corresponding KIR genes, with clear and bright bands for the internal control and KIR genes. The results of detection are fully consistent with the known results.@*CONCLUSION@#The KIR PCR-SSP method established in this study can yield accurate results for the identification of the presence of KIR genes.

Concordance of identifying the presence or absence of KIR genes by Flow-rSSO and PCR-SBT methods: a comparative study / 中国输血杂志

【Objective】 To study the concordance of identifying the presence or absence of KIR genes using flow reverse sequence-specific oligonucleotide probe (Flow-rSSO) hybridization and sequencing based typing-PCR (PCR-SBT) methods. 【Methods】 A total number of 131 cases of DNA samples from Han population were subjected to identify the presence or absence of all 16 KIR genes by Flow-rSSO method, and then sequenced at coding sequence for all 14 functional KIR genes using our in-house KIR PCR-SBT assay. The concordance of identifying the presence or absence of all functional KIR genes by Flow-rSSO and PCR-SBT was analyzed. Samples with inconsistent initial results were re-tested using the Flow-rSSO commercial kits with different Lot number, and further tested using the PCR-SSP commercial kit. 【Results】 The presence or absence of 14 functional KIR genes for 129 of 131 samples were completely in accordance via the PCR-SBT and Flow-rSSO methods. Two samples, one with 3DL1 negative, the other with both 2DS3 and 2DS5 negative initially-identified by Flow-rSSO, were actually all positive tested by PCR-SBT. Further retest by Flow-rSSO commercial kits with different Lot number and PCR-SSP commercial kit indicated that the two samples were all positive, which agreed well with PCR-SBT results. 【Conclusion】 In this paper, the initial test results of the presence or absence of KIR genes identified by Flow-rSSO for 2 samples were wrong, which indicated the importance of carrying out the quality control for reagents in KIR gene testing.

Association of molecular genetic polymorphism of KIR-HLA systems with acute leukemia in Southern Chinese Han / 中华医学遗传学杂志

OBJECTIVE@#To investigate the association of molecular genetic polymorphism of KIR-HLA systems with acute lymphoblastic leukemia (ALL) and acute myelocytic leukemia (AML) in southern Chinese Han.@*METHODS@#A total number of 323 cases of adult ALL patients, 350 adult AML, and 745 random healthy controls were tested by KIR PCR-SSP and HLA-A, -B, -C sequence-based typing (PCR-SBT) methods. The molecular genetic polymorphisms of KIR genes and KIR gene profiles, classⅠ HLA ligands, and KIR receptor +HLA ligand combinations were compared between patient and healthy control groups.@*RESULTS@#A total number of 32 and 33 different kinds of KIR profiles were identified in the ALL and AML patient groups. Compared with the observed frequencies of KIR profiles in healthy controls, the observed frequencies of KIR profile AA1 were significantly lower in both the ALL and AML groups (ALL group: 45.79% vs. 55.30%, Pc=0.004; AML group: 48.27% vs. 55.30%, Pc=0.030). In the ALL group, the observed frequencies of 2DL2 gene and 2DL2+HLA-C1 combination, 2DS2 gene and 2DS2+HLA-C1 combination were significantly higher than those in healthy controls (P<0.05), whereas the frequencies of 2DL3 gene, HLA-A3/A11 ligand and 3DL2+HLA-A3/A11 combination were significantly lower than those in healthy controls. However, no significant differences remained after Bonferroni correction (Pc>0.05). In AML group, the observed frequencies of both 2DS1 and 2DL5 genes were significantly higher than that in healthy controls, whereas the frequencies of HLA-C2 ligand and 2DL1+HLA-C2 combination were significantly lower than that in healthy controls(P<0.05). However, no significant difference existed after Bonferroni correction (Pc>0.05).@*CONCLUSION@#This study revealed some potential susceptibility or protective factors related to acute leukemia in southern Chinese Han, especially the protective factor KIR profile AA1, which might provide new clues and theoretical basis for the pathogenesis of acute leukemia and individualized immunotherapy.

Polymorphism of KIR2DL4 gene among northern Chinese Han population / 中国输血杂志

【Objective】 To investigate the polymorphism of KIR2DL4 gene in northern Chinese Han population. 【Methods】 A total of 327 DNA samples were isolated by magnetic beads from unrelated individuals of northern Chinese Han population. The coding sequence (CDS) of KIR2DL4 were amplified using four pairs of KIR2DL4-specific PCR primers developed by our own KIR sequencing-based typing patent, and each exon of KIR2DL4 carried by the four PCR amplicons was then subjected to DNA Sanger sequencing in both directions. The genotype of each sample was assigned using the Assign 4.7 software. 【Results】 Among 327 individuals, 8 different kinds of KIR2DL4 alleles were detected, with observed frequencies ranked as KIR2DL4*00102 [77.1%(252/327)], *00501 [35.8%(117/327)], *011 [20.2%(66/327)], *00602 [12.5%(41/327)], *00801 [8.6%(28/327)], *00103 [4.9%(16/327)], *00503 [1.5%(5/327)] and *00504 [0.9%(3/327)]. In this study, the 10A type alleles which can encode a full membrane-bound receptor include 2DL4*00102, *00103, *00501, *00503, *00504 and *00602, whereas the 9A type alleles which produce truncated forms of receptor include 2DL4*00801 and *011. The observed frequencies for 10A and 9A type KIR2DL4 alleles were 97.6% (319/327) and 27.8% (91/327), respectively. The ratio of 10A to 9A type was 3.5: 1. The observed frequencies of KIR2DL4 alleles in northern Chinese Han population showed no significant difference compared with southern Chinese Han population (P>0.05). 【Conclusion】 The allelic diversity of KIR2DL4 elucidated in the present study can provide valuable data for KIR-associated disease studies and human evolution.

Epidemiological investigation of allergic rhinitis in Ningxia / 中华耳鼻咽喉头颈外科杂志

Objective@#To investigate the epidemiological characteristics of allergic rhinitis (AR) in Ningxia and to analyze its related factors.@*Methods@#From March to September of 2013, a multi-stage and cluster sampling method was used to investigate the diagnosis and treatment of AR in Ningxia Area (3 years and above). Guidelines for diagnosis and treatment of allergic rhinitis (2009, Wuyishan) was used as the basis for the diagnosis of adult AR, while Guidelines for diagnosis and treatment of pediatric allergic rhinitis (2010, Chongqing) was used as the basis for children. SPSS 16.0 software was used to complete the statistical analysis.@*Results@#The total number of questionnaires was 6 000, and the number of effective questionnaire was 5 236, the recovery rate was 87.27%. With 684 cases diagnosed of AR, the prevalence of AR in Ningxia was 13.06% (684/5 236), including 13.40% (325/2 425) of males, 12.77% (359/2 811) of females. The difference was not statistically significant (χ2=0.456, P>0.05). There was significant difference in the prevalence between Hui and Han [14.35% (452/3 150) vs 11.12% (232/2 086), χ2=11.51, P<0.05]. According to ARIA criteria, persistent AR was 27.63% (189/684), intermittent AR was 72.37% (495/684). The month with highest incidence of AR in Ningxia Area was September, accounting for 71.78% (491/684). The prevalence of urban population was 14.54%, with the prevalence of rural population was 11.90%, and the difference was significant between urban and rural residents (χ2=7.90, P<0.05). The age group with highest prevalence rate was 21~30 years old. The main inhalation allergens were mugwort (68.42%), weeds (58.48%) and ragweed (55.56%). The main dietary allergens were wheat flour (14.33%), peanut (13.74%) and walnut kernel (11.99%). The most common complication was allergic conjunctivitis [82.02% (561/684)].@*Conclusion@#The epidemiology of AR in Ningxia Area is preliminarily understood, which will provide the epidemiological evidence for the prevention and treatment of AR and the formulation of public health policy.

The correlation of allergic rhinitis and trace elements in Ningxia region / 中华耳鼻咽喉头颈外科杂志

<p><b>OBJECTIVE</b>To investigate the correlation of allergic rhinitis and trace elements and to provide a reference for clinical diagnosis, treatment and prevention of allergic rhinitis.</p><p><b>METHODS</b>One hundred and six patients were diagnosed as allergic rhinitis in General Hospital of Ningxia Medical University between January and December in 2010, including 48 cases of perennial allergic rhinitis and 58 cases of seasonal allergic rhinitis. In the same time, one hundred and three healthy volunteers were selected as control. Intravenous blood 3-5 ml were obtained from all subjects both in experimental group and in control group. The content of Ca, Ni, Fe, Mg, Zn, Sr, Mn, Cu, Se in serum and hair was determined by inductively coupled plasma atomic emission spectrometry (ICP-AES). The t-test (SPSS 16.0) was used to compare the results of trace elements in serum between allergic rhinitis and control group.</p><p><b>RESULTS</b>The testing results of trace elements in AR patients serum and normal controls serum were as follows: Cu, Ni (1 002.18 ± 104.62) µg/L, (21.58 ± 5.54) mg/L were super than control group, (832.78 ± 50.45) µg/L, (17.04 ± 4.93) mg/L (t value was 15.545, 5.154, both P < 0.05). But the content of Zn, Se (793.48 ± 46.88) µg/L, (84.25 ± 12.77) µg/L lower than control group (908.53 ± 31.26) µg/L, (98.35 ± 15.21) µg/L (t value was -24.175 and -7.797, both P < 0.05) . The testing results of trace elements in AR patients hair and normal controls hair were as follows: Cu, Ni (42.43 ± 5.03) µg/g, (31.72 ± 5.49) µg/g were super than control group, (23.00 ± 4.45) µg/g, (8.94 ± 7.53) µg/g (t value was -8.633 and 4.236, both P < 0.05). But the content of Zn, Se (92.16 ± 4.54) µg/g , (0.28 ± 0.04) µg/g lower than control group (189.09 ± 8.45) µg/g, (0.39 ± 0.06) µg/g (t value was -28.71 and -8.633, both P < 0.05).</p><p><b>CONCLUSIONS</b>The content of Zn, Se in AR patients serum are lower than that in control group. But the content of Cu, Ni in AR patients serum are super than that in control group. There are no significant difference of trace elements in the serum between pennial allergic rhinitis and seasonal allergic rhinitis.</p>

Expression and implication of CD44 and nm23-H1 proteins in laryngeal carcinoma / 临床耳鼻咽喉头颈外科杂志

OBJECTIVE@#To study the expression of CD44 and nm23-H1 gene proteins and their clinical significance in laryngeal carcinoma.@*METHOD@#The expression of CD44 and nm23-H1 proteins were detected by immunohistochemistry method in 40 cases with laryngeal carcinoma, 20 adjacent carcinoma tissues and 12 cases normal laryngeal mucosa tissues.@*RESULT@#The expression of CD44 and nm23-H1 proteins in laryngeal carcinoma were much higher than that in normal laryngeal mucosa. The expression of CD44 protein in laryngeal carcinoma with metastatic lymph node was higher than that in laryngeal carcinoma without metastatic lymph node, but nm23-H1 protein lower. The expression of CD44 protein was positively correlated with the metastasis, clinical staging and pathological classification but not correlated with T classification of laryngeal carcinoma. The expression of nm23-H1 protein was negative correlation with the metastasis and clinical staging of laryngeal carcinoma.@*CONCLUSION@#CD44 and nm23-H1 gene proteins play an important coordinated regulation role in the carcinogenesis, development and metastasis of laryngeal carcinoma and will probably become the key biological marks in the judging and evaluating prognosis of laryngeal carcinoma.