This study examined the impact of luteinized unruptured follicle (LUF) on endometrial receptivity. The menstrual cycle of 17 LUF patients (LUF group) and 13 ovulatory women (control group) was monitored by measuring LH level in urine and by ultrasonic examination. An endometrial biopsy at the sixth to tenth day after LH surge was taken in all the patients. The expressions of endometrial ER, PR and integrin ανβ3 were immunohistochemically determined. At the same time, the serum levels of E2 and P were detected by chemiluminescence. The results exhibited that (1) The mean serum P level in LUF group (7.32±2.56 ng/mL) was significantly lower than that in control group (11.17±3.17 ng/mL) (P<0.01). But there was no significant difference in the mean serum E2 levels between LUF group (179.35±81.60 pg/mL) and the control group (198.58±75.23 pg/mL) (P>0.05); (2) The mean expression intensities of ER, PR in endometrium of LUF group (183.86±2.43, 167.94±3.04) were significantly higher than those in control group (109.35±6.31, 105.98±4.07) (P<0.01); (3) The mean expression intensities of integrin ανβ3 in endomtrium of LUF patients (114.90±11.38) were significantly lower than those in control group (191.34±1.82) (P<0.01); (4) The change profile of integrin ανβ3 expression in the endometrium of LUF patients was in positive relation with serum P level (r=0.77, P<0.01), but bore no significant relationship with serum E2 level (r=0.01, P>0.05). It was concluded that the depression of serum P levels in LUF patients was closely related to the failure of the down-regulation of ER and PR, and the low expressions of integrin ανβ3 also suggested that the delayed implantation and the impaired endometrial receptivity had impact on embryonic implantation.
In order to explore the effects of Panax notoginoside (PNS) on the expression of transforming growth factor β1 (TGF-β1) and Smad-7 in renal tissues of diabetes, a rat model of diabetic nephropathy was set up by intravenous injection of streptozotocin (STZ). Wistar rats were randomly divided into normal group, diabetic control group, group treated by PNS at low-dosage (PL), group treated by PNS at high-dosage (PH) and group treated by catopril (C), respectively. Fasting blood glucose (FBG), renal index, endogenous creatinine clearance rate (CCr) and urinary albumin (UAlb) in 24 h were examined after 6 weeks. Meanwhile, the expressions of TGF-β1 and Smad7 in renal tissues were immunohistochemically dectected. At the end of the sixth week, FBG, renal index, Ccr, UAlb were all elevated significantly in control group (P<0.01). The expression of TGF-β1 protein was increased while Smad7 protein decreased in renal tissue (P<0.01). However, the treatment with PNS reversed the aforementioned changes in renal tissues of diabetic rats. These results indicate that PNS possess a protective effect on the kidney of diabetic rats and it might protect kidney by inhibiting the expression of TGF-β1 protein and enhancing the expression of Smad7 protein.
Mesenchymal stem cells (MSCs) were induced into a nucleus pulposus-like phenotype utilizing simulated microgravity in vitro in order to establish a new cell-based tissue engineering treatment for intervertebral disc degeneration. For induction of a nucleus pulposus-like phenotype, MSCs were cultured in simulated microgravity in a chemically defined medium supplemented with 0 (experimental group) and 10 ng/mL (positive control group) of transforming growth factor β1 (TGF-β1). MSCs cultured under conventional condition without TGF-β1 served as blank control group. On the day 3 of culture, cellular proliferation was determined by WST-8 assay. Differentiation markers were evaluated by histology and reverse transcriptase-polymerase chain reaction (RT-PCR). TGF-β1 slightly promoted the proliferation of MSCs. The collagen and proteoglycans were detected in both groups after culture for 7 days. The accumulation of proteoglycans was markedly increased. The RT-PCR revealed that the gene expression of Sox-9, aggrecan and type II collagen, which were chondrocyte specific, was increased in MSCs cultured under simulated microgravity for 3 days. The ratio of proteoglycans/collagen in blank control group was 3.4-fold higher than positive control group, which denoted a nucleus pulposus-like phenotype differentiation. Independent, spontaneous differentiation of MSCs towards a nucleus pulposus-like phenotype in simulated microgravity occurred without addition of any external bioactive stimulators, namely factors from TGF-β family, which were previously considered necessary.
The growth inhibition and pro-apoptosis effects of dracorhodin perchlorate on human prostate cancer PC-3 cell line were examined. After administration of 10–80 μmol/L dracorhodin perchlorate for 12–48 h, cell viability of PC-3 cells was measured by MTT colorimetry. Cell proliferation ability was detected by colony formation assay. Cellular apoptosis was inspected by acridine orange-ethidium bromide fluorescent staining, Hoechst 33258 fluorescent staining, and flow cytometry (FCM) with annexin V-FITC/propidium iodide dual staining. The results showed that dracorhodin perchlorate inhibited the growth of PC-3 in a dose- and time-dependent manner. IC50 of dracorhodin perchlorate on PC-3 cells at 24 h was 40.18 μmol/L. Cell clone formation rate was decreased by 86% after treatment with 20 μmol/L of dracorhodin perchlorate. Some cells presented the characteristic apoptotic changes. The cellular apoptotic rates induced by 10–40 μmol/L dracorhodin perchlorate for 24 h were 8.43% to 47.71% respectively. It was concluded that dracorhodin perchlorate significantly inhibited the growth of PC-3 cells by suppressing proliferation and inducing apoptosis of the cells.
This study examined the effect of silencing LRIG3 expression on the proliferation and apoptosis of bladder cancer T24 cells and explored the role of LRIG3 in the tumorigenesis of bladder cancer. Bladder cancer T24 cells were routinely cultured and pSilencer plasmids were employed to construct LRIG3 eukaryotic expression vector of LRIG3-siRNA, i.e., pSilencer-LRIG3-siRNA. After confirmation, the vector was transfected into HEK293 cells to make a replication-deficient adenovirus, pAd-LRIG3-siRNA, which was then introduced into bladder cancer T24 cells. RT-PCR, Western-blotting were performed to detect the levels of LRIG3 mRNA and proteins. Cells number was determined by using MTT test. Hoechst33258 staining, transmission microscopy, flow cytometery were conducted to examine the cell apoptosis. Three groups included a blank control group, a negative control group (containing non-interfering plasmids) and a pAd-LRIG3-siRNA group. Our results showed that the recombinant pAd-LRIG3-siRNA was successfully transfected into the bladder cancer T24 cells. The siRNA formed by the transcription of the recombinant plasmids resulted in significantly reduced expressions of LRIG3 gene and protein and significantly decreased cell proliferation and growth in the pAd-LRIG3-siRNA group as compared with the control group (P<0.01). The siRNA also caused apoptotic changes of some cells, with the apoptosis rate being (17.69±0.75)%, which was significantly different from that of the control group (P<0.01). It was concluded that recombinant pAd-LRIG3-siRNA plasmids could effectively decrease the expression of LRIG3 mRNA and proteins and, to some extent, inhibit the proliferation and promote the apoptosis of bladder cancer T24 cells. Silencing LRIG3 gene might be a novel alternative for the treatment of bladder cancer.
This study examined the effect of MMP9 gene on the biological behaviors of trophoblasts and explore the relation between MMP9 gene and the “superficial implantation of placenta”. In vitro cultured trophoblasts (TEV-1 cells) were transfected with synthesized double-stranded MMP9 RNA (siRNA) by using lipofectamine2000™ technique and the expressions of MMP9 mRNA and protein and the growth and invasiveness of the TEV-1 cells were determined. Our results showed that siRNA transfection could significantly inhibit the expression of MMP9 gene in the TEV-1 cells and the growth and invasiveness of the TEV-1 cells transfected RNA was significantly reduced (P<0.01). We are led to conclude that silencing of MMP9 gene with siRNA can inhibit the growth and invasiveness of trophoblasts and increasing the expression of MMP9 might help prevent and treat preeclampsia.
The aim of this study is to investigate the influence of different posts on the fracture mechanics of endodontically-treated teeth with open apex. Forty-eight human maxillary anterior teeth were collected, and the root was transversely sectioned 12 mm under the cementoenamal junction (CEJ). These samples were then randomly divided into two groups, i.e., minor diameter open apex root (group A) and major diameter open apex root (group B), with mineral trioxide aggregate (MTA) placed into the apical 4 mm in the root canals. Subsequently, both groups were respectively further divided into three subgroups as follows: fiber-post (subgroup 1), metal post (subgroup 2) and non-post (subgroup 3) group. Teeth were restored with a composite resin crown and tested by using a universal testing machine at the rate of 1 mm/min cross-head. Values of the maximum fracture resistance and failure patterns were recorded and compared among all subgroups. In addition, the changes of MTA properties were carefully examined via X-ray photography. Our results indicate that (1) In group A, the mean value of fracture resistance for teeth restored with fiber posts were statistically higher than that with either metal post or non-post; (2) In group B, there was no statistically significant difference in the mean value of fracture resistance among three subgroups; (3) No statistical significance in the mean value of fracture resistance was found between group A and group B; (4) The failure modes of most samples (58%) were irreparable; (5) MTA in two teeth developed cracks after loading tests. In conclusion, endodontically-treated teeth restored with fiber posts are more resistant to fracture than those restored with either metal posts or non-post, and most of the fracture modes are catastrophic in nature.
DNA damage response (DDR) in different cell cycle status of human peripheral blood lymphocytes (PBLs) and the role of H2AX in DDR were investigated. The PBLs were stimulated into cell cycle with phytohemagglutinin (PHA). The apoptotic ratio and the phosphorylation H2AX (S139) were flow cytometrically measured in resting and proliferating PBLs after treatment with camptothecin (CPT) or X-ray. The expressions of γH2AX, Bcl-2, caspase-3 and caspase-9 were detected by Western blotting. DDR in 293T cells was detected after H2AX was silenced by RNAi method. Our results showed that DNA double strand breaks (DSBs) were both induced in quiescent and proliferating PBLs after CPT or X-ray treatment. The phosphorylation of H2AX and apoptosis were more sensitive in proliferating PBLs compared with quiescent lymphocytes (P<0.05). The expression levels of anti-apoptotic proteins Bcl-2 were reduced and cleaved caspase-3 and caspase-9 were increased. No significant changes were observed in CPT-induced apoptosis in 293T cells between H2AX knocking down group and controls. It was concluded that proliferating PBLs were more vulnerable to DNA damage compared to non-stimulated lymphocytes and had higher apoptosis rates. γH2AX may only serve as a marker of DNA damage but exert no effect on apoptosis regulation.
DNA repair processes play a role in the development of drug resistance which represents a huge obstacle to leukemia chemotherapy. Histone H2AX phosphorylation (ser139) (γH2AX) occurs rapidly at the onset of DNA double strand break (DSB) and is critical to the regulation of DSB repair. If DNA repair is successful, cells exposed to anti-neoplastic drugs will keep entering the cycle and develop resistance to the drugs. In this study, we investigated whether γH2AX can be used as an indicator of tumor chemosensitivity and a potential target for enhancing chemotherapy. K562 and multi-drug resistant cell line K562/A02 were exposed to adriamycin (ADR) and γH2AX formed. Flow cytometry revealed that percentage of cells expressing γH2AX was increased in a dose-dependent manner and the percentage of K562/A02 cells was lower than that of K562 cells when treated with the same concentration of ADR. In order to test the potential of γH2AX to reverse drug resistance, K562/A02 cells were treated with PI3K inhibitor LY294002. It was found that LY249002 decreased ADR-induced γH2AX expression and increased the sensitivity of K562/A02 cells to ADR. Additionally, the single-cell gel electrophoresis assay and the Western blotting showed that LY249002 enhanced DSBs and decreased the expression of repair factor BRCA1. These results illustrate chemosensitivity can partly be measured by detecting γH2AX and drug resistance can be reversed by inhibiting γH2AX.
This study investigated the intracellular localization of asparagine synthetase (ASNS) in the relation with chemoresistance in leukemia. pIRES-GFP-ASNS-Flag/Neo expression vector was transiently tansfected into SK-N-MC cells and 297T cells respectively. Immunofluorescence and Western blot analysis were performed for cellular localization of ASNS respectively. U937 cells were treated with L-asparaginase for 48 h and examined for endogenous ASNS expression on plasma membrane by immunofluorescence staining. Immunofluorescence staining showed that the transiently expressed ASNS was partly localized on transfected-SK-N-MC cell surface. Moreover, Western blotting exhibited that ASNS expressed both in cytosol and on plasma membrane of transfected-293T cells. Immunofluorescence staining with anti-ASNS-specific monoclonal antibody revealed that endogenous ASNS was localized on the plasma membrane of U937 cells, except for its distribution in the cytosol. In addition, ASNS exhibited a higher expression on plasma membrane after treatment with L-asparaginase as compared with the untreated cells. It was concluded that the subcellular translocation of ASNS may play an important role in L-asparaginase resistance in leukemia cells.
Autonomic nervous system activation can result in significant changes of atrial electrophysiology and facilitate induction of atrial fibrillation. By recording influence of different concentrations of acetylcholine (ACh) on atrial fibers (AF), we investigated the role of the increased vagal tone in electrical remodeling in atrial fibrillation. Parameters of action potentials and force contraction (Fc) in atrial fibers were recorded by using standard intracellular microelectrode technique and force transducer. It was found that: (1) ACh at 0.1 μmol/L had no significant influence on spontaneous action potentials (SAPs) and Fc (n=6, P>0.05); ACh at both 1.0 and 10.0 μmol/L shortened action potential duration (APD) and Fc of human AF from right atrium (n=6, P<0.05); there was no significant difference in shortening APD between 10.0 and 1.0 μmol/L of ACh; (2) ACh at 0.1 μmol/L had no significant desensitization (n=6, P>0.05), but ACh at 1.0 and 10.0 μmol/L had desensitization (n=6, P<0.05) to SAPs and Fc. The desensitization of ACh on APD in AF was concentration- and time-dependent. It was shown that APD was longer than the control along with extending time of continuous Tyrode’s solution perfusion after desensitization. It is concluded that ACh changes the electrophysiological characteristics of human AF, indicating that increased vagal tone plays a role in the development of a vulnerable substrate for atrial electrical remodeling in atrial fibrillation.
This study examined the relationship between PDGF-induced proliferation of vascular smooth muscle cells (VSMCs) and Nur77 expression and the effect of atorvastatin on VSMC proliferation and Nur77 in PDGF-treated VSMCs. Rat VSMCs were isolated and cultured. After incubation with atorvastatin or Nur77 siRNA, the cells were stimulated with PDGF and detected for BrdU incorporation to measure the proliferation of the VSMCs. Quantitative PCR and Western blotting were used to determine the Nur77 protein and the CREB phosphorylation level, to observe their relations with PDGF-induced VSMC proliferation. Our results showed that PDGF increased the BrdU incorporation in VSMCs, suggesting that it induced the proliferation of the cells. The VSMC proliferation was associated with increased Nur77 expression and elevated CREB phosphorylation. Atorvastatin inhibited the PDGF-induced VSMC proliferation, suppressed Nur77 expression. After silencing of Nur77 gene, the PDGF-induced VSMC proliferation was decreased. It was concluded that PDGF-induced VSMC proliferation was related to the Nur77 expression and CREB phosphorylation. Atorvastatin reduced the Nur77 expression and, at the same time, inhibited the VSMC proliferation.
Adipose tissue is a readily available source of adult stem cells with multipotent properties suitable for tissue engineering and regenerative medical applications. Peptide hydrogel is a novel biomaterial which provides three-dimensional microenvironments for a variety of cells for tissue grafting. In this study, adipose-derived stem cells (ADSCs) were isolated from rats, seeded into the peptide hydrogel polymer scaffolds and cultured in Neurobasal (NB) media supplemented with B27, basic fibroblast growth factor (bFGF) and epidermal growth factor (EGF). Ten days after the culture, some cells were expanded into clonal populations in which the expression of both Nestin and Brdu was detected but only Brdu expression was detected in the cells that were not expanded into clonal populations. Our results suggested that ADSCs in peptide hydrogel polymer scaffolds can be induced to differentiate into cells capable of expressing the neuron-associated markers, self-renewal and self-propagation.
Histopathological examination of testes is important in assessing spermatogenesis and testicular function. Modified Davidson’s fluid (mDF) has been proposed as a superior substitute for Bouin’s fluid (BF) for fixation of adult animal testes. Besides, 4% paraformaldehyde (PFA) has been commonly used to fix testes with convenience. We compared the morphology of the rat testis fixed in 4% PFA, mDF, or BF using hematoxylin and eosin (HE)-stained sections. Fixation in 4% PFA resulted in obvious tissue shrinkage artifacts, especially between seminiferous epithelium cells. Shrinkage artifacts were also observed in the central area of the testes fixed in BF. Use of mDF did not cause shrinkage artifacts between seminiferous tubules, though a small amount can be observed in seminiferous tubules between germ cells. Clarity of nuclear detail in testes fixed in mDF and BF is better compared to 4% PFA. Our study demonstrated that fixation in mDF provided better morphologic details in the rat testis as compared with 4% PFA and BF.
This study investigated the effects and molecular mechanisms of genistein in improving insulin resistance induced by free fatty acids (FFAs) in HepG2 hepatocytes. A model of insulin resistance in HepG2 cells was established by adding palmitic acid (0.5 mmol/L) to the culture medium and the cells were treated by genistein. Glucose consumption of HepG2 cells was determined by glucose oxidase method. The levels of c-jun N-terminal kinase (JNK) phosphorylation, insulin receptor substrate-1 (IRS-1) Ser307 phosphorylation, JNK, IRS-1, phosphatidylinositol-3-kinase p85 (PI-3K p85) and glucose transporter 1 (GLUT1) proteins were detected by Western blotting. The results showed that after the treatment with palmitic acid for 24 h, the insulin-stimulated glucose transport in HepG2 cells was inhibited, and the glucose consumption was substantially reduced. Meanwhile, the expressions of IRS-1, PI-3K p85 protein and GLUT1 were obviously reduced, while the levels of JNK phosphorylation and IRS-1 Ser307 phosphorylation and the expression of JNK protein were significantly increased, as compared with cells of normal control. However, the aforementioned indices, which indicated the existence of insulin resistance, were reversed by genistein at 1–4 μmol/L in a dose-dependent manner. It was concluded that insulin resistance induced by FFAs in HepG2 hepatocytes could be improved by genistein. Genistein might reverse FFAs-induced insulin resistance in HepG2 cells by targeting JNK.
The therapeutic effects of intensive insulin therapy in treatment of traumatic shock combined with multiple organ dysfunction syndrome (MODS) were investigated. A total of 114 patients with traumatic shock combined with MODS were randomly divided into two groups: control group (n=56) treated with conventional therapy, and intensive insulin therapy group (n=58) treated with conventional therapy plus continuous insulin pumping to control the blood glucose level at range of 4.4–6.1 mmol/L. White blood cells (WBC) counts, prothrombin time (PT), serum creatinine (SCr), alanine aminotransferase (ALT), serum albumin and PaO2 were measured before and at the day 1, 3, 5, 7 and 14 after treatment. The incidence of gastrointestinal dysfunction, the incidence of MODS, hospital stay and the mortality were also observed and compared. After intensive insulin therapy, the WBC counts, SCr, ALT and PT were significantly reduced (P<0.05), but the level of serum albumin was significantly increased (P<0.05) at the day 3, 5, 7 and 14. In the meantime, the PaO2 was significantly elevated at the day 3, 5 and 7 (P<0.01) after intensive insulin therapy. The incidence of gastrointestinal dysfunction, the incidence of MODS, the length of hospital stay and the mortality were markedly decreased (P<0.01). The results suggest early treatment with intensive insulin therapy is effective for traumatic shock combined with MODS and can decrease the length of hospital stay and the mortality.
To assess a novel cell manipulation technique of tissue engineering with respect to its ability to augment superparamagnetic iron oxide particles (SPIO) labeled mesenchymal stem cells (MSCs) density at a localized cartilage defect site in an in vitro phantom by applying magnetic force. Meanwhile, non-invasive imaging techniques were use to track SPIO-labeled MSCs by magnetic resonance imaging (MRI). Human bone marrow MSCs were cultured and labeled with SPIO. Fresh degenerated human osteochondral fragments were obtained during total knee arthroplasty and a cartilage defect was created at the center. Then, the osteochondral fragments were attached to the sidewalls of culture flasks filled with phosphate-buffered saline (PBS) to mimic the human joint cavity. The SPIO-labeled MSCs were injected into the culture flasks in the presence of a 0.57 Tesla (T) magnetic force. Before and 90 min after cell targeting, the specimens underwent T2-weighted turbo spin-echo (SET2WI) sequence of 3.0 T MRI. MRI results were compared with histological findings. Macroscopic observation showed that SPIO-labeled MSCs were steered to the target region of cartilage defect. MRI revealed significant changes in signal intensity (P<0.01). HE staining exibited that a great number of MSCs formed a three-dimensional (3D) cell “sheet” structure at the chondral defect site. It was concluded that 0.57 T magnetic force permits spatial delivery of magnetically labeled MSCs to the target region in vitro. High-field MRI can serve as an very sensitive non-invasive technique for the visualization of SPIO-labeled MSCs.
This study examined the differences in tumor formation of three bladder tumor cell lines (BIU-87, T24 and EJ) after subcutaneously transplanted into nude mice, in order to find the best technique for establishing in vivo bladder tumor model. BIU-87, T24 and EJ cells at logarithmic phase were re-suspended in serum-free medium. The cells suspensions of the identical concentration were subcutaneously transplanted into nude mice and then the success rate and tumor growth were compared among the three cell groups. The results of tumor formation were pathologically evaluated. Lung, liver and kidney tissues were also pathologically examined for distant metastasis. The proliferation of the three cells were determined by immunohistochemically detecting the PCNA expression in the tumors. The results showed that the success rates of EJ and T24 cells were significantly higher than that of BIU-87 cells and no distant metastasis was noted among the three groups. The proliferation levels of EJ and T24 cells was significantly higher than that of BIU-87. But at the later stage of tumor formation, as compared with T24 cells, EJ grew more vigorously, soon resulting in the central necrosis of tumor, which affected the measurement of the actual size of the tumors. Moreover, PCNA staining exhibited that the proliferation of EJ and T24 was significantly higher than that of BIU-87 cells. It is concluded that as compared with BIU-87 cells, EJ and T24 cells had higher success rates, with not significant differences in death rate and distant metastasis found among them. There existed no significant difference in tumor formation between EJ and T24 cells and T24 cells do not rupture easily, which makes it a better cell line for the establishment of in vivo bladder tumor model.
This study examined the association of pregnancy with urolithiasis and provided new insights into urolithiasis in pregnancy. A total of 462 subjects were studied from January 2004 to December 2009 in Foshan Maternal and Child Health Hospital, China. Among the 462 subjects, 162 cases of urolithiasis during pregnancy (UPG) were selected as the observation group, 150 cases of no urolithiasis during pregnancy (NUPG) served as pregnancy control group, and 150 cases of no pregnancy (NPG) at reproductive age who took part in physical examination were randomly assigned into non-pregnant control group. At the same time, the patients in observation group were divided into the following sub-groups: no symptomatic urinary calculus (NSUC) and symptomatic urinary calculus (SUC) groups; SUC group was further divided into surgical intervention (SI) and conservative management (CM) groups. The general information and the data of blood and urine were collected and compared among the groups. The results showed that the incidence of urinary calculi in pregnant women was lower than that in non-pregnant women, the formation of urinary stone was associated with the change of metabolism of protein and sugar in pregnant women, and the surgical intervention was a practicable alternative to treat the clinical intractable symptomatic urinary calculi in pregnancy.
This study was to appraise safety and feasibility of laparoscopic approach and investigate the clinical effects of laparoscopic tension-free repair of esophageal hiatal hernia using mesh. From August 2006 to July 2009, 24 patients with esophageal hiatal hernia underwent laparoscopic repair. Twenty-three patients received laparoscopic tension-free repair using mesh, at the same time, Toupet or Dor partial fundoplication was performed. One patient was converted to open surgery. The average operating time was 90 min (70–210 min) and the blood loss was between 10–110 mL. There was no death. The mean postoperative hospital stay was 5 days (3–30 days). During a follow-up period of 12–20 months (mean 15 months), there was no recurrence of the hernia, and no complication with use of mesh. The present study suggested that laparoscopic approach was secure and minimally invasive operation for esophageal hiatal hernia and the use of mesh could reduce recurrence rate.
During placental development, oxygen environment is not only critical for trophoblasts migration and invasion, but also fundamental for appropriate placental perfusion. Cysteine-rich 61 (Cyr61, CCN1) was expressed in the extravillous trophoblasts (EVTs) and decreased in preeclampsia. Its regulatory properties in human first-trimester extravillous trophoblast cell line (TEV-1 cells) upon a low oxygen tension were investigated. The present study examined functional changes involved in adaptation to hypoxia of the TEV-1 cells, using cobalt chloride (CoCl2) as hypoxic mimic. It was found that hypoxia inhibited growth of TEV-1 cells and induced the increase of cell apoptosis (P<0.05). The Cyr61 expression in human EVTs was transcriptionally induced by CoCl2. Inappropriate EVTs apoptosis has been implicated in the failure of trophoblasts to fully invade and modify the uterine environment and Cyr61 down-regulation, potentially leading to preeclampsia.
This study investigated the role of netrin-1 in placental vascular development. In vitro rat aortic ring assay and in vivo Matrigel plug assay were conducted to exmaine the effect of netrin-1 on angiogenesis. Human placental microvascular endothelial cells (HPMECs) were isolated and cultured and their viability, migration and tubular formation were studied, in order to examine the effects of netrin-1. The results showed that netrin-1 potently stimulated neovascularization in a mouse Matrigel plug in vivo and the sprouting of endothelial cells in rat aortic rings in vitro. In addition, netrin-1 enhanced the viability, migration and tube formation of HPMECs. Our study suggested that netrin-1 could significantly promote the formation of blood vessels of human placenta and may be a potential target for developing new therapeutic strategies for placental vasculature-related diseases.
This study examined the effects of retinoic acid (RA), PD98059, SP600125 and SB203580 on the hyperoxia-induced expression and regulation of matrix metalloproteinase-2 (MMP-2) and metalloproteinase-2 (TIMP-2) in premature rat lung fibroblasts (LFs). LFs were exposed to hyperoxia or room air for 12 h in the presence of RA and the kinase inhibitors PD98059 (ERK1/2), SP600125 (JNK1/2) and SB203580 (p38) respectively. The expression levels of MMP-2 and TIMP-2 mRNA were detected by semi-quantitative reverse transcription polymerase chain reaction (RT-PCR). MMP-2 activity was measured by zymography. The amount of p-ERK1/2, REK1/2, p-JNK1/2, JNK1/2, p-p38 and p38 was determined by Western blotting. The results showed that: (1) PD98059, SP600125 and SB203580 significantly inhibited p-ERK1/2, p-JNK1/2 and p-p38 respectively in LFs; (2) The expression of MMP-2 mRNA in LFs exposed to hyperoxia was decreased after treatment with RA, SP600125 and SB203580 respectively (P<0.01 or 0.05), but did not change after treatment with PD98059 (P>0.05). Meanwhile, RA, PD98059, SP600125 and SB203580 had no effect on the expression of TIMP-2 mRNA in LFs exposed to room air or hyperoxia (P>0.05); (3) The expression of pro- and active MMP-2 experienced no change after treatment with RA or SP600125 in LFs exposed to room air (P>0.05), but decreased remarkably after hyperoxia (P<0.01 or 0.05). SB203580 inhibited the expression of pro- and active MMP-2 either in room air or under hyperoxia (P<0.01). PD98059 exerted no effect on the expression of pro- and active MMP-2 (P<0.05). It was suggested that RA had a protective effect on hyperoxia-induced lung injury by down-regulating the expression of MMP-2 through decreasing the JNK and p38 activation in hyperoxia.
This study investigated the effects of hyperoxia on dynamic changes of thioredoxin-1 (Trx1) and thioredoxin reductase-1 (TrxR1) in alveolar type II epithelial cells (AECII) of premature rats. Pregnant Sprague-Dawley rats were sacrificed on day 19 of gestation. AECII were isolated and purified from the lungs of premature rats. When cultured to 80% confluence, in vitro cells were randomly divided into air group and hyperoxia group. Cells in the hyperoxia group were continuously exposed to 95% O2/5% CO2 and those in the air group to 95% air/5% CO2. After 12, 24 and 48 h, cells in the two groups were harvested to detect their reactive oxygen species (ROS), apoptosis, TrxR1 activity and the expressions of Trx1 and TrxR1 by corresponding protocols, respectively. The results showed that AEC II exposed to hyperoxia generated excessive ROS and the apoptosis percentage in the hyperoxia group was increased significantly at each time points as compared with that in the air group (P<0.001). Moreover, TrxR1 activity was found to be markedly depressed in the hyperoxia group in comparison to that in the air group (P<0.001). RT-PCR showed the expressions of both Trx1 and TrxR1 mRNA were significantly increased in AECII exposed to hyperoxia for 12 and 24 h (P<0.01), respectively. At 48 h, the level of Trx1 mRNA as well as that of TrxR1 mRNA in the hyperoxia group was reduced and showed no significant difference from that in the air group (P>0.05). Western blotting showed the changes of Trx1 protein expressions in the hyperoxia group paralleled those of Trx1 mRNA expressions revealed by RT-PCR. It was concluded that hyperoxia can up-regulate the protective Trx1/TrxR1 expressed by AECII in a certain period, however, also cause dysfunction of the cytoplasmic thioredoxin system by decreasing TrxR1 activity, which may contribute to the progression of oxidative stress and cell apoptosis and finally result in lung injury.
This study compared the efficacy of non-penetrating trabecular surgery and trabeculectomy for the treatment of open angle glaucoma. We searched the Cochrane Library, PUBMED (1966 to 2009), Embase (1980 to 2009) and CMB-disk (1979 to 2009) for the randomized clinical trials (RCT) concerning the two treatment strategies. The reports, including the papers listed in bibliographies, were evaluated against a set of quality criteria and the RCTs that satisfied the criteria were selected and subjected to Meta analysis by employing the Cochrane Collaboration’s RevMan 4.5 software package. A total of nine RCTs were included in the study. The analyses of the reports showed that, 12 months after surgery, there was significant difference in the reduction of interocular pressure (IOP) between non-penetrating trabecular surgery and trabeculectomy (Z=6.05 P<0.00001). There also existed statistically significant difference in the reduction of IOP at the censored time between the two procedures (Z=4.92, P<0.00001). Difference in the success rate was also found between the two surgeries (Z=3.82, P=0.0001). It is concluded that, compared with the non-penetrating trabeculectomy, the traditional trabeculectomy could reduce IOP more and had higher success rate while the non-penetrating trabecular surgery is associated with lower postoperative complications.
A simple method has been proposed for the determination of clozapine (CLZ) and chlorpromazine (CPZ) in human urine by dispersive liquid-liquid microextraction (DLLME) in combination with high-performance liquid chromatography-ultraviolet detector (HPLC-UV). All important variables influencing the extraction efficiency, such as pH, types of the extraction solvent and the disperser solvent and their volume, ionic strength and centrifugation time were investigated and optimized. Under the optimal conditions, the limit of detection (LODs) and quantification (LOQs) of the method were 13 and 39 ng/mL for CLZ, and 2 and 6 ng/mL for CPZ, respectively. The relative standard deviations (RSDs) of the targets were less than 5.1% (C=0.100 μg/mL, n=9). Good linear behaviors over the tested concentration ranges were obtained with the values of R2>0.999 for the targets. The absolute extraction efficiencies of CLZ and CPZ from the spiked blank urine samples were 98.3% and 97.8%, respectively. The applicability of the technique was validated by analyzing urine samples and the mean recoveries for spiked urine samples ranged from 93.3% to 105.0%. The method was successfully applied for the determination of CLZ and CPZ in real human urine.