According to Chinese medicine, the atlantoaxial joint is a composite joint composed of tendons and bones, and the stability of the joint depends on the 'tendon-bone balance' involving tendons, ligaments, atlas and axis. Multiple causes of 'tendon off-position, joint subluxation' will lead to joint 'tendon-bone imbalance', which will evolve into atlantoaxial subluxation (AAS), endangering human health. Chinese therapeutic massage (tuina) is a very effective treatment for AAS in adults, but conventional manipulations are prone to ineffectiveness or accidents due to neglect of the causal relationship of the 'tendon-bone imbalance' and inappropriate manipulations. Compared with conventional manipulations, the rational choice of modified manipulations under the guidance of 'tendon-bone balance' theory is more effective and less risky, and more worthy of clinical promotion. From the 'tendon-bone balance' theory, we considered the shortcomings of conventional manipulations, and introduced several modified manipulations that have their own strengths in 'tendon smoothing' and 'bone setting', in order to provide new ideas for treatment of AAS in adults.

This study was aimed to construct model cell line of NPM1-RNAi in HL-60 cells and its resistant line (HL-60/ADR) so as to provide a experimental basis for investigating the potential role of NPM1 gene in leukemia drug resistance. The shRNA targeting to NPM1 was ligated into linear pGCSIL-GFP vector, and transformed into E.coli DH5α. Positive clone was identified by PCR and DNA sequencing. pHelper 1.0, pHelper 2.0 and pGCSIL-GFP-NPM1-shRNA were cotransformed into 293T cells by lentivirus vector system. NPM1-RNAi-LV was transfected into HL-60 and HL-60/ADR cell lines. The efficiency of NPM-RNAi-LV was detected by using real-time quantitative RT-PCR and Western blot. The results showed that the recombinant eukaryotic expression vector pGCSIL-GFP-NPM1-shRNA was constructed. pGCSIL-GFP-NPM1-shRNA was packed into NPM1-RNAi-LV by lentivirus vector system, and transfected into HL-60 and HL-60/ADR cell lines. At mRNA level, the efficiency of NPM1 mRNA knockdown was more than 90% (p < 0.05). At protein level, obvious down-regulation of NPM protein was noted, indicating that NPM1 gene in HL-60 and HL-60/ADR cell lines was knocked down after transfected with NPM1-RNAi-LV. The resistance of HL-60/ADR cell line to adriamycin decreased to a certain degree after NPM1 gene silencing. It is concluded that the model cell lines of NPM1-RNAi in HL-60 and HL-60/ADR are successfully constructed, which can be used for investigating the potential role of NPM1 gene in drug resistance of leukemia.
Cell Line, Tumor ; Doxorubicin ; pharmacology ; Drug Resistance, Neoplasm ; genetics ; Genetic Vectors ; HL-60 Cells ; Humans ; Lentivirus ; genetics ; Nuclear Proteins ; genetics ; RNA Interference

Objective:To investigate the computed tomography (CT) features of gastric neuroendocrine neoplasm (GNEN).Methods:The retrospective and descriptive method was conducted. The clinicopathological data of 30 GNEN patients who were admitted to two domestic medical centers (13 cases in Wenzhou Hospital of Traditional Chinese Medicine and 17 cases in Wenzhou People′s Hospital) from January 2010 to December 2018 were collected. There were 23 males and 7 females, aged (62±4)years, with a range from 27 to 78 years. The patients underwent abdominal CT plain scan and dynamic enhancement scan. Two associate chief radiologists with more than 20 years of work experience observed and analyzed all the images respectively. Observation indicators: (1) CT examination; (2) treatment and postoperative pathological examination; (3) follow-up. Follow-up was conducted using outpatient examination and telephone interview to detect the survival of patients up to December 2018. Measurement data with normal distribution were represented as Mean± SD. Measurement data with skewed distribution were described as M (range). Results:(1) CT examination: of the 30 patients, 14 had the tumor located in the fundus of stomach, 10 had the the tumor located in the body of stomach, and 6 had the tumor located in the antrum. The tumor was elliptical in 18 cases and irregular in 12 cases. There were 15 cases of endogenous type, 13 cases of exogenous type, and 2 cases of intramural type. Patients with G1 neuroendocrine tumor had the maximum diameter of (6.8±1.6)cm, of which 4 cases had the maximum diameter less than 5.0 cm and 4 cases had the maximum diameter of 5.0 to 10.0 cm. Patients with G2 neuroendocrine tumor had the maximum diameter of (8.3±2.7)cm, of which 1 case had the maximum diameter less than 5.0 cm, 4 cases had the maximum diameter of 5.0 to 10.0 cm, and 2 cases had the maximum diameter greater than 10.0 cm. Patients with G3 neuroendocrine carcinoma had the maximum diameter of (17.8±2.2)cm, of which 6 cases had the maximum diameter of 5.0 to 10.0 cm and 9 cases had the maximum diameter more than 10.0 cm. The tumor showed swelling growth in 14 cases and invasive growth in 16 cases. The tumor boundary was clear in 14 cases and unclear in 16 cases. CT plain scan showed homogeneous tumor density in 10 cases and heterogeneous density in 20 cases. Nine patients had iso-density in the tumor parenchymal part, and the CT value was (34.0±3.5)HU. In the 18 cases of low density, the CT value was (16.6±1.4)HU. In the 3 cases of high density, the CT value was (45.3±3.6)HU. Of the 30 patients, 21 cases had small punctate or small round necrotic cyst lesions in the tumor, 10 cases had mesenteric lymph node, peritoneum, liver metastasis and adjacent omentum invasion; 17 cases had abdominal effusion. In the CT enhancement examination, 12 cases showed mild enhancement, and the CT value was (56.5±6.3)HU; 15 cases showed moderate enhancement, and the CT value was (66.0±5.4)HU; 3 cases showed significant enhancement, and the CT value was (76.6±5.8)HU. Seven cases showed homogeneous enhancement and 23 cases had heterogeneous enhancement. There were 8 cases with tortuous vessels. (2) Treatment and postoperative pathological examination: of the 30 patients, 10 cases with mesenteric lymph nodes, peritoneum, liver metastasis and adjacent omentum invasion underwent radical total gastrectomy; 14 cases without surrounding tissue invasion or metastasis underwent radical subtotal gastrectomy; 6 cases with tumor diameter less than 4.0 cm and without surrounding tissue invasion or metastasis underwent endoscopic resection. All the 30 patients were confirmed GNEN by postoperative pathological examination, including 8 cases of G1 neuroendocrine tumor, 7 cases of G2 neuroendocrine tumor, and 15 cases of G3 neuroendocrine carcinoma. Results of immunohistochemical staining showed that 30 patients were positive for synaptophysin, 23 were positive for chromogranin A, and 9 were positive for cytokeratin. (3) Follow-up: all the 30 patients were followed up for 10-80 months, with a median follow-up time of 39 months. The 5-year survival rate of 30 patients was 43.3% (13/30). The 5-year survival rates were 6/8, 3/7 and 4/15 of patients with G1 neuroendocrine tumor, G2 neuroendocrine carcinoma, and G3 neuroendocrine carcinoma.Conclusions:GNEN has the main manifestation as abdominal pain, with G3 as pathological classification, which is common in fundus and body of stomach. The CT findings of GNEN are characterized by swelling or infiltrating growth and round or irregular low-density masses. Tumors are prone to cystic transformation, and showed the mildly to moderately heterogeneous enhancement.

Objective To investigate the impact of salpingectomy with preservation of fallopian tube mesentery on ovarian reserve function. Methods A total of 281 patients with tubal disease who were admitted from January 2020 to March 2021 were collected as research objects. According to the different treatment methods, they were divided into traditional bilateral resection group (n=53, conventional bilateral salpingectomy), traditional unilateral resection group (n=56, conventional unilateral salpingectomy), preservation of bilateral resection group (n=60, bilateral salpingectomy with preservation of fallopian tube mesentery), preservation of unilateral resection group (n=54, unilateral salpingectomy with preservation of fallopian tube mesentery), and control group[n=58, patients with in vitro fertilization-embryo transfer (IVF-ET) but no history of salpingectomy]. The operation time, intraoperative blood loss, and postoperative exhaust time were compared among the groups. Two months after the operation, IVF-ET was performed, and the number of follicles and oocytes, pregnancy rate, and fertilization rate were detected. The levels of anti-mullerian hormone (AMH), follicle-stimulating hormone (FSH), estradiol (E₂), and luteinizing hormone (LH) in peripheral blood were detected after IVF-ET. Follow-up was conducted until March 2023, and pregnancy success rates were recorded for the five groups. Results There were no significant differences in perioperative operation time, intraoperative blood loss, and postoperative exhaust time among traditional bilateral resection group, traditional unilateral resection group, preservation of bilateral resection group, preservation of unilateral resection group (P>0.05). After IVF-ET, there were significant differences in AMH, FSH, E₂, and LH levels among the five groups (P < 0.05). The levels of AMH and E₂ in a rising sequence were traditional bilateral resection group, traditional unilateral resection group, preservation of bilateral resection group, preservation of unilateral resection group and control group, while the levels of FSH and LH in descending sequencing were traditional bilateral resection group, traditional unilateral resection group, preservation of bilateral resection group, preservation of unilateral resection group and control group. Follow-up was conducted until March 2023, and the pregnancy success rates after IVF-ET were 45.28%, 50.00%, 53.33%, 59.26%, and 58.62% in the traditional bilateral resection group, traditional unilateral resection group, preservation of bilateral resection group, preservation of unilateral resection group, and control group, respectively. There was no significant difference in pregnancy success rate after IVF-ET among the five groups (χ²=3.044, P=0.551). Conclusion Compared with traditional salpingectomy, salpingectomy with preservation of fallopian tube mesentery has less impact on ovarian reserve function.

In order to clone the full-length cDNA of a novel EST which is probably related to acute leukemia relapse and to analyse the sequences, the electronic cloning technique combined with RT-PCR was used to clone the full-length cDNA, and the sequences were analyzed by bioinformatics. The results showed that the two novel splicing variants of eIF4E named as splicing variant 1 and 2 of eIF4E were obtained. Bioinformatics analysis showed that variant 1 and 2 exhibited 84% and 47% similarity to eIF4E mRNA, and were localized on eIF4E locus on chromosome 4. The lengths of 2 variants are 1 904 bp and 3 393 bp, encoding 245 amino acids and 132 amino acids respectively. BLAST results showed that the both variants mentioned above contains seven exons. Among them, the sequences of the three exons at 5' end of variant 1, variant 2 and eIF4E mRNA were different from each other. Protein BLAST showed that they are partially different from eIF4E protein. It is concluded that the two novel splicing variants of eIF4E were cloned, and their relation to acute leukemia relapse needs to be further investigated.
Base Sequence ; Cloning, Molecular ; Computational Biology ; DNA, Complementary ; genetics ; DNA, Recombinant ; Eukaryotic Initiation Factor-4E ; genetics ; Humans ; Molecular Sequence Data ; Protein Isoforms ; genetics ; Reverse Transcriptase Polymerase Chain Reaction

The purpose of this study was to compare the differences of the protein expression profiles between human myeloid leukemia K562 cells and adriamycin-resistant K562/A02 cells, as well as to select novel resistance-related proteins in myeloid leukemia by means of proteomics. The total cellular proteins were separated from K562 and adriamycin-resistant K562/A02 cells by using technique of two dimensional difference in gel electrophoresis (2D-DIGE). Differentially expressed proteins were analyzed by matrix-assisted laser desorption ionization/time of flight-mass spectrometry (MALDI-TOF/MS), and by protein database searching. Moreover, the differentially expressed proteins were verified at protein and mRNA levels by Western blot assay and quantitative real time PCR. The results showed that 8 proteins differentially expressed in adriamycin-resistant K562/A02 cells, among them 2 proteins were identified to be down-regulated and 6 to be up-regulated. These identified proteins involved in the cell energy metabolism, cell proliferation, cell apoptosis, signal transduction, gene transcription and translation respectively. The results assayed by Western blot were similar to those detected by 2D-PAGE. Two up-regulated proteins Stathmin and CrkL were selected for verification in K562 and K562/A02 cells. As a result, the results detected by Western blot were identical with results from 2D-DIGE; real time quantitative PCR assay showed that the changes of CrkL at mRNA level were identical with changes at protein level, but no complete identity of Stathmin changes at mRNA level and protein level was observed. It is concluded that the difference of protein expression profile exists in K562 and K562/A02 cells. Stathmin and CrkL proteins may be involved in the drug resistance and suggest a novel clue for the resistant mechanisms in myeloid leukemia, which is worth further to explore.
Adaptor Proteins, Signal Transducing ; metabolism ; Doxorubicin ; pharmacology ; Drug Resistance, Neoplasm ; Humans ; K562 Cells ; Leukemia ; metabolism ; Nuclear Proteins ; metabolism ; Stathmin ; metabolism

This study was aimed to explore the effects of expressing eukaryotic elongation factor 1A1 (eEF1A1) on proliferation and apoptosis in human acute T lymphocytic leukemia (T-ALL) cell line Jurkat with knocked down eEF1A1 gene and its mechanisms. eEF1A1-expressing lentivirus (LV) was constructed and used to transfect the Jurkat cells with knocked down eEF1A1 gene. Then, the expressions of eEF1A1 mRNA and protein were detected by real time PCR(RT-PCR) and Western blot respectively.Cell proliferation, apoptosis and cycle were detected by MTT method, Annexin V-APC labeling and DNA ploidy analysis respectively. The related protein expressions of phosphatidylinositol-3-kinase (PI3K)/serine/threonine kinase (Akt) signaling pathway were detected by Western blot. The results indicated that eEF1A1 mRNA and protein expressions of Jurkat cells with knocked down eEF1A1 gene were re-established by constructing eEF1A1-expression LV. Compared with negative control group (transfected with negative control LV and eEF1A1-shRNA LV), cell proliferation in eEF1A1 expression group was significantly enhanced, cell apoptosis was remarkably inhibited, percentage of cells in G0/G1 phase was significantly reduced alone with increased percentage of cells in S and G2/M phase, and the expression levels of p-Akt (Ser 473), nuclear factor kappa B (NF-κB), p-NF-κB (Ser 468), mammalian target of rapamycin (mTOR) and p-mTOR (Ser 2448) protein significantly increased. It is concluded that eEF1A1 may have a carcinogenic effect in T-ALL cells. eEF1A1 expression has noticeable effects on the proliferation enhancement and apoptosis inhibition of Jurkat cells, which may be mediated by the up-regulation of PI3K/Akt/NF-κB and PI3K/Akt/ mTOR signaling pathway.
Apoptosis ; Cell Proliferation ; Gene Expression ; Humans ; Jurkat Cells ; Peptide Elongation Factor 1 ; genetics ; metabolism ; Proto-Oncogene Proteins c-akt ; metabolism ; RNA, Small Interfering ; genetics ; Signal Transduction

Based on the sequence of BAC (Bacterial Artificial Chromosome) along with the Cre/lox P system, the gene-targeting vectors to multiple loci of the repetitive internal transcribed spacers between rDNA genes in Leghorn chicken were constructed. The key material of multiple loci gene targeting in vivo would be obtained. First, the plasmid of pYLSV-TDN with TK, HRDS2, and Neo genes was constructed. The TK-HRDS2-Neo DNA fragment obtained from the plasmid of pYLSV-TDN was digested by Not I/HindIII and inserted into the upstream of the lox P site of BAC plasmid for obtaining the selective vector of BAC-TDN. The expression vector of pYLVS-GID with EGFP, hIFN genes, and HRDS1 was then obtained. The plasmid of BAC-TDN-VS-GID was obtained by cotransformation of the selective vector of BAC-TDN and the expression vector of pYLVS-GID to E. coli NS3529 through the action of Cre/lox P system. The gene-targeting vector of BAC-TDN-GID to multiple loci of the ITS region in Leghorn chicken was obtained by cleaving the sequence of pYLVS with the homing endonuclease of I -Sce I and ligating with the linker of LS. The insertion and the insert direction of DNA fragments were identified by restriction digestion or PCR and sequencing in each clone. The significance of the technique ofgene-targeting vector to multiple loci are shown as follows. First, the targeting loci were increased to 100 - 300. Second, the problems of unstable expression of inserted genes were partially solved. Third, the need for safety against toxicity integration was resolved. Fourth, the forbidden zone of gene integrating on the repetitive DNA sequences was broken through.
Animals ; Attachment Sites, Microbiological ; genetics ; Chromosomes, Artificial, Bacterial ; genetics ; Cloning, Molecular ; DNA ; genetics ; metabolism ; DNA Restriction Enzymes ; metabolism ; DNA, Ribosomal Spacer ; genetics ; Escherichia coli ; genetics ; Genetic Vectors ; genetics ; Green Fluorescent Proteins ; genetics ; Humans ; Integrases ; genetics ; Interferon-gamma ; genetics ; Polymerase Chain Reaction ; Recombinant Fusion Proteins ; genetics ; Transformation, Genetic

OBJECTIVETo study hPARP1 genetic polymorphism in southern Chinese Han and Miao populations.

METHODSBlood samples from 187 and 210 southern healthy Han and Miao populations were collected. The mutations of exons 12,13,16 and 17 of hPARP1 gene were investigated by PCR-single-strand conformation polymorphism(SSCP).

RESULTSFragments of 253 bp,313 bp,175 bp,362 bp within exons 12,13,16 and 17 respectively of hPARP1 gene were amplified by multiple PCR. An SSCP variant in exons 12,13,16 and 17 of PARP1 gene in 187 healthy Han and 210 healthy Miao individuals was identified. Seven single-base substitutions compared with the sequence of PARP1 gene were identified: a T to C transition in exon 12 (Phe548Ser), a G to T transition in exon 13 (Ala683Ser), a G to T transition in exon 16 (Asp798Tyr), and a A to G transition in exon 17 (His808Arg).

CONCLUSIONThere were polymorphism sites in exons 12,13,16,17 of hPARP1 gene in southern Chinese Han and Miao populations; these results may be useful for the establishment of PARP1 genotyping, and these newly described PARP1 alleles would be advantageous indicators for population studies.

Adult ; Alleles ; China ; Exons ; genetics ; Female ; Gene Frequency ; Genotype ; Humans ; Male ; Poly (ADP-Ribose) Polymerase-1 ; Poly(ADP-ribose) Polymerases ; genetics ; Polymerase Chain Reaction ; Polymorphism, Single Nucleotide ; genetics ; Polymorphism, Single-Stranded Conformational

OBJECTIVETO establish a RP-HPLC method for the determination of calycosin-7-O-beta-D-glucopyranoside in Radix Astragali, and to analyse the calycosin-7-O-beta-D-glucopyranoside content of ten samples of Radix Astragali, collected from different regions.

METHODA Polaris C18(250 mm x 4.6 mm, 5 microns) column was used and a mixture of methanol-water (30:70) was used as the mobile phase at a flow rate of 1.0 mL.min-1. The column temperature was 25 degrees C and the UV detection wavelength was 254 nm.

RESULTThe calibration curve was in good linearity over the range of 0.0106-2.12 micrograms with the regression equation Y = 3035. 97 X - 14.85(r = 0.9999). The average recovery was 95.8% (n = 5, RSD = 1.3%).

CONCLUSIONThe method is simple, quick, sensitive and reproducible. In all of the samples, the calycosin-7-O-beta-D-glucopyranoside contents differ markedly.

Astragalus membranaceus ; chemistry ; classification ; China ; Chromatography, High Pressure Liquid ; Ecosystem ; Glucosides ; analysis ; Isoflavones ; analysis ; Plant Roots ; chemistry ; Plants, Medicinal ; chemistry ; Quality Control ; Species Specificity