[Use of the ETV6/RUNX1 probe to verify the performance of the fluorescence in situ hybridization probe before clinical detection].

Objective: To explore the standardized performance of a FISH probe before clinical detection. Methods: The probe sensitivity and specificity of ETV6/RUNX1 were analyzed via interphase and metaphase FISH in 20 discarded healthy bone marrow samples. The threshold system of the probe was established using an inverse beta distribution, and an interpretation standard was established. Finally, a parallel-controlled polymerase chain reaction detection study was conducted on 286 bone marrow samples from patients at our hospital. The clinical sensitivity, specificity, and diagnostic coincidence rate of ETV6/RUNX1 FISH detection were analyzed, and the diagnostic consistency of the two methods was analyzed by the kappa test. Results: The probe sensitivity and specificity of the ETV6/RUNX1 probe were 98.47% and 100%, respectively. When 50, 100, and 200 cells were counted, the typical positive signal pattern cutoffs were 5.81%, 2.95%, and 1.49%, respectively, and the atypical positive signal pattern cutoffs were 13.98%, 9.75%, and 6.26%, respectively. The clinical sensitivity of FISH was 96.1%, clinical specificity was 99.6%, diagnostic coincidence rate was 99.00%, diagnostic consistency test kappa value was 0.964, and P value was <0.001. Conclusion: For FISH probes without a national medical device registration certificate, standardized performance verification and methodology performance verification can be performed using laboratory developed test verification standards to ensure a reliable and accurate reference basis for clinical diagnosis and treatment.

[Current status and prospects of neoadjuvant therapy for resectable pancreatic cancer].

Pancreatic cancer is a highly malignant tumor. About 75% of patients with pancreatic cancer who underwent radical surgical resection will still experience postoperative recurrence. Neoadjuvant therapy could improve outcomes in patients with borderline resectable pancreatic cancer,has become a consensus;however it is still controversial in resectable pancreatic cancer. Limited high-quality randomized controlled trial studies support the routine initiation of neoadjuvant therapy in resectable pancreatic cancer. With the development of new technologies, such as next-generation sequencing, liquid biopsy, imaging omics, and organoids, patients are expected to benefit from the precision screening of potential candidates for neoadjuvant therapy and individualized treatment strategy.

Shear stress-induced MMP1 and PDE2A expressions in coronary atherosclerosis.

AIM: Biomechanical stress plays an essential role in coronary atherosclerosis (CAS), however, inter-relations between mechanical conditions and gene expressions remain unclear. METHODS: We constructed finite element model of CAS to map human wall shear stress (WSS). Biopsy aortic tissue samples were obtained from 3 CAS patients. Gene expression pattern in CAS was analyzed by GEO datasets. Immunofluorescence staining and western blot confirmed protein expression and localization. RESULTS: Peak WSS was significantly increased in the vessel stenosis of CAS at 0.25 s (mean 55.1 Pa). Analyses results of GSE76275 showed matrix metalloproteinases1 (MMP1) and phosphodiesterase-2A (PDE2A) up-regulation in endothelial shear responsiveness, which was further validated and localized in vascular endothelial cells, smooth muscle cells and other cells by double immunofluorescence staining. Western blotting assay demonstrated up-regulation of MMP1 and PDE2A expression dependent on the WSS. CONCLUSIONS: MMP1 and PDE2A up-regulations rely on increased WSS in development and risk of CAS, suggesting that their elevation may be potential target for diagnosis and treatment (Fig. 3, Ref. 28).

Predictors of catheter-related bladder discomfort after gynaecological surgery.

BACKGROUND: Urinary catheterization is universally used during surgery, and the incidence of postoperative catheter-related bladder discomfort (CRBD) is very high during recovery. We conducted this study to identify the incidence and predictors of postoperative CRBD after gynaecological surgery in the post-anesthesia care unit (PACU). METHODS: This was a prospective observational study. Patients undergoing gynaecological surgery under general anesthesia with intra-operative urinary catheterization were enrolled. We collected the clinical data, incidence and severity of CRBD, and postoperative pain for the patients. Predictive factors of CRBD were analysed by univariate and multivariate analysis. RESULTS: A total of 407 patients were included in this study. The incidence of CRBD after gynaecological surgery was 64.6% (mild CRBD: 22.8%; moderate CRBD: 34.2%; and severe CRBD: 7.6%). Univariate analysis showed that age, type of surgery, type of laparoscopic surgery, additional analgesics, and postoperative pain were influencing factors for CRBD. Based on multivariate logistic regression analysis, age ≥ 50 years, uterus-related laparoscopic surgery, and lack of additional analgesics were independent predictors of moderate or severe CRBD. CONCLUSIONS: This observational study revealed that the incidence of CRBD after gynaecological surgery in PACU was very high. Age ≥ 50 years, uterus-related laparoscopic surgery, and lack of additional analgesics were independent predictors of CRBD. TRIAL REGISTRATION: ChiCTR1800016390. Registered on 30 May 2018.

[Circulating tumor cells in pancreatic cancer patients: progression in the detection methods and clinical application].

Circulating tumor cells (CTC) disseminate from primary tumors by undergoing epithelial mesenchymal transition that allow their entry into the circulation to drive metastatic formation in pancreatic cancer patients.Technological advances in detection and characterization of CTC are conducive to the early diagnosis, differential diagnosis, monitoring disease progression and predicating the probability of canceration or the chemotherapeutic efficacy. Nowadays, detection methods of CTC can be based on immunomagnetic beads technique, cell filtration or microfluidic chips technology, but there are great differences in the sample throughput, CTC recovery rate, purity, and CTC viability among them.Owing to the dilemma in detection methods, the intrinsic relevance between the biological characteristics of CTC and clinical manifestations is still not exactly elucidated. By the improved methodology, next generation sequencing technology and exploring the technique for culturing CTC in vitro and establishing xenotransplanted tumor model in nude mice, more and more biological information will be revealed, and finally, individualized treatment is achieved.

HCCS1 inhibits the stemness of human pancreatic cancer stem-like cells.

OBJECTIVE: SW1990-spheroid enrichment (SW1990-SE) cells were isolated using a new type of consecutive spheroid enrichment in this study. Cell surface markers were determined by flow cytometry for identification. In vivo tumorigenicity was applied by subcutaneous transplantation in nude mice for verifying the stemness characteristics of SW1990-SE cells. MATERIALS AND METHODS: SW1990-SE cells were subjected to lentivirus infection for establishing the SW1990-SE cell line stably low-expressing HCCS1 (SW1990-SE-shHCCS1) and negative control cell line (SW1990-SE-LV3NC). The stemness regulatory effects of HCCS1 on SW1990-SE cells were evaluated by cell counting kit-8 (CCK-8) assay and 96-wells plate single cell cloning assay in vitro. Subcutaneous transplantation in nude mice was conducted for evaluating the in vivo stemness regulation of HCCS1 on SW1990-SE cells.. RESULTS: HCCS1 knockdown in SW1990-SE cells did not markedly change the cell proliferation and doubling time, whereas the in vitro spheroid diameter and single cell cloning efficacy remarkably increased. In vivo experiments showed that HCCS1 knockdown greatly enhanced the tumorigenicity of SW1990-SE cells in nude mice. CONCLUSIONS: This study first obtains the human pancreatic cancer stem-like cells SW1990-SE through consecutive spheroid enrichment. Both in vivo and in vitro experiments verified that HCCS1 knockdown largely enhanced the stemness of SW1990-SE cells. Our study provides an important reference for the research of tumor stem cells.

Comparison of intravenous lidocaine and dexmedetomidine infusion for prevention of postoperative catheter-related bladder discomfort: a randomized controlled trial.

BACKGROUND: Catheter-related bladder discomfort (CRBD) frequently occurs during recovery in patients who undergo intra-operative urinary catheterization. We conducted this study to compare the effect of intravenous lidocaine and dexmedetomidine infusion for preventing CRBD. METHODS: 120 patients undergoing elective open abdominal hysterectomy or hysteromyomectomy requiring urinary bladder catheterization were randomly allocated into three groups of 40 each. Group L received a 2 mg/kg lidocaine bolus followed by infusion of 1.5 mg/kg/h; Group D received a 0.5 µg/kg dexmedetomidine bolus followed by infusion of 0.4 µg/kg/h; Group C received a bolus and infusion of normal saline of equivalent volume. The incidence and different severity (mild, moderate, and severe) of CRBD were assessed on arrival in the postanaesthesia care unit at 0, 1, 2, and 6 h postoperatively. RESULTS: The incidence of CRBD was significantly lower in Group L and Group D compared with Group C at 0, 1, and 2 h. However, there was no significant difference among the three groups regarding the different severity of CRBD at all time points. The requirement of rescue tramadol for CRBD was lower in group L and group D than in group C. The incidence of sedation was significantly higher in Group D compared to Group L and Group C, though no difference in other adverse effects was observed. CONCLUSIONS: Intravenous lidocaine and dexmedetomidine infusion reduced the incidence of CRBD as well as the additional tramadol requirement for CRBD, but had no effect on the different severity of CRBD. TRIAL REGISTRATION: ChiCTR-INR-16009162 . Registered on 5 September 2016.

PKC δ gene can induce macrophages to release inflammatory factors against Mycobacterium tuberculosis infection.

OBJECTIVE: To investigate the effect of PKC δ gene on the anti-tuberculosis activity of macrophages and the mechanism. MATERIALS AND METHODS: Bone marrow cells of PKC δ knockout mice and wild-type mice were cultured and L929 cells were induced to differentiate into macrophages. Lipopolysaccharide (LPS) and trehalose 6,6'-dimycolate (TDM) were used to stimulate macrophages respectively. After 24 and 96 hours, cells and the supernatant were collected to evaluate the inflammatory cytokines produced by macrophages using ELISA method. Real-time PCR was performed to detect the expression of macrophage mRNA level and nitric oxide (NO) production of macrophages was measured by NO assay. RESULTS: The results showed that, after TDB stimulation, IL-1ß, IL-6, and other cytokines, as well as NO produced by macrophages of PKC δ knockout mice, were significantly decreased (p < 0.01) compared with the wild-type mice. In PKC δ knockout macrophages, the above protein-coding genes were also decreased significantly at the transcriptional level (p < 0.01). CONCLUSIONS: PKC δ can enhance the anti-tuberculosis capacity of macrophages by inducing to the release of inflammatory factors by macrophages.

Measurement of the absolute velocity of blood flow in early-stage chick embryos using spectral domain optical coherence tomography.

Doppler optical coherence tomography (OCT) is a noninvasive imaging modality that provides quantitative flow information with high spatial and temporal resolution. However, it is only sensitive to the flow velocity vector parallel to the incident beam. To calculate the absolute velocity, it is necessary to obtain the angle between the incident beam and flow field. In this paper, we describe a practical method to measure the Doppler angle based on the structural information of blood vessels extracted from spectral domain OCT images. In this method, a normal sectional scan of the vessel is performed where the probe beam is perpendicular to the vessel. Next, the axial diameter (Z direction) of the vessel (DA) was measured in the acquired image. For a certain scan in which the probe beam is oblique to the blood vessel, the axial diameter of the vessel (DA') can be measured. Thus, the Doppler angle can be calculated depending on the ratio of DA and DA', and absolute blood flow was determined. We validate this method in a capillary tube as well as in large blood vessels of early-stage chick embryos. This technique is suitable for early-stage embryo blood-flow measurement because most of the blood vessels are easily differentiated from the transparent surrounding structures during that time.

Stimulation of contractions in pregnant human myometrium is associated with 5-HT3 receptors.

BACKGROUND: Serotonin (5-HT) is known to play an important role in regulating uterine contractions. However, the specific receptors involved have not been well characterized. We evaluated whether 5-HT3 receptors exist in human myometrium, and their effects on myometrial contractility when stimulated by a 5-HT3 agonist. METHODS: Four tissue samples taken from patients undergoing hysterectomy (n=2) and elective cesarean delivery (n=2) were used to detect expression of 5-HT3 receptors on the myometrium using western blotting. For isometric tension measurement, another 12 myometrial strips obtained from patients undergoing elective cesarean delivery were randomly divided into a control group (Group CON) and an RS56812 group (Group RG). In increasing doses from 10-7M to 10-4M, RS56812, a 5-HT3 receptor agonist, was used to investigate the contractile effects after bonding to the 5-HT3 receptor, following which the effects of granisetron were assessed. Amplitude, interval and duration of myometrial contractions were recorded. RESULTS: Proteins with a molecular mass of 55kDa, consistent with 5-HT3 receptors, were detected both on non-pregnant and late-pregnant human uteri. RS56812 increased the contractile amplitude at concentrations of 10-6M, 10-5M and 10-4M, achieving maximum effect at 10-5M. A prolonged contractile interval was detected at the concentration of 10-4M. However, RS56812 showed no significant effect on contraction duration. Granisetron did not reverse the contractile effects induced by RS56812. CONCLUSION: 5-HT3 receptors are expressed on non-pregnant and pregnant uteri. RS56812 enhanced myometrial contractions, but this was not affected by granisetron, the mechanism of which requires further investigation.

[Protein-repellent and antibacterial properties of modified orthodontic adhesive].

OBJECTIVE: To develop a novel protein-repellent orthodontic adhesive by incorporating 2-methacryloyloxyethyl phosphorylcholine(MPC). METHODS: MPC was incorporated into a commercially available orthodontic adhesive(Fuji ORTHO) at 0% (control), 1.5%, 3.0%, and 5.0% by mass. Enamel shear bond strength(SBS) was determined. Protein adsorption onto specimens was determined by a micro bicinchoninic acid method. A dental plaque microcosm biofilm model with human saliva as inoculum was used to investigate biofilm viability. RESULTS: The SBS was not reduced in the group(3.0% MPC), compared to the control group. The amount of protein adsorption in the group(3.0% MPC) was (0.46±0.06) µg/cm(2) and (4.57 ± 0.42) µg/cm(2) in the control group. Lactic acid production of biofilms in the group(3.0% MPC) was (7.12±1.03) mmol/L and (12.16±1.24) mmol/L in the control group. CONCLUSIONS: MPC based orthodontic adhesive greatly reduced the protein adsorption and bacterial adhesion, without compromising enamel shear bond strength.

Differential expression profiling of microRNAs in para-carcinoma, carcinoma and relapse human pancreatic cancer.

PURPOSE: To explore the altered different expression of miRNAs and the mechanisms underlying the relapse and metastasis of pancreatic cancer. MATERIALS AND METHODS: The most differentially expressed miRNAs were analyzed by gene ontology (GO) term analysis, Kyoto encyclopedia of genes and genomes (KEGG) pathway analysis and protein interaction analysis. The potentially regulated target genes of the most differentially expressed miRNAs were also analyzed further by GO term analysis and KEGG pathway analysis, and quantitated by qRT-PCR. RESULTS: In total, we found 12 miRNAs displayed at least a 30-fold increase or decrease in expression of carcinoma and relapse vs. para-carcinoma human pancreatic cancer (C/R vs. P). In addition, our study found that pancreatic cancer was related to pathways in cancer, including Jak-STAT signaling pathway, MAPK signaling pathway and PPAR signaling pathway. CONCLUSIONS: The differential expressed miRNAs and their predicted target genes that involved in Jak-STAT signaling pathway, MAPK signaling pathway and PPAR signaling pathway indicating their potential roles in pancreatic carcinogenesis and progress.

Relaxant effects of metoclopramide and magnesium sulfate on isolated pregnant myometrium: an in vitro study.

BACKGROUND: Metoclopramide and magnesium sulfate are extensively used agents in obstetrics. In this study, the relaxant properties of metoclopramide and magnesium sulfate on pregnant myometrium, together with the possible reversing influences of oxytocin and cabergoline (a dopamine D2 receptor agonist), were investigated. METHODS: Myometrial strips from 24 parturients were randomly allocated to four groups: control (Group CON), magnesium sulfate and oxytocin (Group MSO), metoclopramide and oxytocin (Group MEO), and metoclopramide and cabergoline (Group MEC). Myometrial strips were mounted on a myograph bathed in Krebs buffer. Saline (Group CON) and five incremental doses of magnesium sulfate (Group MSO) or metoclopramide (Groups MEO and MEC) were sequentially microinjected into the bath. Subsequently, oxytocin (Groups CON, MSO and MEO) or cabergoline (Group MEC) was microinjected into the bath. The myometrial contractile characteristics after each drug injection, including contractile force, interval and duration, were analyzed. RESULTS: Magnesium sulfate was more potent for prolonging myometrial contractile interval than reducing contractile force. Metoclopramide relaxed myometrial contractions by inhibiting contractile force and prolonging contractile interval in a concentration-dependent manner. Oxytocin reversed both the inhibited contractile force and the prolonged contractile interval caused by a high concentration of magnesium sulfate but accelerated the contractile interval and had no significant effect on the contractile force suppressed by metoclopramide. The relaxant effects of metoclopramide were completely reversed by cabergoline. CONCLUSIONS: Both magnesium sulfate and metoclopramide relaxed myometrial contractions, and exhibited different responses to subsequent oxytocin treatment. The relaxant mechanism of metoclopramide may be via blockade of dopamine D2 receptor, which requires further investigation.

First Report of Powdery Mildew Caused by Oidium neolycopersici on Tomato in China.

Tomato powdery mildew can cause remarkable reduction in fruit size and quality (4). In March of 2008, powdery mildew appeared as circular, white colonies on leaves, petioles, and stems of tomato plants grown in greenhouses in Shangqiu, Henan Province, China. The pathogenic fungus had unbranched conidiophores with an average length of 58.4 µm and width of 5.1 µm. Conidia were hyaline, elliptical, and were borne singly. Average length and width of conidia were 30.6 and 15.1 µm, respectively. Germ tubes were straight and formed at the ends or very close to the ends of conidia. Chasmothecium was not found in the collected samples. Different tomato cultivars and species, including Lycopersicon esculentum Mill (cvs. Moneymaker, Micro-Tom, Zaofen, Fenguo, and Zhongza series), L. peruvianum cv. LA2172, and L. hirsutum cv. G1.1560, were inoculated with a conidial suspension with a concentration of 5 × 104 conidia/ml. Plants developed powdery mildew symptoms as early as 4 days after inoculation. Susceptible symptoms developed on all L. esculentum cultivars, while L. peruvianum LA2172 and L. hirsutum G1.1560 displayed complete resistance, which is similar to the results of Bai et al 2004 (1) and Lindhout and Pet 1990 (3). Morphological characteristics of the pathogen on susceptible genotypes were similar to those from naturally infected plants. On the basis of the characteristics of the asexual stage, the pathogen was identified as an isolate of Oidium neolycopersici L. Kiss, which was confirmed by internal transcribed spacer (ITS) sequence analysis. PCR amplification and sequencing of the ITS region were performed with primers ITS1 and ITS4. The nucleotide sequence was assigned GenBank Accession No. EU486992, which had a 100% homology with 10 ITS sequences of O. neolycopersici in GenBank (Accession Nos. EU047559 to 047568) (2). In Asia, the spread of this pathogen has been recently reported in Japan (2). To our knowledge, this is the first report of tomato powdery mildew in China. Voucher specimens are available at the Specimen Center in the Department of Life Science, Shangqiu Normal University. References: (1) Y. Bai et al. Mol. Plant-Microbe. Interact. 18:354, 2005. (2) T. Jankovics et al. Phytopathology 98:529, 2008. (3) P. Lindhout and G. Pet. Tomato Gen. Coop. Rep. 40:19, 1990. (4) J. M. Whipps et al. Plant Pathol. 47:36, 1998.

Alteration in the copy number of mitochondrial DNA in leukocytes of patients with mitochondrial encephalomyopathies.

OBJECTIVES: We investigated whether mutation of mitochondrial DNA (mtDNA) affects the copy number of the mitochondrial genome in patients with mitochondrial myopathy encephalopathy with lactic acidosis and stroke-like episodes (MELAS) and those with myoclonic epilepsy with ragged-red fiber (MERRF) syndromes. MATERIALS AND METHODS: Forty-eight Taiwanese patients with MELAS syndrome and 20 patients with MERRF syndrome were recruited in this study. RESULTS: In relation to controls, the copy numbers of mtDNA in leukocytes of patients with MELAS or MERRF syndrome were significantly higher at a young age but lower at an advanced age. In addition, MELAS patients harboring higher proportions of mtDNA with A3243G transition had lower mtDNA copy numbers. The MELAS or MERRF patients with multi-system disorders had lower mtDNA copy numbers in leukocytes. Furthermore, higher proportions of mtDNA with 4977 bp deletion were found in leukocytes of MERRF patients with multi-system involvement. CONCLUSION: In leukocytes, alteration in the copy number of mtDNA is related to the proportion of mtDNA with a point mutation or large-scale deletion, which may serve as a biomarker in the pathogenesis and disease progression of MELAS and MERRF syndromes.

Mitochondrial theory of aging matures--roles of mtDNA mutation and oxidative stress in human aging.

Mitochondrial theory of aging, a variant of free radical theory of aging, proposes that accumulation of damage to mitochondria and mitochondrial DNA (mtDNA) leads to aging of humans and animals. It has been supported by the observation that mitochondrial function declines and mtDNA mutation increases in tissue cells in an age-dependent manner. Age-related impairment in the respiratory enzymes not only decreases ATP synthesis but also enhances production of reactive oxygen species (ROS) through increased electron leakage in the respiratory chain. Human mtDNA, which is not protected by histones and yet is exposed to high levels of ROS and free radicals in the matrix of mitochondria, is susceptible to oxidative damage and mutation in tissue cells. In the past decade, more than one hundred mtDNA mutations have been found in patients with mitochondrial disease, and some of them also occur in aging human tissues. The incidence and abundance of these mutant mtDNAs are increased with age, particularly in tissues with great demand for energy. On the other hand, recent studies have revealed that the ability of the human cell to cope with oxidative stress is compromised in aging. Comparative analysis of gene expression by microarray technology has shown that a number of genes related to oxidative stress response are altered in aging animals. We discovered that the transcripts of early growth response protein-1, growth arrest and DNA damage-inducible proteins and glutathione S-transferase genes are increased in response to oxidative stress in human skin fibroblasts. Moreover, the activities of Cu,Zn-SOD, catalase and glutathione peroxidase decrease with age, whereas Mn-SOD activity increases with age up to 65 years and slightly declines thereafter in skin fibroblasts. Such an imbalance in the function of antioxidant enzymes may result in excess production of damaging ROS in the cell. This notion is supported by the observation that intracellular levels of H2O2 and oxidative damage to DNA and lipids are significantly increased with age of the fibroblast donor. Furthermore, the mitochondrial pool of reduced glutathione declines and DNA damage is enhanced in aging tissues. Taken together, these observations and our previous findings that mtDNA mutations and oxidative damage are increased in aging human tissues suggest that mitochondrial theory of aging is mature.

Oxidative stress in human aging and mitochondrial disease-consequences of defective mitochondrial respiration and impaired antioxidant enzyme system.

Respiratory function of mitochondria is compromised in aging human tissues and severely impaired in the patients with mitochondrial disease. A wide spectrum of mitochondrial DNA (mtDNA) mutations has been established to associate with mitochondrial diseases. Some of these mtDNA mutations also occur in various human tissues in an age-dependent manner. These mtDNA mutations cause defects in the respiratory chain due to impairment of the gene expression and structure of respiratory chain polypeptides that are encoded by the mitochondrial genome. Since defective mitochondria generate more reactive oxygen species (ROS) such as O2- and H2O2 via electron leak, we hypothesized that oxidative stress is a contributory factor for aging and mitochondrial disease. This hypothesis has been supported by the findings that oxidative stress and oxidative damage in tissues and culture cells are increased in elderly subjects and patients with mitochondrial diseases. Another line of supporting evidence is our recent finding that the enzyme activities of Cu,Zn-SOD, catalase and glutathione peroxidase (GPx) decrease with age in skin fibroblasts. By contrast, Mn-SOD activity increases up to 65 years of age and then slightly declines thereafter. On the other hand, we observed that the RNA, protein and activity levels of Mn-SOD are increased two- to three-fold in skin fibroblasts of the patients with CPEO syndrome but are dramatically decreased in patients with MELAS or MERRF syndrome. However, the other antioxidant enzymes did not change in the same manner. The imbalance in the expression of these antioxidant enzymes indicates that the production of ROS is in excess of their removal, which in turn may elicit an elevation of oxidative stress in the fibroblasts. Indeed, it was found that intracellular levels of H2O2 and oxidative damage to DNA and lipids in skin fibroblasts from elderly subjects or patients with mitochondrial diseases are significantly increased as compared to those of age-matched controls. Furthermore, Mn-SOD or GPx-1 gene knockout mice were found to display neurological disorders and enhanced oxidative damage similar to those observed in the patients with mitochondrial disease. These observations are reviewed in this article to support that oxidative stress elicited by defective respiratory function and impaired antioxidant enzyme system plays a key role in the pathophysiology of mitochondrial disease and human aging.

Increases of mitochondrial mass and mitochondrial genome in association with enhanced oxidative stress in human cells harboring 4,977 BP-deleted mitochondrial DNA.

In order to investigate the effect of aging- and disease-associated deletion of mtDNA on cellular functions, we used cytoplasm fusion to construct a series of the cybrids harboring varying proportions of mtDNA with 4,977 bp deletion from skin fibroblasts of a patient with chronic progressive external ophthalmoplegia. The cybrids were grown in the Dulbecco's modified Eagle medium supplemented with 5% fetal bovine serum, 100 microg/ml pyruvate and 50 microg/ml uridine. The population doubling time was longer for the cybrids containing higher proportions of 4,977 bp-deleted mtDNA. In addition, we found that the respiratory function was decreased with the increase of mtDNA with 4,977 bp deletion in the cybrids. Since impairment of the respiratory system of mitochondria increases the electron leak of the respiratory chain, we further determined the oxidative stress in these cybrids. The results showed that the specific contents of 8-hydroxy 2'-deoxyguanosine and lipid peroxides of the cybrids harboring > 65% of the 4,977 bp-deleted mtDNA were significantly increased as compared with those of the cybrids containing undetectable mutant mtDNA. On the other hand, we found that the mitochondrial mass and the relative content of the mitochondrial genome in the cybrids harboring 4,977 bp-deleted mtDNA were higher than those of the cybrids containing only wild type mtDNA. The relative content of mtDNA was increased 17% and 30%, respectively, in the cybrids harboring 17% and 56% of mtDNA with 4,977 bp deletion. Moreover, both mitochondrial mass and mtDNA content were concurrently increased by treatment of the cybrids with 180 microM of hydrogen peroxide. Taken these findings together, we conclude that increase of mitochondrial mass and mtDNA are the molecular events associated with enhanced oxidative stress in human cells with impaired respiratory function caused by mtDNA deletion.

[A study on cultivation techniques for Glycyrrhiza uralensis fisch. in Daxing County area, Beijing].

OBJECTIVE: To study the suitable cultivation techniques for Glycyrrhiza uralensis in the sandy soil in Daxing county area, the Beijing. METHOD: Small plot trial and Large-scale cultivation in the fields. RESULT AND CONCLUSION: Uralensis can be cultivated in the sandy soil in Daxing County area, Beijing.

[Studies permeation of puerarin and its phospholipid complex through mouse skin in vitro].

OBJECTIVE: To investigate the skin permeation of puerarin and its phospholipid complex and compare the differences between their permeation rates and cumulative permeation amounts. METHOD: Performing a test of permeation through the mouse skin in vitro in an improved Franz diffusion cell. RESULT: The cumulative permeation amount of phospholipid complex was higher than that of puerarin in the first hour and then increased slowly, meanwhile the permeation rate of puerarin rose higher than that of the complex. CONCLUSION: Phospholipid complex of puerarin can permeate through the mouse skin rapidly up to a certain amount in a short time, then begins to release drug slowly and lastingly.