Spinal infection caused by Prevotella intermedia:analysis of misdiagnosis and literature review / 中国感染控制杂志

Objective To explore the clinical characteristics and treatment scheme of patients with spinal infection caused by Prevotella intermedia(P.intermedia).Methods Clinical diagnosis and treatment processes of a patient with spinal infection caused by P.intermedia admitted to the spinal surgery department of a hospital were summa-rized,and relevant literature was retrieved from database for reviewing.Results The patient,a 50 year old male,was admitted to the hospital due to"lumbago pain complicated with pain in double lower extremities for 2 months".The lesion tissue was taken for metagenomic next-generation sequencing(mNGS)detection,which detected P.in-termedia,and the patient was diagnosed with P.intermedia spondylitis.After treatments with open lesion clea-rance,tube rinsing+autologous bone transplantation fusion internal fixation,intravenous drip of ceftriaxone sodium and metronidazole,as well as metronidazole rinsing,infection was under control.A total of 16 available papers were retrieved,together with this case,a total of 17 patients were included,with 7 males and 10 females.The main risk factors were diabetes and history of corticosteroid use(35.3％).The most common invasion sites were lumbar ver-tebra(n=12)and thoracic vertebra(n=6).13 cases were positive for pathogen culture,3 cases were positive for molecular detection,and 1 case was positive for staining microscopy.17 patients received anti-anaerobic bacteria treatment,with 14 cases receiving combined surgical treatment.One case died,with a mortality of 5.9％;5 cases had partial neurological impairment,with a disability rate of 29.4％.The survival rate of patients who received treatment of anti-anaerobic bacteria combined with surgery was 92.8％,3 patients only with anti-anaerobic bacteria treatment but without surgery were all cured.Conclusion P.intermedia is an opportunistic pathogeanic bacteria which often causes infection in immunocomprised individuals and is prone to be misdiagnosed.It is recommended to perform mNGS detection to identify the pathogen as early as possible and seize the opportunity for treatment to reduce mortality.

Effects of percutaneous vertebropIasty with different bone cement voIumes on the adjacent intervertebraI disc in a rabbit modeI of osteoporosis / 中国组织工程研究

BACKGROUND: With the development of clinical practices, some complications of vertebroplasty are gradually highlighted. Considering the thermal reaction, toxicity and space-occupying filling of bone cement, vertebroplasty with bone cement injection is likely to cause some impacts on the surrounding vertebrae, especially the adjacent ones. However, little is reported on the effect of different volumes of bone cement on degeneration of the adjacent vertebrae. OBJECTIVE: To determine the effects of different bone cement volumes in percutaneous vertebroplasty on adjacent intervertebral discs in a rabbit model of osteoporosis. METHODS: Thirty 5-month-old New Zealand rabbits were randomly assigned into five groups: sham, iohexol, low-dose bone cement, middle-dose bone cement and high-dose bone cement groups. Osteoporosis models were established by ovariectomy combined with glucocorticoid injection. After modeling, sham puncture at the fifth lumbar vertebrae was done in the sham group, and true puncture at the fifth lumber vertebra was done in the iohexol group followed by injection with 0.2 mL of iohexol. Different volumes of bone cement (0.1, 0.2, 0.3 mL) were injected in the low-, middle- and high-dose bone cement groups, respectively. After 12 weeks, rabbits in these three groups were killed to take the intact L4-5segments that were divided into two parts: one for TUNEL staining to observe cell apoptosis and to calculate apoptotic index, and the other for real-time PCR detection of relative expression of interleukin-1, type II collagen and matrix metalloproteinase-7. RESULTS AND CONCLUSION: The cell apoptosis index in the low-, middle- and high-dose bone cement groups were significantly different from that in the sham and iohexol groups, but there was no significant difference among three bone cement groups. The relative expression of interleukin-1 mRNA showed a significant difference between the high-dose group and sham group. The relative expression of type II collagen and matrix metalloproteinase-7 in the three bone cement groups was significantly different from that in the sham and iohexol groups, and the expression in the high-dose bone cement group was also significantly different from that in the low- and middle-dose bone cement groups. To conclude, after percutaneous vertebroplasty, injected bone cement certainly impacts the adjacent vertebrae in the rabbit model of osteoporosis, but not in a dose-dependent manner. Moreover, a dramatic aggravation of degeneration of the adjacent vertebrae is developed when high-dose bone cement is injected.

Effect of sphingosine 1-phosphate/sphingosine 1-phosphate receptor signal pathway on function of neutrophils / 中国实验血液学杂志

The aim of this study was to examine the priming effect of sphingosine 1-phosphate (S1P) on fMLP-activated neutrophils, mainly to detect the neutrophil respiratory burst products, and to investigate the signaling pathway involved in S1P activity. Flow cytometry was used to evaluate the new isolated neutrophil; the superoxide anion output was detected indirectly by cytochrome C reduction in respiratory burst; the dihydro-rhodamine 123 was used to detect the intensity of respiratory burst; the signal transduction pathways of neutrophil respiratory burst were explored by Western blot. The results showed that after pretreated with S1P, the level of superoxide anion released by fMLP-activated neutrophils significantly increased; the Rhodamine 123 mean fluorescence intensity in S1P primed fMLP-activated neutrophils group was significantly higher than that in fMLP treatment group; PI3K and Akt proteins involved in the signal pathway of neutrophil respiratory burst. It is concluded that S1P is a new priming reagent, which primes respiratory burst of fMLP-activated neutrophils; this signal pathway may be that S1P interacts with its receptor, activates PI3K, then activates Akt-transmitting signals through NADPH oxidase, finally results in the respiratory burst.

The role of third-party tolerogenic dendritic cells in the prevention of acute graft-versus-host-disease following allogeneic bone marrow transplantation in mice / 中华血液学杂志

<p><b>OBJECTIVE</b>To explore the biological characteristic of third-party-derived tolerogenic DC(tDC) and the influence of third-party-derived tDC on acute graft-versus-host-disease (aGVHD) following allogeneic bone marrow transplantation (allo-BMT) in mice.</p><p><b>METHODS</b>tDC from bone marrow cells of D1 mice was cultured with low doses of GM-CSF, IL-10 and TGF-β1D1. The phenotype, expression of cytokines and function associated molecules were identified with FACS and RT-PCR. Mixed lymphocyte reaction was applied to analyze the influence of third-party-derived tDC on allo-CD4(+)T cells proliferation in vitro. Different doses of D1-tDC were adoptive transferred in the aGVHD model in allogeneic BMT which B6 mice as donors and D2 mice as recipients. Survival time, clinical GVHD score and the levels of Th1/2 cytokines in serum were monitored after allo-BMT using the aGVHD model as control.</p><p><b>RESULTS</b>tDC expressed lower levels of MHC II and co-stimulatory molecules, such as CD80, CD86 and CD40, even when stimulated by LPS. The results by RT-PCR indicated that tDC expressed low levels of IL-12p40 and high levels of immunosuppressive molecules, such as IL-10, TGF-β, Fas Ligand, indoleamine 2, 3-dioxygenase (IDO) and arginase. In the allogeneic MLR, third-party tDC suppressed allo-CD4(+)T cells proliferation, which was relative to the dose of tDC. In the B6→D2 mouse model, all aGVHD mice died within 18 days. Remarkably, if 10(4) third-party tDC were transferred, 60% mice survived at least 60 days. When the doses of tDC were reduced to 10(3) cells, only 20% of mice survived day 60, and when increased tDC to 10(5), all of the mice died within day 37 after allo-BMT. The cytokine levels in serum indicated that 10(4) tDC-treated mice secreted in vivo high level of IL-10 21d after BMT (P < 0.05), the levels of IL-10 in 10(3), 10(4) and 10(5) tDC-treated mice were (114.23 ± 7.78), (646.18 ± 212.02), (121.97 ± 10.47) ng/L, respectively.</p><p><b>CONCLUSION</b>Third-party tDC could suppress allo-CD4(+)T cells proliferation in vitro and prevent aGVHD in allogeneic BMT mode, which may be mediated by modulating tolerogenic cytokines secretion, such as IL-10. And this effect was associated with the dose of tDC. Adoptive therapy by transfusing third-party tDC cultured with low doses of GM-CSF, IL-10 and TGF-β1 could significantly prolong the survival of recipients and prevent aGVHD in allogeneic BMT.</p>

A preliminary study on the influence of human plasma exosomes-like vesicles on macrophage Wnt5A-Ca²+ pathway / 中华血液学杂志

<p><b>OBJECTIVE</b>To study the influence of human plasma exosomes-like vesicles on the regulatory function of macrophages.</p><p><b>METHODS</b>The exosomes-like vesicles were purified from healthy donors plasma with a series of high-speed centrifugation and ultrafiltration. Macrophages were derived from cultured human blood monocytes. The molecular markers of macrophages were assayed by FACS. After cultured with exosomes-like vesicles, the changes of macrophages cytoplasma Ca(2+), and related genes and proteins were assayed by FACS, RT-PCR and Western Blot, respectively.</p><p><b>RESULTS</b>After cultured with exosomes-like vesicles, mean fluorescent intensity (MFI) of macrophages cytoplasma Ca(2+) was increased. The vesicles enhanced macrophages to express cytokines genes, the expression of IL-1β and TNF-α genes being increased by 0.85 and 1.69 times respectively at 2 h, and that of IL-6 gene 3.7 times compared with the control at 8 h. However, the vesicles inhibited the expression of macrophages IL-10 gene, had no influence on the Frizzled5 receptor expression and could induce CaMKII phosphorylation.</p><p><b>CONCLUSIONS</b>Exosomes-like vesicles can up-regulat macrophages expression of inflammatory cytokines genes, and increase the secretion of inflammatory cytokines by activating the Wnt5A-Ca(2+) signaling pathway.</p>

A proliminary study on the regulatory function of human plasma exosomes-like vesicles / 中华血液学杂志

<p><b>OBJECTIVE</b>To identify the exosomes-like vesicles from the plasma and study their biologic characteristics and regulatory effect.</p><p><b>METHODS</b>The exosomes-like vesicles were purified from healthy donors plasma with a series of high-speed centrifugations and ultrafiltration. Morphology was identified by transmission electron microscopy and biologic characteristics by Western blot and flow cytometry. CD4(+)T cells and CD4(+)CD25(+)CD127low Treg cells were purified from peripheral blood mononuclear cells (PBMCs) by Magnetic cell sorting. After exosomes-like vesicles cultured with CD4(+)T cells or CD4(+)CD25(+)CD127low Treg cells, cell proliferation and apoptosis were assayed. Phosphorylated β-catenin level in Wnt signaling by phosflow.</p><p><b>RESULTS</b>Exosomes-like vesicles from plasma were similar to previously described exosomes in shapes and size and expressed exosome marker proteins CD63 and CD81 as well as the MHC-II molecule, costimulatory molecules CD86 etc. After co-cultured with CD4(+) T cells, exosomes-like vesicles inhibited the proliferation of the T cells in a dose-dependent manner. After Treg cells cultured with exosomes-like vesicles for 14 days, the survival rate of the Treg cells was 57.07%, while that of the control Treg was 30.91%. Frizzled receptors 2, 3, 4and LRP6 gene mRNA expressed (the relative gray value was 48.50, 34.84, 23.85, 49.73) in the Treg cells by RT-PCR, and Wnt molecular expressed in exosomes-like vesicles. After Treg cells co-cultured with exosomes-like vesicles, the MFI of phosphorylated β-catenin decreased (from 20.06 ± 2.99 to 12.41 ± 2.08), and the expression of Bcl-2 mRNA was upregulated significantly (the relative gray value from 0.45 to 84.97).</p><p><b>CONCLUSIONS</b>Exosomes-like vesicles existed in human plasma and express immune regulatory molecules. They can suppress the proliferation of activated CD4(+) T cells induce their apoptosis and pro-long the survival of natural Treg cells via Wnt signaling pathway.</p>

Immunity mechanism of exosomes derived from dendritic cells / 中国实验血液学杂志

To confirm the mechanism of exosomes as tumor vaccines inducing immunity response, dendritic cells (DCs) were induced from human peripheral blood mononuclear cells, while exosomes were isolated from DC loaded tumor antigen. The effect of exosomes on priming T cell proliferation was analysed under conditions with or without DCs, or DCs at different mature stages. The function of exosomes in immunity was detected through block test after blocking some molecules (CD11a, CD11b, CD11c, CD54, MFG-E8 and CD83). The effect of DCs on embedded exosomes was observed by confocal microscopy, the effect of blocking surface molecules on exosomes on DC-embedding exosomes was assayed by flow cytometry. The results indicated that both exosomes derived from imDC (imDex) and exosomes derived from mDC (mDex) could not prime T cells without DC or with imDC. The exosomes derived from mDC induced with different cytokines (LPS, TNF-alpha, CpG, CD40L) were no significant difference in concentrations but were different in effect. The immunity function of exosomes depended on CD11a, CD11b, CD11c, CD54, MFG-E8 and CD83 molecules, the effect of priming T cells is reduced when these molecules were blocked. Confocal microscopy and FACS assay showed that blocking CD11a and CD54 could inhibit exosome-targeted DC and DC-embedded exosomes. It is concluded that the exosomes target DCs through their surface molecules, therefore results in immune response of T cells.

Immune tolerance induced by exosomes derived from regulatory dendritic cells of mice / 中国实验血液学杂志

The study was aimed to explore the roles of exosomes derived from regulatory dendritic cells of mice in the induction of immune tolerance. Immature DC (iDC) from mouse bone marrow cells and regulatory DCs (rDC) were induced by treating iDC with TGF-beta1 and IL-10. The phenotype of regulatory DCs and normal DCs were assayed by flow cytometry. Exosomes from immature DCs (iDex) and regulatory DCs (rDex) were isolated by ultracentrifugation and ultrafiltration. A skin transplantation model was established with the recipients BALB/c mice and the donor C57BL/6 mice. Recipients were divided into PBS control group, iDex group (injection 10 microg iDex of donor C57BL/6 mice via tail vein at days 7 and 3 before skin transplantation), rDex group (injection 10 microg rDex of donor C57BL/6 mice via tail vein at days 7 and 3 before skin transplantation). The capacity of the donor mice and the unrelated allogeneic donor mice to stimulate allogeneic T lymphocyte proliferation was examined by mixed lymphocyte culture (MLR). The results showed that TGF-beta1 and IL-10 could down-regulate the expressions of costimulatory molecules, including CD80, CD86 and CD40. The graft mean survival time (MST) in control group, iDex group and rDex group was 7.8, 10.7 and 18.8 days, respectively. There was significant difference in MST between iDex group and control group (p<0.05), and between rDex group and iDex group (p<0.01). The results of MLR assays indicated donor-specific hyporeactivity especially in rDex group, while the tolerant B/C mice were still immunocompetent to unrelated allogeneic DBA mouse. It is concluded that injection iDex or rDex of donor mice via tail vein before skin transplantation induces immunotolerance, and the effect of rDex is more significant.

A preliminary study on the biological characteristics and function of exosomes derived from dendritic cells / 中华血液学杂志

<p><b>OBJECTIVE</b>To establish a method for isolating exosomes from dendritic cells (DC), and to analyse its biological characteristics and function in antitumor immunity.</p><p><b>METHODS</b>Immature DCs (im-DC) from human peripheral blood mononuclear cells were loaded with the antigen of K562 tumor cells, then exosomes were secreted from imDC and lipopolysaccharide (LPS) induced mature DC (mDC). The exosomes from imDC and mDC were isolated separately by ultracentrifugation and ultrafiltration. The exosomes diameter was determined, their profile was observed by electron microscope, and the surface molecules were detected by Western blot. To analyse the effect of exosomes on antitumor immunity, the proliferation, IFN-gamma expression, CD69 up-regulation and cytotoxicity of antigen-specific T cells were measured.</p><p><b>RESULTS</b>Exosomes were small flattened sphere vesicles with an average diameter of 72.3 nm and expressed CD80, CD86, HLA-DR, FasL, CD54 and MFG-E8 molecules. As compared to immature exosomes, exosomes from mDC were proved to express more CD80 and less MFG-E8, to be more potent for inducing antigen-specific T cells proliferation and immunity respond in vitro: at its optimum concentration, the absorption value of T cell proliferation test was 0.50 +/- 0.01, CD69 was up-regulated and (13.4 +/- 5.8)% of T cells was in proliferating, (22.8 +/-2.4)% of T cells expressed IFN-gamma, and (21.3 +/-8.6)% of tumor cells were killed.</p><p><b>CONCLUSION</b>A simple and quick method to isolate and analyse exosomes is established. The exosomes can induce antitumor immunity respond.</p>

Proliferation of natural killer T cells in umbilical cord blood and peripheral blood and their different phenotypes / 中国实验血液学杂志

Purpose of this study was to establish an effective method in vitro to proliferate natural killer T (NKT) cells from umbilical cord blood (UCB) and peripheral blood (PB), and to study their different phenotype. Mononuclear cells (MNC) from UCB and PB were cultured in the presence of IL-2 (100 U/ml), with or without alpha-Galcer. TCR Valpha24 Vbeta11 double positive natural killer T-cells (NKT cells) and their other phenotypes were determined by flow cytometry. The results showed that after expansion for 7 days, TCRValphabeta(+) NKT cells from UCB-MNCs increased by (8.74 +/- 4.37) x 10(2) times as much, but most of them did not express NK1.1 and its TCR Vbeta11(+) was higher than TCR Valpha24(+). After expansion for 14 days, TCR Valphabeta(+) NKT cells from PB-MNCs increased by (3.72 +/- 2.01) x 10(2) times, the expression of NK1.1 was high and its TCR Vbeta11(+) was almost equal to TCR Valpha24(+). It is concluded that human TCR Valpha24 Vbeta11 double positive NKT cells can expand by addition of alpha-Galcer. The proliferation efficiency in UCB-MNCs is greater than that in PB-MNCs. Most of the UCB-NKT is NK1.1(-), while the PB-NKT is NK1.1(+), a new subset of NKT cells.

Study on the targeting effects of M1-GS RNA on K562 cells / 中华血液学杂志

<p><b>OBJECTIVE</b>To determine the effects of M1-GS RNA (M1 RNA) on bcr-abl mRNA and oncoprotein after M1 RNA with guide sequence (M1-GS RNA) targeting the oncogene was transfected into K562 cells.</p><p><b>METHODS</b>pAVGS4 (an eukaryocyte expression vector containing M1-GS RNA sequence) and pNAV-1 (as the control) were transfected into K562 cells by X-tremeGENE Q2. Total RNA was extracted at 24, 48, 72 and 96 hours after transfection. Then RT-PCR was done to compare the products at different time point. After collecting pAVGS4-transfected cells and the control cells at 48 and 96 hours after transfection, total protein was extracted and quantified. Change of P210 was determined by Western blot. Colony formation was analyzed at 96 hours after transfection.</p><p><b>RESULTS</b>RT-PCR based on transfected cells at different time point showed that the amount of bcr-abl mRNA began to decrease at 24 hours and reduced to 9.2% and 2.5% respectively at 48 and 72 hours after transfection. Western blot showed that the expression of P210 in the pAVGS4 group reduced to 10.4% of the control at 48 hours and 6.7% of the control at 96 hours after transfection. The inhibition rate of colony formation was 81.3% after K562 cells were transfected by pAVGS4.</p><p><b>CONCLUSION</b>pAVGS4 can efficiently destroy bcr-abl mRNA in K562 cells. The transcript level of bcr-abl mRNA was reduced with the time after transfection. The expression of P210 was decreased significantly at 48 and 96 hours after transfection. K562 cell colony formation was prominently inhibited.</p>

Effect of HMBA on differentiation and apoptosis of HL-60 and U937 cells and its mechanism / 中华血液学杂志

<p><b>OBJECTIVES</b>To explore the effect of hexamethylene bisacetamide (HMBA) on the differentiation and apoptosis of HL-60 and U937 cells, and its mechanism.</p><p><b>METHODS</b>Flow cytometry was used to evaluate the expressions of cellular surface antigen CD(11b), CD(14), apoptotic marker Annexin V, cell cycle distribution and endocytic antigen cyclin D, cyclin E and p27. Changes of c-myc, Rb, Bcl-2 gene mRNA levels were detected by RT-PCR.</p><p><b>RESULTS</b>After 72 hours of HMBA treatment, CD(11b) expressions increased significantly, apoptosis increased under high-dose HMBA, cells were arrested in G(0)/G(1) phase and reduced cyclin E, increased cyclin D and p27 were significant in a dose-dependent manner in HL-60 and U937 cells. RT-PCR showed that c-myc and bcl-2 mRNA was significantly down-regulated and Rb mRNA up-regulated in HL-60 and U937 cells.</p><p><b>CONCLUSION</b>HMBA can induce the differentiation of HL-60 and U937 cells, while apoptosis of these cell is induced only by high dose of HMBA. The possible mechanism of HMBA inducing differentiation might be related to the changes of cell cycle regulators and certain proliferation and differentiation related genes.</p>

Experimental study of K562 cell apoptosis induced by siRNA / 中华血液学杂志

<p><b>OBJECTIVES</b>To construct a siRNA expression vector pBCR6 that produces siRNA against bcr/abl mRNA and detect apoptosis rate of K562 cells after pBCR6 transfection.</p><p><b>METHODS</b>Template sequence for siRNA was designed, synthesized and inserted into an expression vector pSilencer1.0-U6. Restriction analysis and sequencing were performed to verify the pBCR6 vector. Then pBCR6 was transfected into K562 cells by X-tremeGene Q2. pSilencer1.0-U6 was used as the control. At different time point after transfection, apoptosis rate was determined by Tunel and Annexin V+ PI with FCM.</p><p><b>RESULT</b>pBCR6 was verified by restriction analysis and sequencing. The apoptosis rate of K562 cells markedly increased at 48 and 72 hour after transfected with pBCR6, and increased in a time-dependent manner [the apoptosis rate of transfected K562 cells was (47.80 +/- 1.63)% at 72 hrs, whereas the control group was (6.67 +/- 0.37)%, P < 0.0001] No prominent change in apoptosis rate was found in the control.</p><p><b>CONCLUSION</b>The siRNA expression vector against bcr/abl mRNA was successfully constructed. The pilot study showed that pBCR6 could effectively induce K562 cells apoptosis. siRNA may be a new tool for molecular target therapy for chronic myelogenous leukemia.</p>

Effects of hexamethylene bisacetamide on cell cycle and expression of its regulatory proteins in HL-60 cells / 中国实验血液学杂志

Hexamethylene bisacetamide (HMBA) is referred as a differentiation-inducer for the clinical treatment of acute myeloid leukemia and myelodysplastic syndrome. However, the molecular mechanism of the effects of HMBA on myeloid leukemic cells remains unknown. In this study, the effects of HMBA on cell cycle and expression of cell cycle regulatory proteins in HL-60 cell were investigated in order to explore its pharmacological mechanism. The altered distribution of cell cycle and expression of its regulatory proteins (cyclin D, cyclin E and p27) in HL-6 0 cell induced by HMBA were analyzed by flow cytometry. The effects on transcription for mRNA of CKI p15, p16 and p27 in HL-60 cell were further studied by RT-PCR. The results showed that HMBA could mainly commit HL-60 cell to G0/G1 arrest and the significantly decreased endocytic cyclin E protein and increased cyclin D/p27 protein after HMBA treatment were found. There was no expression of p15, p16 mRNA in untreated HL-60 cell and 3 mmol/L of HMBA could make them expressed after exposed for 24 h or 48 h respectively. The expression of p27 mRNA was positive and no obviously different in untreated HL-60 cells exposed for 24 h, 48 h and 72 h. These results suggested that one of the pharmacological mechanisms of HMBA was to elevate the expression of p27 and reduce the cyclin E expression as well as to activate the expression of p15, p16 gene mRNA, that arrested cell at G0/G1 and exerted its effects of anti-proliferation.

The biological activity of MHC classII transactivator ribonuclease P: a novel approach for hepatic transplantation rejection / 中华肝脏病杂志

<p><b>OBJECTIVE</b>This paper studied the effect of RNaseP against CIITA on repressing class II MHC (MHCII) expression.</p><p><b>METHODS</b>It was constructed that M1-RNA with guide sequences (GS), recognizing the 629 site of CIITA (M1-629-GS), by PCR from pTK117 plasmid, then was cloned into psNAV (psNAV-M1-629-GS). CIITA target gene was obtained from Raji cell by RT-PCR, and then inserted into pGEM-7zf (+) (pGEM-800). psNAV-M1-629-GS and pGEM-800 were transcribed and then mixed up and incubated in vitro. Stable transfectants of hepatocyte with psNAV-M1-629-GS by nanometer were tested for MHCII induction by recombinant human interferon-gamma (IFN-gamma). mRNA abundance of CIITA was measured by RT-PCR.</p><p><b>RESULTS</b>It showed that M1-629-GS could exclusively cleave pGEM-800 that formed a base pair with the GS. When induced with IFN-gamma, the expression of HLA-DR, -DP, -DQ on psNAV-M1-629-GS+ hepatocyte was (1.01+/-0.51)%, (4.37+/-1.28)%, (1.98+/-0.42)% respectively, was down-modulated 90.65%, 89.11% and 65.32% compared with control, while the mRNA content of CIITA reduced significantly (P<0.01).</p><p><b>CONCLUSION</b>M1-629-GS could effectively repress MHCII expressing through cleaving CIITA mRNA. These results provided insight into the future application of it as a new nucleic acid drug against the rejection of hepatic transplantation.</p>

Inhibition effect of CIITA anti-sense cDNA on the expression of MHCII molecules of Hela cells / 中华血液学杂志

<p><b>OBJECTIVE</b>To investigate the inhibiting effect of classII major histocompatibility complex transactivator (CIITA) anti-sense cDNA on MHCII molecules expression.</p><p><b>METHODS</b>CIITA antisense cDNAs (arII1, arII2 and arII3) were obtained from Raji cell by RT-PCR, and then inserted into the pcDNA3.1B plasmid. The stable transfectants of Hela cell were tested for CIITA protein by immunohistochemistry, and classic MHCII molecules (HLA-DR, -DP, -DQ) induction through recombinant human IFN-gamma by flow cytometry. mRNAs of CIITA, invariant chain and classic MHCII molecules were detected by RT-PCR.</p><p><b>RESULTS</b>The effect of arII2 was the best. When induced with IFN-gamma, the expression of CIITA protein was inhibited by 87.23%, and that of HLA-DR, -DP and -DQ was almost totally inhibited on arII2 positive Hela cells. The mRNA contents of CIITA, HLA-DR, -DP, -DQ and invariant chain were decreased significantly (P < 0.05).</p><p><b>CONCLUSIONS</b>arII2 inhibits CIITA and thus the family of MHCIImolecules regulated by CIITA, therefore, it provides a novel approach to graft versus host disease study in hematopoietic stem cell transplantation.</p>

Clinical and laboratory studies of expression of cyclin A in leukemia cells / 中国实验血液学杂志

Uncontrolled cell proliferation is the basic feature of cancer. Some of the prime cell cycle regulators are involved directly in tumorigenesis. Cyclin A, one of the G(1)/S cyclin, can cause transformation. The purpose of this research was to investigate whether cyclin A overexpression was involved in leukemogenesis and proliferation of leukemia cells. The expression of cyclin A at S-phase in leukemia cell line HL-60, blast cells of acute leukemia patients, bone marrow cells of outpatients without malignant hematological disease and peripheral blood cells of healthy donors was investigated by simultaneous indirect immunofluorescence staining of intracellular antigen and DNA. To further investigate whether cyclin A played as a key molecular in cell proliferation, HL-60 cells were exposed to different concentrations of hexamethylene bisacetamide (HMBA). MTT dye absorbance of living cells and cell cycle analysis were adopted to evaluate growth arrest. Differentiation was evaluated by detection of the change of expression of CD11b and CD33 on cell surface. The results showed that overexpression of cyclin A was only found among specimens from acute leukemia and leukemia cell line. There was no elevated cyclin A detection for cyclin A among specimens from outpatients and healthy donors. In HMBA interference experiment, HMBA was able to induce growth arrest and monocytic macrophage differentiation of HL-60 cells in a dose-dependent manner, and all these changes were associated with a marked down-regulation of cyclin A expression. In conclusion, aberrant overexpression of cyclin A at S-phase was only found in leukemia cell lines and blast cells from acute leukemia. The dose-dependent effect of HMBA on cell growth and differentiation of HL-60 cell line which was consistent with the decrease of cyclin A expression in these cells suggested that the molecular mechanisms of HMBA inducement involved downregulation of cyclin A expression.

Activation of random-donor platelet concentrates and platelet prepared from random-donor apheresis collections during storage / 中国实验血液学杂志

The activation of random-donor platelet concentrates and platelets prepared from random-donor apheresis collections (plateletpheresis) during storage were studied. Percentage of CD62p staining and mean channel fluorescence (MCF) of CD41 of two kinds of platelets during storage on day 0, day 1, day 3 and day 5 were determined by flow cytometry. The results showed that percentages of CD62p staining and MCF of CD41 in plateletpheresis were (18.91 +/- 6.25)%, (19.48 +/- 8.27)%, (22.82 +/- 6.06)%, (56.71 +/- 11.79)% and (8.09 +/- 2.38)%, (8.13 +/- 2.45)%, (8.44 +/- 2.51)%, (19.87 +/- 6.13)%, while the results of platelet concentrates were (30.65 +/- 12.33)%, (31.46 +/- 11.86)%, (32.51 +/- 13.05)%, (63.55 +/- 13.27)% and (10.33 +/- 4.37)%, (11.09 +/- 6.61)%, (13.46 +/- 9.69)%, (24.41 +/- 10.15)%, respectively. The platelet count and pH value were also determined. The platelet number, pH value, percentage of CD62p staining and MCF of CD41 had no significant difference within 3 days of platelet storage. The platelet number and pH value decreased significantly (P < 0.001), while percentages of CD62p staining and MCF of CD41 increased significantly (P < 0.001) on day 5 of storage. It is concluded that the quality of plateletpheresis is better than platelet concentrate.