Second-trimester serum IL-1, IL-6, IL-8, TNF-a, CRH levels in prediction of preterm labor / 上海预防医学

[Objective]To determine the relationship between maternal serum IL-1, IL-6, IL-8, TNF-a and CRH levels and preterm labor in asymptomatic women at 24 to 28 weeks' gestation. [Methods] We performed a prospective cohort study from January 2015 to December 2017 for asymptomatic women at 24 to28 weeks' gestation and maternal serum. According to the pregnancy outcome, 100 cases of preterm labor were selected as experimental group and another 100 cases of term labor as control group. The levels of serum IL-1, IL-6, IL-8, TNF-a and CRH were measured by ELISA. The partial membranes of all cases were examined pathologically after labor. [Results] The levels of serum IL-1, IL-6 and IL-8 in preterm labor group were significantly higher than those in the control group (P <0.05). The levels of serum IL-6and IL-8 in preterm labor with chorioamnionitis were significantly higher than those without chorioamnionitis and term labor (P < 0.05). The level of IL-1 in preterm labor without chorioamnionitis was significantly higher than that of term labor (P<0.05). The levels of serum TNF-a and CRH had no difference between the groups (P>0.05). Receiver-operating characteristic curves demonstrated that serum IL-1, IL-6 and IL-8 predicted preterm labor. The area under the curve (AUC) of IL-1 was 0.843. The AUC of IL-6 was0.675 and IL-8 was 0.55. [Conclusion] Maternal serum IL-1, IL-6 and IL-8 in asymptomatic women at24 to 28 weeks' gestation are useful markers for prediction of preterm labor.

Preparation of DJ-1 overexpressing cell model by using lentiviral vector / 基础医学与临床

Objective To prepare gene overexpressing cell model of human wild-type DJ-1 and its L166P mutant, and to investigate the role of lentiviral vector in gene overexpressing cell model .Methods Wild type DJ-1 and L166P mutant DJ-1 lentiviral vector plasmids were respectively constructed .After sequencing and comparing cor-rectly, the plasmid was amplified and transfected into HEK 293T cell line.Expression of WT DJ-1 and L166P mu-tant DJ-1 in cell lines was detected by fluorescence and Western blot .After determining the accurate expression of the target protein, a large amount of HEK293T cells was transfected and packaged to produce lentiviral particles. The PC12 cells were infected with the titer of virus supernatant.The fluorescence intensity of GFP and the expres-sion of target protein were observed by fluorescence microscope and Western blot method ,and the infection effi-ciency of the virus was determined .Results Lentiviral vectors carrying wild type DJ-1 and its mutants were suc-cessfully constructed .The virus vector can be transfected into HEK 293T cells and the target protein can be correctly expressed.The viral titers of LV-DJ-1 and LV-DJ-1/L166P were 2×109 TU/mL and 2×108 TU/mL, respectively. Virus supernatant can efficiently infect PC 12 cells, and most cells can express target proteins .The protein expres-sions of exogenous wild-type DJ-1 and L166P mutants were 315% and 285% of endogenous content ,respectively. Conclusions Lentivirus vector can infect cells efficiently , and it is a good way to prepare gene over expressing cell model.A cell model overexpressing DJ-1 or its L166P mutant is successfully prepared .The model can be used for subsequent DJ-1 function research .

Xuefu zhuyu oral liquid intervened stress-stimulated depression model rats / 中国中西医结合杂志

<p><b>OBJECTIVE</b>To observe effects of xuefu zhuyu Oral Liquid (XZOL) on the brain behavior and monoamine neurotransmitter 5-HT, and brain derived neurotrophic factor (BDNF) content on depression model rats.</p><p><b>METHODS</b>Male SD rats were randomly divided into the control group, the model group, the XZOL group, and the Deanxit Tablet group, 12 in each group. The depressive rat model was established by chronic unpredictable mild stress method. XZOL was administered to rats in the XZOL group by gastro-gavage, while Deanxit Tablet was given to those in the Deanxit Tablet group by gastrogavage. The intervention lasted for two weeks. The behavioral changes were observed by sucrose water consumption test and open-field test. The 5-HT and BDNF contents were detected using ELISA.</p><p><b>RESULTS</b>After chronic stress stimulus, experimental rats in the model group might have abnormal behavioral changes and lowered 5-HT content, showing statistical difference when compared with the control group (P <0.01). No obvious change in stimulated rats' behavior after intervention of XZOL and Deanxit Tablet. 5-HT content was not obviously reduced (P>0.05). Besides, XZOL was superior to Deanxit Tablet in increasing the 5-HT content (P<0.05). But the brain BDNF level of rats in the model group was not statistically different from that of rats in the model group (P >0.05), while the brain BDNF level of rats in the XZOL group and the Deanxit Tablet group was lower than that of rats in the model group (P <0.01).</p><p><b>CONCLUSIONS</b>Stress can lead to behavioral changes and lowered 5-HT content of rats. The intervention of XZOL could fight against depression-induced behavioral changes and increase 5-HT content. But it did not significantly affect the brain BDNF level. We inferred that it might not effect through the BDNF pathway.</p>

SUDHL-4 cell culture in vitro and establishment of mouse tumor model / 中国实验血液学杂志

This study was designed to investigate the biological and immunological characteristics of a human diffuse large B-cell lymphoma (DLBCL) cell line SUDHL-4, and to establish a mouse model for human DLBCL. SUDHL-4 cells were cultured under different conditions. The morphology and in vitro expression of B-cell and tumor-related markers were detected by microscopy and flow cytometry respectively. To establish the transplanted tumor, the cells were injected subcutaneously into SCID mice. Tumor formation and its histomorphology were analyzed. The results showed that the expression of B cell/tumor-related markers was found on cultured SUDHL-4 cells. A stable mouse model of human DLBCL was successfully established in SCID mice by subcutaneous injection of 10(7) SUDHL-4 cells. Tumor tissue from mice exhibited similar histologic manifestation to those of human DLBCL. It is concluded that the SUDHL-4 cells represent a high consistency in immunological characteristics with human DLBCL. Transplantation of SUDHL-4 cells provides a syngeneic mouse model for the study of human DLBCL.

Clinical study on childhood acute lymphoblastic leukemia diagnosed and treated with 04 Protocol in Chongqing, China / 中华儿科杂志

<p><b>OBJECTIVE</b>To analyze the clinical and laboratory data from acute lymphoblastic leukemia (ALL) patients and the results of treatment using 04 Protocol (suggested by the Pediatric Hematology Group of Chinese Medical Association in 2004).</p><p><b>METHODS</b>This study included 88 children with ALL below the age of 18 years during the period from October 1, 2004 to June 30, 2007. Minimal inhibitory concentration (MIC) and clinical risk classification were done and the new chemotherapy regimen was used according to the protocol. Patients were stratified into low-risk (LR), medium-risk (MR), and high-risk (HR) groups. Life table method was used to estimate survival rate and statistical analysis was done by using software SPSS for Windows.</p><p><b>RESULTS</b>From October 2004 to June 2007, 88 childhood ALL patients were treated with the 04 Protocol. Sixty-three (91.30%) patients attained complete remission (CR) and 17 patients lost to follow up. The overall 4-year-event-free survival (EFS) rate (+/- SE) was (59.73 +/- 7.22)%. EFS was (75.60 +/- 9.71)% in the LR (n = 30), (65.50 +/- 11.69)% in the MR (n = 20) and (44.03 +/- 12.36)% in the HR. Relapse occurred in 18.18% of patients. Seven (7.95%) of 88 patients with ALL died during he induction therapy. Infection was the most common cause of death.</p><p><b>CONCLUSION</b>The outcome of patients treated with the 04 Protocol was favorable. Clinical risk classification and the leukemia cells of D19 are independent predictors of prognosis of ALL. High dose methotrexate played an important role in prevention and treatment of central nervous system leukemia. The mortality rate of this chemotherapeutic protocol during induction therapy was high.</p>

Astrocytes protect MN9D neuronal cells against rotenone-induced oxidative stress by a glutathione-dependent mechanism / 生理学报

Astrocytes maintain homeostasis of neuronal microenvironment, provide metabolic and trophic support to neurons and modulate neuronal responses to injury. Rotenone specifically inhibits mitochondrial complex I, and long exposure to rotenone may increase the risk for Parkinson's disease (PD) and cause Parkinsonism. However, little is known about the role of astrocytes in the process of rotenone-induced dopaminergic neuron injury. In order to investigate this issue, we used MN9D cells as a cell model of dopaminergic neurons and rotenone as a toxin to initiate mitochondrial deficiency. MN9D cells treated with the normal medium or astrocyte-conditioned medium (ACM) were exposed to different concentrations of rotenone for different time followed by cell viability measurement by MTT assay. Besides, various concentrations of ACM and temporally different treatments were devised to evaluate protective efficiency of ACM. Growth curve of cells in the normal medium or ACM was continuously assessed by cell counting for 8 d. The influence of rotenone and ACM on cellular oxidative stress was determined by DCFH-DA staining followed by flow cytometric analysis. Glutathione (GSH) content after treatment of ACM or rotenone was measured by GSH assay kit. Our results showed that rotenone decreased viability of MN9D cells in a dose-dependent manner and ACM treatment significantly attenuated rotenone toxicity at each concentration. No significant difference in growth rate was observed between the normal medium and ACM treatment. Four concentrations of ACM, namely 1/3ACM, 1/2ACM, 2/3ACM and pure ACM, all displayed protection, increasing cell viability to (124.15+/-0.79)%, (126.59+/-0.82) %, (125.84+/-0.61) % and (117.15+/-1.63) % of the cells exposed directly to rotenone, respectively. Treatment with ACM through the whole experiment except the initial 24 h, 24 h before or at the same time of rotenone addition all exerted protective effects, with cell viability being (110.11+/-2.52)%, (113.30+/-2.36) %, (114.42+/-2.00)% of the cells exposed directly to rotenone, respectively. Conversely, ACM treatment 12 h after rotenone addition had no protective effect, with cell viability being (102.54+/-1.36)% of the cells exposed directly to rotenone. Moreover, ACM treatment up-regulated GSH level in MN9D cells nearly twofold. Incubation with 100 nmol/L rotenone for 24 h depleted GSH level by nearly two thirds of the control, but ACM treatment mitigated the drop of GSH level, maintaining its content at (147.83+/-0.63)% of the control. Consistent with GSH change, rotenone administration resulted in a positive rate of 96.24% of DCF staining, implying a great extent of oxidative stress, whereas treatment with ACM reduced the extent of oxidative stress to a positive rate of 78.31%. Taken together, these findings suggest that astrocytes protect MN9D cells from oxidative stress caused by rotenone, and GSH partially accounts for the protection. Therefore, astrocytes may play a protective role in the process of PD.

The Influence of ?-Synuclein Overexpression on Mitochondrial Membrane Structure with Atomic Force Microscopy / 中国生物工程杂志

Objective:To identify the effect of ?-synuclein overexpression on mitochondrial membrane structure with atomic force microscopy. Methods:?-syn expression was mediated by AAV (adeno-associated viral vector) and Recombinant AAV/?-syn and AAV/LacZ viral particles were stereotaxically injected in the left side of rat substantia nigra (SN) for rat model of ?-synuclein overexpression. Mitochondria were isolated from rats SN of Brain. Mitochondria were analysis with JC-1 staining,atomic force microscopy and Western blot. Results:By 16 weeks post-infection of AAV-?-syn,the level of ?-syn increased about 2 times in mitochondrial fraction with Western blot and mitochondrial membrane potential (??) decreased with JC-1 staining. Furthermore,mitochondria swelling and porous like structure formed on the mitochondrial membrane with atomic force microscopy. Conclusion:The data suggested that ?-syn could accumulate in mitochondria,might form mitochondrial membrane pores and lead to ?? decreases. ?-syn might lead to mitochondrial dysfunction in Parkinson's disease.

Therapeutic Benefit of TH,AADC,and GCH-I Genes for Parkinson's Disease in Rat Model / 中国生物工程杂志

Parkinson's disease(PD) is a common neurodegenerative disorder with no effective protective treatment,characterized by a massive degeneration of dopaminergic neurons in the substantia nigra(SNpc) and the subsequent loss of their projecting nerve fibers in the striatum.The major neurochemical manifestation of this disorder is the loss of the neurotransmitter dopamine(DA) in the striatum as a result of the progressive degeneration of the dopaminergic neurons in the substantia nigra.There have been significant progresses in recent years reporting on the use of mesenchymal stem cells(MSCs)in gene therapy,with specific application towards PD.MSCs,a kind of multipotent adult progenitor cells,are considered as a useful vehicle for cell and gene therapy because of their multiple differentiation potentiality and self-transplantation.The present study was focused on treating rat model of PD using human tyrosine hydroxylase gene(hTH),human aromatic L-amino acid decarboxylase gene(hAADC) and human GT Pcyclohydrolase I gene(hGCH-I) engineered MSCs,in order to provide a better understanding about the application of these cells in the therapeutic benifit of PD.The gene of hTH,hAADC and hGCH-I were introduced via recombinant adeno-associated virus(rAAV) infection into the MSCs in vitro.The genetically modified MSCs expressing hTH,hAADC and hGCH-I were transplanted into the striatum of PD rat models.The behavior,the nigra-striatal level of DA and its metabolite were detected.The results of present study were shown as follows:hTH,hAADC,hGCH-I and LacZ gene were transfected into MSCs with adeno-associated virus vectors.The HEK293 packaging cells(ATCC) were transfected with the plasmids of pAAV-hTH,pAAV-hAADC,pAAV-hGCH-I,pAAV-LacZ,pAAV-RC,pHelper by using calcium phosphate precipitation.Titer was detected using HT1080 cells.Viral particles were collected and used to infect MSCs.The purified modified MSCs expressing the three kinds of genes were selected separately and were grafted in the striatum of the PD model rats in the lesion side.The MSCs genetically modified suvived well 12 weeks after transplantation.The improvements of the behavior were observed every week after transplantation.Compared with the control group,the rounds of asymmetric rotation after apomorphine administration decreased in the groups double or triple genes engineered MSCs grafted(p

Overexpression of ?-Synuclein Gene Caused Dopaminergic Neuron Damage in Substantia Nigra of Rats / 中国生物工程杂志

Objectiive:The present study was designed to explore whether overexpression of human wild ?-synuclein in rat brain caused selective dopaminergic neuron loss in substantia nigra and aimed to find out a new method to make a rat model of Parkinson's disease(PD).Methods:The human wild ?-synuclein gene was induced into the rat brain by Adeno-Associated Virus(AAV) vector.The overexpression of ?-synuclein was detected by realtime PCR.The behavior of rats were recorded every 4 weeks after the viral particle injection.TH immunohistochemistry were performed at 4,8,12 and 16 weeks post-injection as well as the dopamine(DA),3,4-dihydroxypheny-lacetic acid(DOPAC) of striatum were determined by high performance liquid chromatography coupled with electrochemical detection.Results:Realtime PCR results revealed a significant overexpression of ?-synuclein in the injected hemisphere.By 8 weeks post injection,a significant loss of the dopaminergic neurons was observed.34% of the dopaminergic neurons were lost after 12 weeks,and about 60% cells loss after 16 weeks.The DA and DOPAC levels in the striatum decreased about 15% 12 weeks after injecting viral particle carried ?-synuclein gene and 30% decreased after 16 weeks.The AAV-?-synuclein-treated rats developed a type of motor impairment,i.e.,head position bias,compatible with this magnitude of nigrostriatal damage.Conclusion:All the results showed that overexpression of human wild ?-synuclein caused selective dopaminergic neuron loss and mimic a symptom of human PD in rats.This may be a new methed to make rat PD model which can offer new opportunities for the study of pathogenetic mechanismsand exploration of new therapeutic targets of particular relevance to human PD.

Study on the Role of ?-synuclein in Mitochondria Dysfunction Caused by Small Dosage Rotenone / 中国生物工程杂志

Mitochondrial dysfunction has been implicated in the aetiology of sporadic Parkinson's disease but its role in the disease mechanism remains unclear.To investigate the effect of synuclein on mitochondrial dysfunction induced by rotenone.The human dopaminergic SH-SY5Y cells were used as a cell model.The cells over-expressed the wild-type ?-synuclein were treated with complex I inhibitor rotenone.The cell viability,complex I activity,Mitochondrial swelling and O2-content were tested at different time point-1w,2w,4w after rotenone treated.CCK-8 test results showed that the cell viability of overexpressed ?-synuclein(SH-SY5Y-Syn)was much lower than the control group(SH-SY5Y-Ctr).After administrating with rotenone about 1w or 2w the cell viability of SH-SY5Y-Syn became higher than that of SH-SY5Y-Ctr.On the 4th week the results were contrary to the first 2 weeks.Similar results were got when test the mitochondrial function.In the first 2 weeks after roteoone administrating,the mitochondrial function of SH-SY5Y-Syn was better than that of SH-SY5Y-Ctr.This suggest that the ?-synuclein could protect the mitochondrial against the injury induced by rotenone in the early stage-1w,2w,while this effect disappeared in the final stage-4w.

Overexpression of alpha-synuclein in SH-SY5Y cells partially protected against oxidative stress induced by rotenone / 生理学报

Both genetic and environmental factors are involved in the pathogenesis of Parkinsonos disease (PD). Epidemiological studies showed that environmental factors shared with the common mechanisms of resulting in alpha-synuclein aggregation by inhibiting complex I of mitochondria and leading to oxidative stress. To investigate the relationship between alpha-synuclein and oxidative stress, we used human dopaminergic SH-SY5Y cells transfected with alpha-synuclein-enhanced green fluorescent protein (EGFP). alpha-synuclein gene expression was determined by immunocytochemistry and real-time quantitative PCR. Both SH-SY5Y and alpha-synuclein overexpressed SH-SY5Y (SH-SY5Y/Syn) cells were treated with various concentrations of rotenone for different time. Cell viability and oxidative stress were detected by MTT assay and DCF assay. Superoxide dismutase (SOD) activity was assessed with xanthine peroxidase method. Cell apoptosis was detected with flow cytometry. Results showed that alpha-synuclein gene was constantly overexpressed in SH-SY5Y/Syn cells. After treatment with rotenone, both cell viability and complex I activity in these cells were reduced in a concentration-dependent manner. Oxidative stress was also found in these cells. Compared with SH-SY5Y cells, SOD activity in SH-SY5Y/Syn cells was increased distinctly (P<0.05) and alpha-synuclein significantly attenuated rotenone-induced cell apoptosis. These results suggest that the alpha-synuclein overexpression in SH-SY5Y cells has a tendency to partially resist oxidative stress induced by rotenone and this response may assist cell survival.

Effect of Tpo and/or IL-11 gene modified stromal cells on the expansion of CD34+ CD38- hematopoietic primitive progenitor cells / 中华血液学杂志

<p><b>OBJECTIVE</b>To investigate the effects of Tpo and/or IL-11 gene modified stromal cells on the expansion of CD(34)(+) hematopoietic stem/progenitor cells in cord blood.</p><p><b>METHODS</b>Retroviral vectors containing Tpo or IL-11 gene were constructed and used to transfect the stromal cell line HFCL. Tpo and/or IL-11 mRNA was assayed by Northern blot. Non-modified stromal cells were used, CD(34)(+) hematopoietic stem/progenitor cells from cord blood were expanded on gene-modified stromal cells for 7 days. The phenotype of CD(34)(+)CD(38)(-) primitive progenitors was detected by flow cytometry.</p><p><b>RESULTS</b>HFCL expressed Tpo and/or IL-11 mRNA after transfected by the retroviral vectors. The percentages of CD(34)(+)CD(38)(-) primitive progenitors in the cultures of Tpo, IL-11 and Tpo + IL-11 modified HFCL were (1.8 +/- 0.24)%, (1.62 +/- 0.23)%, and (2.45 +/- 0.28)%, respectively, which were higher than that in the control [(0.8 +/- 0.23)%].</p><p><b>CONCLUSION</b>The stromal cells modified by Tpo and/or IL-11 gene were able to enhance ex vivo expansion of CD(34)(+) and CD(34)(+)CD(38)(-) hematopoietic stem/progenitor cells from cord blood.</p>