Research progress of DNA methylation in idiopathic male infertility / 中国医师杂志

Male infertility is a multifactorial disease, of which the cause of male infertility cannot be determined, which is called idiopathic male infertility, and the incidence rate is gradually rising. Because its cause is unknown, it has become a major problem in the department of reproductive endocrinology. With the in-depth study of epigenetics, the diagnosis and treatment of idiopathic male infertility also has a new direction, especially the important role of DNA methylation in spermatogenesis and embryonic development. More and more gene fragments and loci have been found by scholars, which makes it possible to achieve accurate identification and targeted treatment. This article reviews and summarizes the research progress of DNA methylation related to idiopathic male infertility in recent years, aiming to further elaborate the pathogenesis of idiopathic male infertility and provide new ideas for clinical diagnosis and treatment.

Will male semen quality improve with environmental quality? / 亚洲男科学杂志(英文版)

Wenzhou has improved its environmental quality because of comprehensive environmental remediation; nevertheless, the semen quality of infertile males remains unclear. This study determined whether better environmental quality improved semen quality in this region. We recorded semen quality data from 22 962 infertile males from January 2014 to November 2019 at the Center for Reproductive Health of The First Affiliated Hospital of Wenzhou Medical University (Wenzhou, China). Patients were predominantly 30-35 years old (33.1%) and workers (82.0%), with high school education or lower (77.6%); more than a half of the patients (52.6%) were Wenzhou household registration; and most patients (77.5%) had abnormal semen quality. Patients who were older than 40 years and workers, and those with Wenzhou household registration, had significantly worse semen quality (all P < 0.05). From 2014 to 2019, progressive sperm motility, total sperm motility, and semen volume showed increasing linear trends in all patients (P = 0.021, 0.030, and 0.005, respectively), yet normal sperm morphology showed a linearly decreasing trend (P = 0.046). Sensitivity analyses for subgroups yielded similar results. In conclusion, the improvement of environmental quality and better function of the accessory glands are associated with progressive sperm motility, total sperm motility, and semen volume. Normal sperm morphology is influenced by occupational exposures and personal lifestyle and does not improve with environmental quality.

A Inibição do Metabolismo da Glicose por miR-34a e miR-125b Protege contra a Morte Celular de Cardiomiócitos Causada por Hiperglicemia / Inhibiting Glucose Metabolism By miR-34a and miR-125b Protects Against Hyperglycemia-Induced Cardiomyocyte Cell Death

Resumo Fundamento: É sabido que a resistência à insulina e a hiperglicemia são causas patológicas importantes no desenvolvimento de cardiomiopatia diabética (CMD). Entretanto, seus mecanismos moleculares precisos na patogênese da CMD ainda não estão claros. Objetivos: Estudos recentes revelam que os microRNAs (miRNAs) desempenham papéis essenciais na patogênese da CMD. Este projeto tem o objetivo de determinar os papéis de miR-34a e miR-125b na morte celular de cardiomiócitos causada por hiperglicemia. Métodos: Cardiomiócitos primários de ratos foram isolados e expostos a concentrações de glicose normais e altas. A viabilidade das células foi medida utilizando-se o ensaio MTT. As expressões de miR-34a e miR-125b foram detectadas por qRT-PCR. Alvos potenciais de miR-34a e miR-125b foram previstos pelo www.Targetscan.org, e validados a partir de tecidos cardíacos humanos. Um p<0,05 foi considerado significância estatística. Resultados: Demonstra-se neste estudo que o miR-34a e o miR-125b têm resposta celular reduzida no coração humano diabético. Além disso, os dados in vitro de cardiomiócitos primários de ratos demonstraram que o tratamento com glicose alta em curto prazo estimula a expressão de miR-34a e miR-125b. Demonstrou-se que, em condições de glicose alta, os cardiomiócitos de ratos apresentaram metabolismo de glicose intracelular, e a captação de glicose e a produção de lactato aumentaram significativamente. Foi identificado que as principais enzimas metabólicas da glicose, hexoquinase 2 (HK2) e lactato desidrogenase-A (LDHA) eram alvos diretos de miR-125b e miR-34a, respectivamente. A superexpressão de miR-125b e miR-34a poderia evitar a morte de celular de cardiomiócitos causada por hiperglicemia. Por fim, a recuperação de HK2 e LDHA em cardiomiócitos com superexpressão de miR-125b e miR-34a restaurou a sensibilidade de cardiomiócitos à hiperglicemia. Conclusões: Nossos resultados propõem um mecanismo molecular para proteção cardiovascular diabética mediada por microRNA e contribuirão para o desenvolvimento de estratégias de tratamento de disfunção cardiovascular associada a diabetes.

Abstract Background: It is well-known that insulin resistance and hyperglycemia are important pathological causes for the development of diabetic cardiomyopathy (DCM). However, its precise molecular mechanisms in the pathogenesis of DCM remain unclear. Objectives: Recent studies reveal that microRNAs (miRNA) play essential roles in the pathogenesis of DCM. This project aimed to determine the roles of miR-34a and miR-125b in hyperglycemia-induced cardiomyocyte cell death. Methods: Rat primary cardiomyocytes were isolated and exposed to normal and high concentrations of glucose. Cell viability was measured using MTT assay. Expressions of miR-34a and miR-125b were detected by qRT-PCR. Potential targets of miR-34a and miR-125b were predicted from www.Targetscan.org and validated from human heart tissues. A statistical significance of p<0.05 was considered. Results: The present study shows that miR-34a and miR-125b are downregulated in a human diabetic heart. Moreover, in vitro data from rat primary cardiomyocytes showed that short-term high glucose treatment stimulates miR-34a and miR-125b expressions. Under high glucose, it was found that rat cardiomyocytes displayed increased intracellular glucose metabolism, and glucose uptake and lactate production were significantly increased. It was also found that the key glucose metabolic enzymes, Hexokinase 2 (HK2) and Lactate dehydrogenase-A (LDHA), were direct targets of miR-125b and miR-34a, respectively. Overexpression of miR-125b and miR-34a could prevent hyperglycemia-induced cardiomyocyte cell death. Finally, the restoration of HK2 and LDHA in miR-125b and miR-34a overexpressed cardiomyocytes recovered the cardiomyocytes' sensitivity to hyperglycemia. Conclusion: Our results proposed a molecular mechanism for the microRNA-mediated diabetic cardiovascular protection and will contribute to developing treatment strategies for diabetes-associated cardiovascular dysfunction.

Effect of hot or warm property on skin toxicity of essential oil as penetration enhancer and its mechanism / 中国中药杂志

To compare the effect of hot or warm property of Chinese medicine(CM) on the skin toxicity of essential oils(EOs) as penetration enhancer in vitro and in vivo, and explore the mechanism. EOs were extracted from WIM of Bichengqie(Litseae Fructus), Dingxiang(Flos Syzygii Aromatici), Huajiao(Pericarpium Zanthoxyli Bungeani), and Xiaohuixiang(Fructus Foeniculi) with warm property, and Ganjiang(Rhizoma Zingiberis), Gaoliangjiang(Rhizoma Alpiniae Officinari), Hujiao(Fructus Piperis), and Wuzhuyu(Fructus Evodiae Rutaecarpae) with hot property, respectively. Then the in vitro toxicity was evaluated by human keratinocyte cytotoxicity. In vivo skin irritation potency was also evaluated through pathological observation after topical administration. The components, especially those located in stratum corneum, were analyzed by GC-MS. The main components, namely monoterpenes and sesquiterpenes, of EOs extracted from CM with hot property,were detected for the interaction with keratino-lipid ceramide 3 by molecular simulation technology; and the interaction energy value was calculated based on the optimal conformation. It was found that the skin cell toxicity of EOs from CM with hot property was significantly higher than that of EOs from CM with warm property. However, there was no significant difference between them by in vivo skin irritation evaluation. Whether from CM with hot property or warm property, EOs showed a significant reduced toxicity compared with azone. Sesquiterpenes(33.56%±19.38%) were found to be one of the main components in EOs from CM with hot property, while almost no sesquiterpenes was found in EOs from CM with warm property. After topical administration of EOs from CM with hot property, sesquiterpenes were demonstrated to be prone to locate in stratum corneum. The results of molecular simulation also revealed that the interaction between sesquiterpenes and ceramide 3 was significantly stronger than that of monoterpenes(P<0.01). In conclusion, the location of sesquiterpenes in stratum corneum resulted in the significant difference between in vitro skin cell toxicity and in vivo skin irritation potency. The EOs from CM with hot property shall be taken into account for further development of potent penetration enhancer.

Use of contrast-enhanced ultrasound in evaluating the efficacy and application value of microwave ablation for adenomyosis

Aim: This study aims to assess the use of contrast-enhanced ultrasonography (CEUS) and dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) in the evaluation of percutaneous microwave ablation (PMWA) of localized adenomyosis. Materials and Methods: Sixty-six patients with single-onset adenomyosis who underwent PMWA at the Liaocheng Tumor Hospital of Shandong Province from January 2013 to February 2019 were enrolled. Venous CEUS and DCE-MRI examinations were performed before and 1-2 days after the surgery. The ablation rates calculated by CEUS and DCE-MRI were compared and analyzed for accuracy. Results: After microwave ablation (MWA), CEUS showed that the volume and ablation rate of the ablated zone were 52.03 ± 28.39 cm3 and 90.90% ±6.61%, respectively. By DCE-MRI, the ablation volume and ablation rate of adenomyosis were 52.20 ± 28.65 cm3 and 90.88% ±6.32%, respectively. Dysmenorrhea was significantly relieved within 3 months of the operation, and nonmenstrual hemoglobin levels were significantly improved at 3 and 6 months after the operation (P < 0.05). All 66 cases of adenomyosis were treated using PMWA. Postoperatively, 17 patients reported a change in vaginal fluid; however, no special treatment was required as this disappeared 2-11 days after surgery. Conclusions: CEUS can accurately evaluate the ablation rate of localized adenomyosis treated with MWA, which is consistent with DCE-MRI. It is convenient and easy to perform ablation of adenomyomas, with incomplete ablation and angiography, and is a method worthy of clinical promotion

Pharmacological network-based study on interventional mechanism of Gu-Chang-Zhi-Xie pills for treatment of irritable bowel syndrome / 药学学报

This study was designed to explore the interventional mechanism involving "multi-components, multi-targets and multi-pathways" of Gu-Chang-Zhi-Xie pills (GCZX) for treatment of irritable bowel syndrome (IBS) using pharmacological network technology. Firstly, 96 active ingredients from GCZX pills were screened by ADME parameter filtration and chemical space principal component analysis, and the targets of anti-IBS function were predicted using PharmMapper online database. Secondly, AutoDock Vina was used to validate the docking between the active ingredients and predicted disease targets, and to establish the corresponding relationship between "pharmacodynamic molecules and target proteins". Finally, the target elements were mapped into the KEGG biological pathway by CluoGO plug-in, which further elucidates the potential relationship between the key targets and the mechanism of action of Gu-Chang-Zhi-Xie pills for treatment of IBS. The results showed that most of the top 11 key pharmacodynamic molecules were isoquinoline alkaloids, which mainly acted on inflammatory or pain targets, with different degrees of anti-inflammatory and analgesic effects. A total of 39 key targets were identified, including TPH1, TNF-α, IL-6, IFN-γ, MAO-A and IL-10. These targets were mapped to 29 KEGG pathways, of which the P-value of 5-HT signaling pathway was the smallest. Therefore, the pharmacodynamic molecules mainly act on 6 core targets and may play a major role in the regulation of 5-HT signal synthesis or transport pathway. This study sets an example for drug development and mechanistic investigation using innovative technology.

Research progress of anti-tumor mechanism of effective components of angelica dahurica / 国际中医中药杂志

Angelica dahurica is diaphoretic, commonly used in the treatment of cold, wet, and itchy rubella nasosinusitis. Studies have shown that the effective components of Angelica dahurica can be used in the treatment of malignant tumors. This paper summarizes the related literature home and abroad recently, and regards the effective components of Angelica dahurica as a major role in treatment of tumor by inhibiting tumor cell proliferation, promoting apoptosis, inhibiting tumor metastasis, inhibiting platelet aggregation, enhancing immunity, and anti-tumor drug resistance. Meanwhile, the paper finds the shortcomings of the present researches, and hopes to provide reference for the future experiment and clinical research.

Vacuum sealing drainage with anterolateral thigh perforator flap for repair foot soft tissue defect / 中国骨伤

<p><b>OBJECTIVE</b>To investigate clinical effect of vacuum sealing drainage(VSD) with anterolateral thigh perforator flap for repair foot soft tissue defect.</p><p><b>METHODS</b>From September 2014 to September 2016, 45 patients with foot soft tissue defect repaired by VSD with anterolateral thigh perforator flap, including 31 males and 14 females aged from 17 to 55 years with an average of 36 years old. Thirty patients were dorsal soft tissue defects, and 15 patients were plantar soft tissue defects. Sizes of soft tissue defect ranged from 7 cm×10 cm to 11 cm×18 cm, the wound was contaminated seriously and assisted with deep soft tissue exposure. Emergency surgical wound debridement with VSD treatment were performed, and wound surface was cleaned after 5 to 7 days, free anterolateral thigh flap were repaired. Sizes of flap ranged from 9 cm×12 cm to 13 cm×21 cm. Appropriate method was used to repair flap donor area.</p><p><b>RESULTS</b>All 45 free propeller flap were survived without skin edge necrosis, ulcerative infection. Forty patients were followed up from 3 to 24 months with an average of 13 months. Flap showed good color, beautiful appearance, good elasticity, soft texture, but without obvious bloated, the wood of leg healed well.</p><p><b>CONCLUSIONS</b>VSD with anterolateral thigh perforator flap for repair foot soft tissue defect could effective control wound soft tissue infection, receive reliable flap blood supply and skin flap could repaired at stage I. It is an effective method for repairing foot soft tissue defects.</p>

Establishment of a Novel Method for Screening Epidermal Growth Factor Receptor Tyrosine Kinase Inhibitor Resistance Mutations in Lung Cancer / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Drug resistance to targeted therapies occurs in lung cancer, and resistance mechanisms related to epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) are continuously being discovered. We aimed to establish a novel method for highly parallel multiplexed detection of genetic mutations related to EGFR TKI-resistant lung cancer using Agena iPLEX chemistry and matrix-assisted laser desorption ionization time-of-flight analysis on the MassARRAY mass spectrometry platform.</p><p><b>METHODS</b>A review of the literature revealed 60 mutation hotspots in seven target genes (EGFR, KRAS, PIK3CA, BRAF, ERBB2, NRAS, and BIM) that are closely related to EGFR TKI resistance to lung cancer. A total of 183 primers comprised 61 paired forward and reverse amplification primers, and 61 matched extension primers were designed using Assay Design Software. The detection method was established by analyzing nine cell lines, and by comparison with LungCarta™ kit in ten lung cancer specimens. EGFR, KRAS, and BIM genes in all cell lines and clinical samples were subjected to Sanger sequencing for confirming reproducibility.</p><p><b>RESULTS</b>Our data showed that designed panel was a high-throughput and robust tool, allowing genotyping for sixty hotspots in the same run. Moreover, it made efficient use of patient diagnostic samples for a more accurate EGFR TKIs resistance analysis. The proposed method could accurately detect mutations in lung cancer cell lines and clinical specimens, consistent with those obtained by the LungCarta™ kit and Sanger sequencing. We also established a method for detection of large-fragment deletions based on single-base extension technology of MassARRAY platform.</p><p><b>CONCLUSIONS</b>We established an effective method for high-throughput detection of genetic mutations related to EGFR TKI resistance based on the MassARRAY platform, which could provide more accurate information for overcoming cancers with de novo or acquired resistance to EGFR-targeted therapies.</p>

Detecting the spectrum of multigene mutations in non-small cell lung cancer by Snapshot assay / 癌症

As molecular targets continue to be identified and more targeted inhibitors are developed for personalized treatment of non-small cell lung cancer (NSCLC), multigene mutation determination will be needed for routine oncology practice and for clinical trials. In this study, we evaluated the sensitivity and specificity of multigene mutation testing by using the Snapshot assay in NSCLC. We retrospectively reviewed a cohort of 110 consecutive NSCLC specimens for which epidermal growth factor receptor (EGFR) mutation testing was performed between November 2011 and December 2011 using Sanger sequencing. Using the Snapshot assay, mutation statuses were detected for EGFR, Kirsten rate sarcoma viral oncogene homolog (KRAS), phosphoinositide-3-kinase catalytic alpha polypeptide (PIK3CA), v-Raf murine sarcoma viral oncogene homolog B1 (BRAF), v-ras neuroblastoma viral oncogene homolog (NRAS), dual specificity mitogen activated protein kinase kinase 1 (MEK1), phosphatase and tensin homolog (PTEN), and human epidermal growth factor receptor 2 (HER2) in patient specimens and cell line DNA. Snapshot data were compared to Sanger sequencing data. Of the 110 samples, 51 (46.4%) harbored at least one mutation. The mutation frequency in adenocarcinoma specimens was 55.6%, and the frequencies of EGFR, KRAS, PIK3CA, PTEN, and MEK1 mutations were 35.5%, 9.1%, 3.6%, 0.9%, and 0.9%, respectively. No mutation was found in the HER2, NRAS, or BRAF genes. Three of the 51 mutant samples harbored double mutations: two PIK3CA mutations coexisted with KRAS or EGFR mutations, and another KRAS mutation coexisted with a PTEN mutation. Among the 110 samples, 47 were surgical specimens, 60 were biopsy specimens, and 3 were cytological specimens; the corresponding mutation frequencies were 51.1%, 41.7%, and 66.7%, respectively (P = 0.532). Compared to Sanger sequencing, Snapshot specificity was 98.4% and sensitivity was 100% (positive predictive value, 97.9%; negative predictive value, 100%). The Snapshot assay is a sensitive and easily customized assay for multigene mutation testing in clinical practice.

Impact of smoking status and pathologic type on epidermal growth factor receptor mutations in lung cancer / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Epidermal growth factor receptor (EGFR) mutations in lung carcinomas can make the disease more responsive to the treatment with tyrosine kinase inhibitors. We aimed to evaluate the prevalence of EGFR mutations in a large series of lung carcinomas.</p><p><b>METHODS</b>We examined 1195 consecutive lung cancer patients for EGFR mutations in exons 18, 19, and 21 using direct sequencing of polymerase chain reaction products. A detailed smoking history was obtained. Patients were categorized as never smokers (< 100 lifetime cigarettes), former smokers (quit > 1 year ago), or current smokers (quit < 1 year ago).</p><p><b>RESULTS</b>There were EGFR mutations in 9 (4.5%) of 201 squamous carcinomas, in 1 (2%) of 50 large cell carcinomas, and in 1 (2.3%) of 44 small cell carcinomas that were investigated. Three hundred and twenty-seven mutations were found in the series of 858 adenocarcinomas (38.1%). Among 858 lung adenocarcinomas, we detected EGFR mutations in 250 (48.6%) of 514 never smokers, 39 (33.9%) of 115 former smokers, and 38 (16.6%) of 229 current smokers. Significantly fewer EGFR mutations were found in people who smoked for more than 15 pack-years (P = 0.0002) or stopped smoking less than 15 years ago (P = 0.033) compared with individuals who never smoked.</p><p><b>CONCLUSIONS</b>Adenocarcinoma is the most frequent EGFR mutation pathologic type in lung cancer. The likelihood of EGFR mutations in exons 18, 19 and 21 decreases as the number of pack-years increases. Mutations were less common in people who smoked for more than 15 pack-years or who stopped smoking cigarettes less than 15 years ago. These data can assist clinicians in assessing the likelihood of exons 18, 19, or 21 EGFR mutations in Chinese patients with lung cancer when mutational analysis is not feasible.</p>

Isolation, culture and identification of rabbit bone marrow-derived vascular endothelial progenitor cells / 中国实验血液学杂志

The aim of study was to set up a suitable method of isolation, culture and identification of endothelial progenitor cells (EPC) derived from rabbit bone marrow. Density gradient centrifugation was used to isolate mononuclear cells from bone marrow, the isolated mononuclear cells were cultured with specific culture medium for EPCs. EPCs were identified by cellular morphologic observation, immunohistochemistry testing, flow cytometry and the function test of taking up Dil-ac-LDL and FITC-UEA-1. The results indicated that the newly isolated bone marrow-derived mononuclear cells exhibited a round appearance, following culture for 48 hours, adherent cells grew in colony cluster, presenting with round or irregular appearance, and nuclear division was obvious. On day 7, flaky cell colonies mutually connected together, presenting with spindle-shaped cells. Immunohistochemistry testing in the EPCs showed CD133(+), CD34(+), VIII factor(++), KDR(++); flow cytometry testing showed that the positive rate of CD133 was (18.23+/-7.12)%, the positive rate of CD34 was 47.71+/-14.85%, the positive rate of CD31 was (71.61+/-13.51)%, the positive rate of KDR was (87.24+/-11.40)%. And more than 80% EPC could take up both Dil-acLDL and FITC-UEA-1. It is concluded that the mononuclear cells isolated from bone marrow by density gradient centrifugation can differentiate into EPCs under special culture situation.

Mutation of plakophilin-2 gene in arrhythmogenic right ventricular cardiomyopathy / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Arrhythmogenic right ventricular cardiomyopathy (ARVC) is one of the leading causes of sudden cardiac death. Recent studies have shown that ARVC, which is an inheritable genetic change, results from mutations in genes encoding desmosomal proteins. Plakophilin-2 is an important component of the desmosome. Because the full range of genetic variations related to ARVC is unknown and no related studies of the Chinese population have been reported, we aimed to investigate the genetic variation of plakophilin-2 in ARVC patients from the Southern Region of China.</p><p><b>METHODS</b>Genomic DNA was isolated from peripheral blood samples of all 34 ARVC patients, who were screened through a clinical evaluation. They were used to detect variations in the sequences of the plakophilin-2 genes by polymerase chain reaction amplification in combination with direct sequencing.</p><p><b>RESULTS</b>In exon-1 of the plakophilin-2 gene, a deletion mutation (c.145_148 del GACA) was found in one family pedigree. The mutation was also found in exon-2, 4, and 11 of the plakophilin-2 gene. The QT interval dispersion of the ECG was considerably longer in the mutation group than in the non-mutation group of ARVC patients, and this result was statistically significant (P < 0.05).</p><p><b>CONCLUSION</b>We discovered a plakophilin-2 mutation that prolongs the QT interval dispersion in the southern Chinese ARVC population.</p>

Predictors of mortality in critically multiple trauma patients after damage control surgery / 中华外科杂志

<p><b>OBJECTIVE</b>To investigate the efficiency of damage control surgery (DCS) and predictors of mortality in critically multiple trauma patients.</p><p><b>METHODS</b>From May 1998 to February 2007, DCS were carried out in 27 patients with critically multiple trauma. Of the patients 15 cases survived (survival group) and 12 cases died (dead group). The surgical complications, causes of death, demographic, physiologic and medical parameters were collected and compared between the two groups. Multiple logistic regression analysis were performed to identify possible predictors of mortality.</p><p><b>RESULTS</b>The incidence of surgical complications was 37.0 percent, and the intra-abdominal infections was the most frequent (18.5%). The overall mortality rate was 44.4 percent. The most common causes of death was multiple organ dysfunction syndrome (50.0%). With respect to predicting mortality, statistically significant differences was found in parameters as age, injury severity score (ISS), initial temperature and base excess (BE), estimated blood loss, initial ICU temperature and length of hospital stay. Older age, increased absolute value of initial BE and lower initial ICU temperature were determined as independent predictors of mortality on multiple logistic regression analysis.</p><p><b>CONCLUSIONS</b>There is a comparable high morbidity and mortality rate in severely injured patients managed with DCS. Increased age, a larger absolute value of initial BE and lower initial ICU temperature could independently predict death of the patients.</p>

Construction and expression of RU486-inducible eukaryotic vector carrying red fluorescent protein / 南方医科大学学报

<p><b>OBJECTIVE</b>To construct an inducible eukaryotic vector carrying red fluorescent protein (DsRed) and evaluate the regulation of DsRed gene expression in vitro.</p><p><b>METHODS</b>The vector pRS17-RUDsRed containing DsRed gene, promoter and RU486-inducible system was constructed using molecular biological methods. To minimize potential interference, the two transcriptional elements were spaced with a 1.6 kb insulator. Fluorescence microscopy and flow cytometry were used to observe the activation of this regulatable vector after transfection in MFC cells.</p><p><b>RESULTS</b>The vector was identified by digestion with different restriction enzymes, sequencing and PCR. In the absence of RU486, the cells transfected with the vector exhibited very low DsRed protein expression, and the addition of RU486 induced efficient DsRed expression in the cells.</p><p><b>CONCLUSION</b>The RU486-inducible eukaryotic vector carrying DsRed protein allows effective regulation of the target gene expression in vitro, which provides a useful tool for gene regulation and gene therapy studies.</p>

Effect of intra-bone marrow infusion of allogeneic mesenchymal stem cells on reconstruction of marrow mesenchymal stem cells in rat HSCT models / 中国实验血液学杂志

This study was aimed to investigate the effect of intra-bone marrow (IBM) injection of allogeneic mesenchymal stem cells (MSCs) on reconstruction of bone marrow MSCs (BM-MSCs) in rats that received hematopoietic stem cell transplantation (HSCT), and to detect the donor MSCs in the hosts for clarifying the effect mechanism of donor MSCs. Wistar female rats conditioned with lethal dose 60Co gamma-ray irradiation were co-transplanted with F344 female fetal and neonatal peripheral blood (FNPB) and BrdU-labeled MSCs separated from bone marrow mononuclear cells of F344 male rats. The donor MSCs were infused by IBM injection in bilateral tibia or intravenous injection (IV), while the FNPB were all via IBM route. The survival rate, engraftment level of HSCs and recovery of BM-MSCs of recipients were monitored. The ratio of BrdU-labeled MSCs in recipient rats was calculated by immunofluorescence assay (IFA) and the Y chromosomes were examined by PCR. The results showed that the recipient rats of the two co-transplantation groups were all alive at day 60 after transplantation. There was no significant difference between these two groups on the survival rates or the engraftment levels of HSCs, but each of them was much better than that of the FNPB group. At day 30 after transplantation, the proliferation ability of recipients' BM-MSCs was still below normal, while that of the FNPB (IBM)+MSC (IBM) group was the best of all the experiment groups (p<0.01). At 60 days, the donor MSCs coexisted with host MSCs in only a few recipient rats examined by IFA, while the Y chromosomes could be detected in all the recipient rats in the two cotransplantation groups. It is concluded that the infusion of allogeneic MSCs can accelerate the recovery of HSCT recipients' BM-MSCs. The IBM route is safe and more effective than intravenous infusion.

Effect of free hemoglobin on hemorrhagic shock in pigs: TNF-alpha, IL-6 expressions in serum and rates of MODS after the blood transfusion / 中国实验血液学杂志

This study was aimed to investigate the expressions of tumor necrosis factor-alpha (TNF-alpha), Interleukin-6 (IL-6) in serum and the incidence of multiple organ dysfunction syndrome (MODS) in pigs with hemorrhagic shock after the blood transfusion simultaneously combined with different doses of free hemoglobin (FHb) so as to provide guidance of banked blood with high concentration of FHb during war injury through understanding effect of FHb on the animals. The different doses of FHb were given intravenously during the recovery of pig from shock, the vital signs and functional changes of vital organs were monitored and the incidence of MODS was determined, as well as the serum specimens were collected and the TNF-alpha, IL-6 levels in serum were detected by ELISA. The results showed that there were statistical differences of serum levels of TNF-alpha and IL-6 in pigs after FHb 10 mg/kg infusion, as compared to shock control group. There was significantly difference of the serum levels of TNF-alpha, IL-6 after FHb 15 mg/kg infusion, compared to the control group. The incidence of MODS increased significantly. It is concluded that the blood infusion containing high dose (more than 10 mg/kg) of FHb influences significantly on the cytokines in pigs with hemorrhagic shock, and increases damage of cytokines to vital organs and the incidence of MODS. The tolerance dose of the pigs to free hemoglobin is about 10 mg/kg or so. The infusion of blood with less than 10 mg/kg is relatively safe for pig in hemorrhagic shock.

Clinical management of abdominal trauma / 中华创伤杂志(英文版)

<p><b>OBJECTIVE</b>To improve the prognosis of patients with abdominal trauma.</p><p><b>METHODS</b>Between January 1993 and December 2005, 415 patients were enrolled in this research. The patients consisted of 347 males and 68 females with mean age of 36 years (ranging from 3-82 years). All abdominal traumas consisted of closed traumas (360 cases, 86.7%) and open traumas (55 cases, 13.3%).</p><p><b>RESULTS</b>A total of 407 cases (98.1%) were fully recovered from trauma and the other 8 cases (1.9%) died of multiple injuries. The mean injury severity score (ISS) of all patients was 22 while the mean ISS of the patients who died in hospital was 42. Postoperative complications were seen in 9 patients such as infection of incisional wounds (6 cases), pancreatic fistula (2 cases) and intestinal fistula (1 case). All these postoperative complications were cured by the conservative treatment.</p><p><b>CONCLUSION</b>Careful case history inquisition and physical examination are the basic methods to diagnose abdominal trauma. Focused abdominal ultrasonography is always the initial imaging examination because it is non-invasive and can be performed repeatedly with high accuracy. The doctors should consider the severity of local injuries and the general status of patients during the assessment of abdominal trauma. The principle of treatment is to save lives at first, then to cure the injuries. Unnecessary laparotomy should be avoided to reduce additional surgical trauma.</p>

The cytotoxicity of CIK/NK cells stimulated by K562-DC fusion vaccines in NOD/SCID mice model for human erythroleukemia / 中华血液学杂志

<p><b>OBJECTIVES</b>To study the in vivo efficacy and the safety of cord blood derived CIK/NK cells stimulated by K562-dendritic cells (DC) fusion vaccines in NOD/SCID mice model for human erythroleukemia.</p><p><b>METHODS</b>DC and CIK /NK cells were both derived from cord blood mononuclear cells. DC were fused with inactivated K562 leukemia cell by PEG to produce K562-DC fusion vaccines. K562-DC fusion vaccines were co-cultured with CIK/NK cells to prepare K562-DC fusion vaccine stimulated CIK/NK cells. NOD/SCID mice were inoculated with 1 x 10(6) K562 cells. 24 hours later, 1 x 10(7) vaccines stimulated CIK/ NK cells and 1 x 10(7) CIK/NK cells were transfused into the NOD/SCID mice. NOD/SCID mice without inoculation of K562 cells were used as control group. CD13 and CD56 positive cells were assayed by flow cytometry.</p><p><b>RESULTS</b>All the leukemia NOD/SCID mice without therapy died within 39 days, tumor was found in 5 of 8 mice. One of 8 leukemia mice treated with K562-DC fusion vaccines stimulated CIK/NK cells died at the 65th day, the anti-tumor response rate was 87.5%. Two of the leukemia mice treated with CIK/NK cells died at the 56th and 65th day respectively, the anti-tumor response rate was 75%. There was no significant difference in survival time between these two groups, and both survivals were longer than that of the control group. There was no significant difference in CD13 positive cells in the survival mice between these two groups, and both of that were less than that of the control mice. There was no significant difference in CD56 positive cells between the two treated groups and the control group.</p><p><b>CONCLUSIONS</b>Cord blood derived CIK/ NK cells stimulated by inactivated tumor cells retain the cytotoxicity and do not develop tumor in vivo.</p>

Isolation and culture of human embryonic AGM derived HSPCs in hematopoietic culture systems created by AGM stromal cells / 中国实验血液学杂志

This study was purposed to isolate human embryonic AGM derived HSPCs and investigate the effect of AGM stromal cells on AGM-derived HSPCs. Immunohistochemical sections of human AGM tissue were investigated for CD34, Flk-1 and VEGF expression. Human AGM-derived single cells were isolated and seeded onto pre-treated feeder of human AGM stromal cells (hAGMS3 and hAGMS4) by direct contact and non-contact co-culture in Transwell culture system. Growth characteristics of HSPCs with cobblestone area-forming cells (CAFCs) were observed and number of cobblestone area (CA) was counted. Indirect immunofluorescent assay was used to detect CD34 and Flk-1 expression on the surface of suspended cells as well as CAFCs in contact co-culture system. The cells after culture for 2 weeks were collected from both contact and non-contact co-culture systems for CFU assay. The result showed that hematopoietic cells in AGM tissue expressed CD34 and Flk-1. Both of the hematopoietic culture systems could produce CFCs. Nevertheless, direct contact co-culture produced CD34(+)Flk-1(+) CAFC and more CFUs than those from indirect non-contact culture (hAGMS3 system: 1647 +/- 194 vs 389 +/- 31, p < 0.05; hAGMS4 system: 1586 +/- 75 vs 432 +/- 35, p < 0.05). It is concluded that there were CD34(+)Flk-1(+) HSCs in human embryonic AGM region. The hematopoietic co-culture systems composed of AGM-derived HSPCs and AGM stromal cells are successfully established, both direct contact and Transwell non-contact co-culture can expand AGM-derived definitive HSPCs. Cell-cell contact between AGM-derived HSPCs and AGM stromal cells are of most importance to maintain and expand AGM-HSPCs.