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In this study, aiming at the characteristics of Abelmoschus manihot (L.) bast lignin content is high and degumming difficulty, it introduces a new type of degumming methods. Microwave-assisted treatment was used in the degumming process of Abelmoschus manihot (L.) Medik. The effect of NaOH concentration, microwave-assisted treatment time and temperature on the residual gum content in Abelmoschus manihot (L.) Medik fibers were investigated. The result showed that the optimal conditions by microwave-assisted treatment were 50min of processing time,10g/L of NaOH concentration and 95°C of temperature when the residual gum content could reach to 0.96%. Compared with the traditional process, this method significantly shortened the time from 90-240min to 50min. Developed a highly efficient, low energy consumption degumming process, and the Abelmoschus manihot (L.) fiber after the degumming process was be applied to the development and utilization of textile products.

This study was purposed to investigate the clinical value of HLA matching(low and high resolution) and its effect on outcome of the patients received umbilical cord blood transplantation(UCBT). Sequence-specific oligonucleotide probe (SSOP) , sequence-based typing (SBT) and sequence-specific primers(SSP) were used to perform high resolution HLA matching for HLA-A, -B, -Cw, -DRB1, -DQB1 and low resolution for HLA-A, B, DRB1 among 34 patients with hematologic malignancies who received unrelated UCB transplantation and grafts. The effects of HLA matching (low or high resolution ) on leading engraftment, hematopoietic reconstitution, graft-versus-host disease (GVHD) and infection after UCB transplantation were analyzed by comparison. The results showed that the median of total nucleated cells (TNC) of transplanted cord blood was 6.0×10(7)/kg, The time of neutrophil recovery was significantly shortened when more than 5×10(7)/kg TNC were transplanted (P0.05). The HLA-(6-10)/10 group of high resolution HLA matching was better than the HLA (4-5)/10 group in the respect of leading engraftment, the time of platelet recovery and the rate of acute GVHD (P0.05). In contrast, HLA-I+II locus, HLA-DRB1 or HLA-DQB1 locus mismatch could prolong the platelet engraftment time (P0.05). There was statistical difference in the time of platelet recovery, the rate of acute GVHD between the HLA (5-6)/6 group of low resolution HLA matching and the HLA (3-4)/6 group after UCB transplantation (P0.05), but the mismatch locus of HLA with low resolution did not correlate with the time of platelet recovery (P0.05). It is concluded that the high resolution HLA matching between patients received unrelated UCB transplantation and grafts may contribute to select the better UCB, that has important clinical value to promote hematopoietic reconstitution and to reduce the complications after UCB transplantation.

To find out the distributed characteristics of KIR2DL1 alleles frequencies and the recognition HLA-C ligand in the Chinese Han population.The 111 patients and 116 donors from CMDP were performed the KIR2DL1 high-resolution typing and KIR genotyping using sequence-based testing (SBT) and PCR-SSP methods.A total of 224 individuals with KIR2DL1 locus was predominantly observed and accounted for 98.68% (224/227). There were 3 different KIR2DL1 alleles, including KIR2DL1*00302, *00201 and *00401 alleles polymorphism. The most common phenotype observed were KIR2DL1*00302 (84.82%, 380/448), KIR2DL1*00201 (12.05%, 54/448) and KIR2DL1*00401(3.13%,14/448), present at allele genotype frequencies of 61.04%,6.22% and 1.58% respectively. The allele homozygotic types of KIR2DL1*00302 and KIR2DL1*00302 were the most frequent in 6 KIR2DL1 allele by high resolution typing. The allele heterozygous types of KIR2DL1*00302 and KIR2DL1*00401 presented statistically different in haplotypes A/A and B/x (P=0.001), and KIR2DL1*00401 lacked of all A/A haplotype. The KIR2DL1*00302 and KIR2DL1*00201 allele had significant positive associations between different KIR pairs of KIR2DS1, KIR2DL3, KIR2DS4 and KIR3DL1/S1 using linkage disequilibrium analysis (P0.01), respectively. In the receptor-ligand of KIR/HLA model after allo-HSCT, KIR2DL1*00302 alleles correlated with their HLA-C2 group ligands. KIR2DL1*00302 and HLA-C*06:02 was the most common combination ligand model, but KIR2DL1*00302 and HLA-C*01:02 was the most frequent mismatch ligand model with the development of NK cell-induced alloreactivity, meanwhile there was statistically significant difference of frequency distribution (P0.05).The KIR2DL1*00302 was the most frequent allele in Chinese Han population. The KIR2DL1 high resolution typing would be beneficial for predicting donor NK cells all activity after hematopoietic stem cell transplantation and selecting suitable donors.

To explore the influence of the killer cell immunoglobulin like receptor (KIR) gene polymorphism on cytomegalovirus (CMV) infection and pathogenesis after hematopoietic stem cell transplantation (HSCT).The KIR genotype was determined by sequence-specific primer polymerase chain reaction (PCR-SSP) in 138 pairs of donors and recipients before HSCT during October, 2005 and May, 2011. Posttransplant monitoring for CMVpp65 antigen was performed by indirect immune histochemically assays since week 2 after transplantation. The differences between CMV positive group and negative group, inhibitive and active KIR of donors and recipients, and KIR haplotype frequency of donors and recipients were analyzed.There were no significant differences in frequency of KIR gene and haplotype AA, AB, BB between the donors and recipients. The frequencies of 2DS2 and 2DS4 * 003-007 of donors in CMV positive group were obviously lower than those in CMV negative group with significant differences (8% vs 16% , P = 0.0420; 3% vs 13%, P = 0.0050). There was no significant difference in KIR gene between CMV positive group and CMV negative group. The CMV infection rates of haplotype AA, BB, AB donors were 64.38%, 36.84% and 50.00%, while CMV infection rates of haplotype AA, BB, AB recipients were 53.73%, 46.15% and 51.72%, respectively. The CMV infection rate was higher in the patients received KIR haplotype AA donor than in those received KIR haplotype BB donor (36.84% vs 64.38%, P = 0.0299). 2DS4 x 003-007 and haplotype BB of donor were found associated with CMV infection in multifactor analysis.KIR genotypes of donors are associated with CMV infection after HSCT.

To study the impact of various human leukocyte antigen (HLA) high resolution typing mismatching of donor-recipient pairs on prognosis of unrelated donor hematopoietic stem cell transplantation.835 donor-recipient pairs of CMDP data from 2005 to 2010 were analyzed retrospectively. HLA-A, B, C, DRB1 and DQB1 typing were performed using SBT, SSOP and SSP methods. The diseases involved in acute myeloid leukemia (AML) (n = 288), acute lymphoid leukemia (ALL) (n = 227), chronic myeloid leukemia (CML) (n = 187), myelodysplastic syndrome (MDS) (n = 52), non-hodgkin's lymphoma(NHL) (n = 25), aplastic anemia(AA) (n = 42) and thalassemia (n = 14). Of 835 donor-recipient pairs, 362 were completely matched, 159 had a mismatch for a single allele, 125 had a mismatch for a single antigen, 95 had mismatched for both single allele and single antigen, 29 were mismatched at double allele, 20 at double antigen, 45 at multiple allele and antigen. The follow-up assessment was completed before March 2011.HLA-matched pairs had higher overall survival (OS) than HLA-mismatched pairs (79.83% vs 73.15%), but there was no statistically significant differences (P0.05). HLA mismatch for a single allele plus a single antigen was a significantly risk factor for OS, disease free survival (DFS) and transplant-related mortality (TRM). The OS from high to low in different diseases were thalassemia, AA, CML, MDS, AML, NHL, and ALL. OS of HLA locus mismatch were DRB1 (94.4%), DQB1 (83.3%), B (75%), A (74.4%) and C (71.4%), respectively. OS of single allele mismatch at HLA locus from high to low were DRB1, C, A, B and DQB1.HLA-A, B, C locus mismatch were statistically significantly associated with lower OS and grade II-IV acute GVHD compared with HLA-matched pairs (P0.05). The donor-recipient pairs with HLA-B*15:01/B*15:05, DRB1*12:01/DRB1*12:02, C*04:01/C*03:04, DQB1*03:02/DQB1*03:03 alleles mismatch were given priority. But the donor-recipient pairs with HLA-B*39:01/B*39:05, C*15:02/C*14:02, C*08:01/C*03:04, C*07:02/C*15:02 alleles mismatch were risk factors for influence of OS and aGVHD.The high resolution typing for HLA-A, B, C, DRB1, DQB1 can be identified nonpermissive mismatch, which is beneficial for the selection of a suitable donor improves survival on unrelated donor HSCT.

To analyze the allele frequencies and polymorphism of human leukocyte antigens (HLA) -A, B, Cw, DRB1 and DQB1 between donors-recipients on high-resolution typing; and to analyze the matching and mismatching proportion between donors and recipients.HLA high-resolution types were determined by sequence based typing (SBT), sequence specific oligonucleotide probe (SSOP) and sequence specific primer (SSP) on 2540 unrelated Chinese Han individuals including 1168 recipients and 1372 donors, then statistical analyses were carried out.Forty-four HLA-A alleles were detected, and among them the frequencies of A*1101, A*2402, A*0201, A*0207, A*3303, A*0206 and A*3001 exceeded 0.05, and accounted for 80.4%. Eighty-one HLA-B alleles were detected, and the frequencies of B*4001, B*4601, B*5801, B*1302 and B*5101 exceeded 0.05, and accounted for 43.0% of total. There were 44 HLA-Cw alleles, among them the frequencies of Cw*0702, Cw*0102, Cw*0304, Cw*0801, Cw*0602, Cw*0303, Cw*0302 and Cw*0401 exceeded 0.05, and were 80.3% of total. There were 61 HLA-DRB1 alleles, the frequencies of DRB1*0901, DRB1*1501, DRB1*1202, DRB1*0803, DRB1*0701, DRB1*0405, DRB1*0301 and DRB1*1101 exceeded 0.05, and were 70.1% of total. Finally, 22 HLA-DQB1 alleles were detected, the frequencies of DQB1*0301, DQB1*0303, DQB1*0601, DQB1*0602, DQB1*0202, DQB1*0302, DQB1*0401, DQB1*0502 and DQB1*0201 exceeded 0.05, and they were 87.4% of total. All the five loci were of heterozygote deficiency. The HLA-A, B and DRB1 loci conformed to Hardy-Weinberg equilibrium (HWE) (P0.05); but HLA-Cw and HLA-DQB1 loci did not (P0.05). Except several particular genotypes, all the five loci conformed to HWE. After comparing data between donors and recipients, only 22.4% of recipients found HLA matched donors (10/10); 24.6% of recipients found single HLA allele mismatched donors (9/10); 26.3% of recipients had two HLA alleles mismatched donors (8/10).The characteristics of allele frequencies and polymorphism of HLA-A, B, Cw, DRB1 and DQB1 on high-resolution typing in Chinese Han population is valuable for donor searching in unrelated hematopoietic stem cell transplantation, and it provides genetic basis for donor registry and usage of donor resource for Chinese Marrow Donor Program.

To detect the frequencies of KIR2DS4 alleles in Chinese Han population and to study the impact of KR2DS4 alleles on clinical outcomes of HLA identical unrelated allo-HSCT.A Sequence-Based Testing (SBT) and TOPO TA cloning system for identifying and distinguishing alleles of the KIR2DS4 gene were established. A total of 150 Chinese-Han individuals, including 75 leukemia patients who received allo-HSCT and their HLA high-resolution typing identical unrelated donors (URD) were entered this study. The patients underwent transplantation for CML (n = 24), AML (n = 19), ALL (n = 29) and other malignancies (n = 3).The majority (139) of the 150 samples (92.7%) were positive for KIR2DS4. Sequencing of the whole length coding region of this gene identified four of the 12 known KIR2DS4 alleles, KIR2DS4*00101, *003, *004, and *007. 2DS4*00101 was the most frequent, being found in 109 of the 139 individuals (78.4%). The ratio of deleted to non-deleted versions of KIR2DS4 was approximately 1:2. Three novel KIR2DS4 alleles were identified. Transplantations from KIR haplotype B/x donors showed significantly higher overall survival rates than those from KIR haplotype A/A donors [RR 3.1 (95%CI 1.1 - 8.6), P = 0.007]. There was a lower overall survival rates in recipients when their donors carried two 2DS4 full-length allele (2DS4*001) than those carried less (0 or 1) 2DS4*001 allele (P = 0.031). In the haplotype A/A group, a higher risk of acute GVHD (aGVHD) [RR 9.0 (95%CI 1.2 - 66.9), P = 0.01], especially grade III-IV aGVHD (P = 0.006), was seen when the donor was homozygous for the full-length KIR2DS4*00101 allele.The development and application of the described SBT 2DS4 allele typing method highlights the diverse nature of the KIR gene family and displays the existence of KIR polymorphism that remains uncharacterized. Our findings suggest that KIR typing for 2DS4 be beneficial for selecting suitable donors.

To explore the relationship between CMV reactivation and KIR haplotype or HLA-Cw genotype in patients after unrelated-donor hematopoietic stem cell transplantation (HSCT).From January 2003 to December 2008 the HLA-Cw/KIR genotype of 48 patient-donor pairs were determined by polymerase chain reaction with sequence specific primers (PCR-SSP) and sequence specific nucleotide (PCR-SSOP). Posttransplant CMV reactivation was performed by immune histochemically assay.Of 48 patients, 15 were transplanted from unrelated donors with an antigen mismatch for HLA Cw and 33 patient-donor pairs were matched for HLA-Cw. The CMV reaction rate was 66.7% for HLA-Cw mismatch group and 48.5% for HLA-Cw match group (chi(2) = 1.39, P = 0.2375). Thirty-seven donor-patients pairs belonged to group C1 and 11 to group C2, and CMV reaction rate was 64.9% in group C1 and 18.2% in group C2 (chi(2) = 18.13, P0.0001). Twenty-six patients received graft from KIR haplotype A (group A donor) and 22 from KIR haplotype B donors (group B donor) and CMV reaction rate was 57.7% in group A donor and 50.0% in group B donor (chi(2) = 0.28, P = 0.5941). The number of donor activating KIRs (aKIRs) was less than that of recipient aKIRs in 34 patient-donor pairs in which the CMV reaction rate was 70.6%, and the number of donor aKIRs was more than that of recipient aKIRs in 14 patient-donor pairs in which the CMV reactivation was 14.3%. There was a significan difference between the two group (chi(2) = 12.44, P = 0.0004).KIR and HLA-Cw genotypes influence the rate of CMV reactivation following non-T cell deleted unrelated donor hematopoietic cell transplantation.

To establish a real-time quantitative reverse transcriptase polymerase chain reaction (RQ-RT-PCR) for quantitative detection of the common molecular markers that have affirmative clinical significance in the acute and chronic leukemia patients, and evaluate its significance in diagnosing leukemias and monitoring minimal residual disease (MRD).Primers and TaqMan probes were designed for detecting various fusion transcripts and normal abl gene was used as the internal control. The expression level of fusion transcripts in 202 newly diagnosed leukemias were determined.In absolute quantity, expression level of the fusion transcripts in various leukemias was b3a2 (b2a2) 47614.63, e1a2 98847.53, AML1-ETO 300029.51, PML-RAR alpha 25506.28, respectively, while in relative quantity to abl, the levels were 1.05, 0.91, 5.33 and 0.55, respectively.The relative quantification of gene expression level by using RQ-RT-PCR to abl control gene is more accurate and direct viewing. Different levels of transcription of corresponding fusion genes are found in various subtypes of leukemias at diagnosis, among which the level of AML1-ETO was higher and PML-RAR alpha lower.

To study the biological function of killer cell immunoglobulin-like receptor (KIR) and the role of donor inhibitory KIR and recipient genetic background in HLA matched unrelated hematopoietic stem cell transplantation (HSCT).HLA genotype of 51 patients (ALL 18 cases, CML 15 cases, AML 10 cases and others 8 cases) and their respective matched unrelated donors from Database of China Marrow Registration was determined by polymerase chain reaction sequence oligonucleotide probes (PCR-SSOP) and sequence specific primers (PCR-SSP). The KIR genotype was determined by PCR-SSP.All the patients and the donors expressed KIR2DL1, KIR2DL2/L3, KIR2DL4, KIR3DL2 and KIR3DL3. 96.7% individuals expressed KIR3DL1. Among them, 21.57% of KIR was completely identical, while 78.43% was not. Of the non-identical KIRs, 25.49% were the recipient's KIR genotype containing the donor's ones, and 27.45% was the donor's containing the recipient's. 74.62% of donor's KIR2DL1 lacked recipient's C2 ligand, 5.91% of donor's KIR2DL2/L3 lacked recipient's C1 ligand, 19.74% of donor's KIR3DL1 lacked recipient's Bw4 ligand and 54.91% of donor's KIR3DL2 lacked recipient's A3, A11 ligand.KIR genotype and HLA class I antigen are inherited independently. KIR2DLI and KIR3DL2 of donors may cause alloreactivity of NK cell. The mismatch of KIR/HLA in donor-recipient plays a very important role in matched unrelated allo-HSCT. The outcome of HSCT can be better predicted by the model of the presence of KIRs on the donor' sNK cells and the absence of corresponding KIR ligand in the recipient's HLA.

To investigate the expression of AML1/ETO9a isoform in the acute myeloid leukemia (AML)-M2 patients.Expressions of AML1/ETO fusion gene and AML1/ETO9a isoform were detected by using reverse transcriptase-polymerase chain reaction (RT-PCR) in leukemia patients, MDS patients, leukemia cell lines and healthy subjects. Karyotype was studied by R-banding technique.In 30 newly diagnosed AML-M2 patients 15 were found to express AML1/ETO9a isoform, while the rest including 20 AML-M2CR, 18 other subtypes of AML, 5 chronic myelogenous leukemia (CML), 3 myelodysplastic syndromes (MDS), 3 leukemia cell lines (NB4, KG-1, K562) and 5 healthy subjects were AML1/ETO9a negative. Among the 15 AML/ETO9a isoform expressing cases, 13 were demonstrated t(8;21) translocation and AML1/ETO expression.Isoform AML1/ETO9a was correlated to AML/M2, and it may promote the development of leukemia in combination with the AML1/ETO fusion gene.