The novel GATA1-interacting protein HES6 is an essential transcriptional cofactor for human erythropoiesis.

Normal erythropoiesis requires the precise regulation of gene expression patterns, and transcription cofactors play a vital role in this process. Deregulation of cofactors has emerged as a key mechanism contributing to erythroid disorders. Through gene expression profiling, we found HES6 as an abundant cofactor expressed at gene level during human erythropoiesis. HES6 physically interacted with GATA1 and influenced the interaction of GATA1 with FOG1. Knockdown of HES6 impaired human erythropoiesis by decreasing GATA1 expression. Chromatin immunoprecipitation and RNA sequencing revealed a rich set of HES6- and GATA1-co-regulated genes involved in erythroid-related pathways. We also discovered a positive feedback loop composed of HES6, GATA1 and STAT1 in the regulation of erythropoiesis. Notably, erythropoietin (EPO) stimulation led to up-regulation of these loop components. Increased expression levels of loop components were observed in CD34+ cells of polycythemia vera patients. Interference by either HES6 knockdown or inhibition of STAT1 activity suppressed proliferation of erythroid cells with the JAK2V617F mutation. We further explored the impact of HES6 on polycythemia vera phenotypes in mice. The identification of the HES6-GATA1 regulatory loop and its regulation by EPO provides novel insights into human erythropoiesis regulated by EPO/EPOR and a potential therapeutic target for the management of polycythemia vera.

Health economics study of paliperidone palmitate in the treatment of schizophrenia: a 12-month cohort study.

BACKGROUND: To analyze the economic benefits of paliperidone palmitate in the treatment of schizophrenia. METHODS: We collected 546 patients who met the diagnostic criteria for schizophrenia according to the 《International Statistical Classification of Diseases and Related Health Problems,10th》(ICD-10). We gathered general population data such as gender, age, marital status, and education level, then initiated treatment with paliperidone palmitate. Then Follow-up evaluations were conducted at 1, 3, 6, 9, and 12 months after the start of treatment to assess clinical efficacy, adverse reactions, and injection doses. We also collected information on the economic burden before and after 12 months of treatment, as well as the number of outpatient visits and hospitalizations in the past year to analyze economic benefits. RESULTS: The baseline patients totaled 546, with 239 still receiving treatment with paliperidone palmitate 12 months later. After 12 months of treatment, the number of outpatient visits per year increased compared to before (4 (2,10) vs. 12 (4,12), Z=-5.949, P < 0.001), while the number of hospitalizations decreased (1 (1,3) vs. 1 (1,2), Z = 5.625, P < 0.001). The inpatient costs in the direct medical expenses of patients after 12 months of treatment decreased compared to before (5000(2000,12000) vs. 3000 (1000,8050), P < 0.05), while there was no significant change in outpatient expenses and direct non-medical expenses (transportation, accommodation, meal, and family accompanying expenses, etc.) (P > 0.05); the indirect costs of patients after 12 months of treatment (lost productivity costs for patients and families, economic costs due to destructive behavior, costs of seeking non-medical assistance) decreased compared to before (300(150,600) vs. 150(100,200), P < 0.05). CONCLUSION: Palmatine palmitate reduces the number of hospitalizations for patients, as well as their direct and indirect economic burdens, and has good economic benefits.

Comprehensive proteomic analysis reveals dynamic phospho-profiling in human early erythropoiesis.

Normal early erythropoiesis depends on the precise regulation of protein expression and phosphorylation modification. Dysregulation of protein levels or modification contributes to erythroid disorders. To date, the dynamics of protein phosphorylation profiling across human erythroid development is not fully understood. Here, we characterized quantitative proteomic and phosphoproteomic profiling by tandem mass-tagging technology. We systemically built phospho-expression profiling and expression clusters of 11 414 phosphopeptides for human early erythropoiesis. The standardization methods for multitier integrative analyses revealed multiple functional modules of phosphoproteins (e.g., regulation of the G2/M transition) and active phosphorylated signalling (e.g., cell cycle-related pathways). Our further analysis revealed that CDK family members were the main kinases that phosphorylate substrates in erythroid progenitors and identified that CDK9 played an important role in the proliferation of erythroid progenitors. Collectively, our phosphoproteomic profiling, integrative network analysis and functional studies define landscapes of the phosphoproteome and reveal signalling pathways that are involved in human early erythropoiesis. This study will serve as a valuable resource for further investigations of phosphatase and kinase functions in human erythropoiesis and erythroid-related diseases.

The regulation of NONO by USP11 via deubiquitination is linked to the proliferation of melanoma cells.

Ubiquitin-specific protease 11 (USP11) has been implicated in the regulation of DNA repair, apoptosis, signal transduction and cell cycle. It belongs to a USP subfamily of deubiquitinases. Although previous research has shown that USP11 overexpression is frequently found in melanoma and is correlated with a poor prognosis, the potential molecular mechanism of USP11 in melanoma remains indefinitive. Here, we report that USP11 and NONO colocalize and interact with each other in the nucleus of melanoma cells. As a result, the knockdown of USP11 decreases NONO levels. Whereas, overexpression of USP11 increases NONO levels in a dose-dependent manner. Furthermore, we reveal that USP11 protects NONO protein from proteasome-mediated degradation by removing poly-ubiquitin chains conjugated onto NONO. Functionally, USP11 mediated melanoma cell proliferation via the regulation of NONO levels because ablation of USP11 inhibits the proliferation which could be rescued by ectopic expression of NONO protein. Moreover, a significant positive correlation between USP11 and NONO concentrations was found in clinical melanoma samples. Collectively, these results demonstrate that USP11 is a new deubiquitinase of NONO and that the signalling axis of USP11-NONO is significantly involved in melanoma proliferation.

Unexpected role for p19INK4d in posttranscriptional regulation of GATA1 and modulation of human terminal erythropoiesis.

Terminal erythroid differentiation is tightly coordinated with cell-cycle exit, which is regulated by cyclins, cyclin-dependent kinases, and cyclin-dependent kinase inhibitors (CDKI), yet their roles in erythropoiesis remain to be fully defined. We show here that p19INK4d, a member of CDKI family, is abundantly expressed in erythroblasts and that p19INK4d knockdown delayed erythroid differentiation, inhibited cell growth, and led to increased apoptosis and generation of abnormally nucleated late-stage erythroblasts. Unexpectedly, p19INK4d knockdown did not affect cell cycle. Rather, it led to decreased expression of GATA1 protein. Importantly, the differentiation and nuclear defects were rescued by ectopic expression of GATA1. Because the GATA1 protein is protected by nuclear heat shock protein family (HSP) member HSP70, we examined the effects of p19INK4d knockdown on HSP70 and found that p19INK4d knockdown led to decreased expression of HSP70 and its nuclear localization. The reduced levels of HSP70 are the result of reduced extracellular signal-regulated kinase (ERK) activation. Further biochemical analysis revealed that p19INK4d directly binds to Raf kinase inhibitor PEBP1 and that p19INK4d knockdown increased the expression of PEBP1, which in turn led to reduced ERK activation. Thus we have identified an unexpected role for p19INK4d via a novel PEBP1-p-ERK-HSP70-GATA1 pathway. These findings are likely to have implications for improved understanding of disordered erythropoiesis.

Targeting c-met receptor tyrosine kinase by the DNA aptamer SL1 as a potential novel therapeutic option for myeloma.

Hepatocyte growth factor (HGF)/c-met pathway activation has been implicated in the pathogenesis of multiple myeloma (MM), and blocking this pathway has been considered a rational therapeutic strategy for treating MM. Aptamers are single-stranded nucleic acid molecules that fold into complex 3D structures and bind to a variety of targets. Recently, it was reported that DNA aptamer SL1 exhibited high specificity and affinity for c-met and inhibited HGF/c-met signaling in SNU-5 cells. However, as the first c-met-targeted DNA aptamer to be identified, application of SL1 to myeloma treatment requires further investigation. Here, we explore the potential application of SL1 in MM. Our results indicated that c-met expression is gradually increased in MM patients and contributes to poor outcomes. SL1 selectively bound to c-met-positive MM cells but not to normal B cells and suppressed the growth, migration and adhesion of MM cells in vitro in a co-culture model performed with HS5 cells, wherein SL1 inhibited HGF-induced activation of c-met signaling. In vivo and ex vivo fluorescence imaging showed that SL1 accumulated in the c-met positive tumour areas. In addition, SL1 was active against CD138+ primary MM cells and displayed a synergistic inhibition effect with bortezomib. Collectively, our data suggested that SL1 could be beneficial as a c-met targeted antagonist in MM.

Conformational Heterogeneity and FRET Data Interpretation for Dimensions of Unfolded Proteins.

A mathematico-physically valid formulation is required to infer properties of disordered protein conformations from single-molecule Förster resonance energy transfer (smFRET). Conformational dimensions inferred by conventional approaches that presume a homogeneous conformational ensemble can be unphysical. When all possible-heterogeneous as well as homogeneous-conformational distributions are taken into account without prejudgment, a single value of average transfer efficiency 〈E〉 between dyes at two chain ends is generally consistent with highly diverse, multiple values of the average radius of gyration 〈Rg〉. Here we utilize unbiased conformational statistics from a coarse-grained explicit-chain model to establish a general logical framework to quantify this fundamental ambiguity in smFRET inference. As an application, we address the long-standing controversy regarding the denaturant dependence of 〈Rg〉 of unfolded proteins, focusing on Protein L as an example. Conventional smFRET inference concluded that 〈Rg〉 of unfolded Protein L is highly sensitive to [GuHCl], but data from SAXS suggested a near-constant 〈Rg〉 irrespective of [GuHCl]. Strikingly, our analysis indicates that although the reported 〈E〉 values for Protein L at [GuHCl] = 1 and 7 M are very different at 0.75 and 0.45, respectively, the Bayesian Rg2 distributions consistent with these two 〈E〉 values overlap by as much as 75%. Our findings suggest, in general, that the smFRET-SAXS discrepancy regarding unfolded protein dimensions likely arise from highly heterogeneous conformational ensembles at low or zero denaturant, and that additional experimental probes are needed to ascertain the nature of this heterogeneity.

Conformations of a Metastable SH3 Domain Characterized by smFRET and an Excluded-Volume Polymer Model.

Conformational states of the metastable drkN SH3 domain were characterized using single-molecule fluorescence techniques. Under nondenaturing conditions, two Förster resonance energy transfer (FRET) populations were observed that corresponded to a folded and an unfolded state. FRET-estimated radii of gyration and hydrodynamic radii estimated by fluorescence correlation spectroscopy of the two coexisting conformations are in agreement with previous ensemble x-ray scattering and NMR measurements. Surprisingly, when exposed to high concentrations of urea and GdmCl denaturants, the protein still exhibits two distinct FRET populations. The dominant conformation is expanded, showing a low FRET efficiency, consistent with the expected behavior of a random chain with excluded volume. However, approximately one-third of the drkN SH3 conformations showed high, nearly 100%, FRET efficiency, which is shown to correspond to denaturation-induced looped conformations that remain stable on a timescale of at least 100 µs. These loops may contain interconverting conformations that are more globally collapsed, hairpin-like, or circular, giving rise to the observed heterogeneous broadening of this population. Although the underlying mechanism of chain looping remains elusive, FRET experiments in formamide and dimethyl sulfoxide suggest that interactions between hydrophobic groups in the distal regions may play a significant role in the formation of the looped state.

Polycation-π interactions are a driving force for molecular recognition by an intrinsically disordered oncoprotein family.

Molecular recognition by intrinsically disordered proteins (IDPs) commonly involves specific localized contacts and target-induced disorder to order transitions. However, some IDPs remain disordered in the bound state, a phenomenon coined "fuzziness", often characterized by IDP polyvalency, sequence-insensitivity and a dynamic ensemble of disordered bound-state conformations. Besides the above general features, specific biophysical models for fuzzy interactions are mostly lacking. The transcriptional activation domain of the Ewing's Sarcoma oncoprotein family (EAD) is an IDP that exhibits many features of fuzziness, with multiple EAD aromatic side chains driving molecular recognition. Considering the prevalent role of cation-π interactions at various protein-protein interfaces, we hypothesized that EAD-target binding involves polycation- π contacts between a disordered EAD and basic residues on the target. Herein we evaluated the polycation-π hypothesis via functional and theoretical interrogation of EAD variants. The experimental effects of a range of EAD sequence variations, including aromatic number, aromatic density and charge perturbations, all support the cation-π model. Moreover, the activity trends observed are well captured by a coarse-grained EAD chain model and a corresponding analytical model based on interaction between EAD aromatics and surface cations of a generic globular target. EAD-target binding, in the context of pathological Ewing's Sarcoma oncoproteins, is thus seen to be driven by a balance between EAD conformational entropy and favorable EAD-target cation-π contacts. Such a highly versatile mode of molecular recognition offers a general conceptual framework for promiscuous target recognition by polyvalent IDPs.

[Mycoplasma pneumoniae infection and drug resistance in children: an analysis of 1026 cases].

OBJECTIVE: To investigate the Mycoplasma pneumoniae (MP) infection and drug resistance in children with respiratory tract infection and to provide a rational basis for the clinical diagnosis and treatment of MP infection. METHODS: Throat swabs were collected from 3529 children with respiratory tract infection, who visited the pediatric outpatient department or received treatment in the pediatric ward of our hospital from September 2010 to September 2011. The swabs were cultured to detect MP. The drug sensitivity of MP to azithromycin, roxithromycin, erythromycin, acetylspiramycin and clarithromycin was evaluated. RESULTS: Of the 3529 children with respiratory tract infection, 1026 (29.07%) were MP-positive. There were cases of MP infection in all four seasons of the year but infection rates in summer and autumn were significantly higher than in spring and winter (P < 0.05). The infection rate in females was higher than in males (30.43% vs 28.32%; P > 0.05). The infection rate was negatively correlated with age in these children, and there were significant differences in the infection rate among all age groups (P < 0.05). For macrolide antibiotics suitable for children, the cultured MP developed the highest resistance to roxithromycin, followed by erythromycin, acetylspiramycin, clarithromycin, and azithromycin, with significant differences among them (P < 0.01). CONCLUSIONS: MP infection rate is very high among children with respiratory tract infection. The incidence of MP infection is relatively low among school-age children and children are more susceptible to MP infection in summer and autumn than in spring and winter. Throat swabs should be cultured and drug sensitivity tests should be performed as early as possible in children with respiratory tract infection, so that proper intervention can be undertaken in time to reduce drug-resistant strains of MP.

Small-Angle X-ray Scattering Signatures of Conformational Heterogeneity and Homogeneity of Disordered Protein Ensembles.

An accurate account of disordered protein conformations is of central importance to deciphering the physicochemical basis of biological functions of intrinsically disordered proteins and the folding-unfolding energetics of globular proteins. Physically, disordered ensembles of nonhomopolymeric polypeptides are expected to be heterogeneous, i.e., they should differ from those homogeneous ensembles of homopolymers that harbor an essentially unique relationship between average values of end-to-end distance REE and radius of gyration Rg. It was posited recently, however, that small-angle X-ray scattering (SAXS) data on conformational dimensions of disordered proteins can be rationalized almost exclusively by homopolymer ensembles. Assessing this perspective, chain-model simulations are used to evaluate the discriminatory power of SAXS-determined molecular form factors (MFFs) with regard to homogeneous versus heterogeneous ensembles. The general approach adopted here is not bound by any assumption about ensemble encodability, in that the postulated heterogeneous ensembles we evaluated are not restricted to those entailed by simple interaction schemes. Our analysis of MFFs for certain heterogeneous ensembles with more narrowly distributed REE and Rg indicates that while they deviate from MFFs of homogeneous ensembles, the differences can be rather small. Remarkably, some heterogeneous ensembles with asphericity and REE drastically different from those of homogeneous ensembles can nonetheless exhibit practically identical MFFs, demonstrating that SAXS MFFs do not afford unique characterizations of basic properties of conformational ensembles in general. In other words, the ensemble to MFF mapping is practically many-to-one and likely nonsmooth. Heteropolymeric variations of the REE-Rg relationship were further showcased using an analytical perturbation theory developed here for flexible heteropolymers. Ramifications of our findings for interpretation of experimental data are discussed.

[An epidemiologic survey on blood lead levels in preschool children living in towns of Hunan Province].

OBJECTIVE: To investigate blood lead levels (BLLs) and influencing factors of BLLs among preschool children living in towns of Hunan Province. METHODS: A total of 2 044 preschool children (1,108 boys and 936 girls) from towns of 12 regions in Hunan Province were enrolled by a cluster sampling between September 2008 and June 2009. The average age of the children was 4.4 ± 1.1 years (range 2 to 6 years). BLLs were determined using the atomic absorption spectrographic method. The influencing factors of BLLs were investigated using a standard questionnaire and logistic regression analysis. RESULTS: The mean BLLs of the children were 81.9 ± 34.5 µg/L. BLLs more than 100 µg/ L were noted in 482 children (23.58%). Of the 482 children, 472 (23.09%) showed BLLs of 100-199 µg/L and 10 (0.49%) showed BLLs ≥ 200 µg/L. There were significant differences in the prevalence of elevated BLLs (≥ 100 µg/L) among different age groups (P < 0.01). The prevalence of elevated BLLs in boys (28.99%) was significantly higher than that in girls (21.98%) (P < 0.01). There were significant differences in the prevalence of elevated BLLs in children from different regions (P < 0.01). The logistic regression analysis showed that the male (OR = 1.449, P < 0.01), father's occupational lead exposure (OR = 1.314, P < 0.01)and maternal frequent use of hair dyes (OR = 1.678, P < 0.05) were risk factor for elevated BLLs. CONCLUSIONS: The prevalence of elevated BLLs is higher in preschool children living in towns of Hunan Province and is associated with a child's region and age. The male, father's occupational lead exposure and maternal frequent use of hair dyes are risk factor for elevated BLLs.

Responses of Periphyton Microbial Growth, Activity, and Pollutant Removal Efficiency to Cu Exposure.

Periphyton is an effective matrix for the removal of pollutants in wastewater and has been considered a promising method of bioremediation. However, it still needs to be verified whether periphyton can maintain microbial activity and pollutant removal efficiency when dealing with the influence with complex components, and the underlying mechanisms of periphyton need to be revealed further. Herein, this study investigated the microbial growth, activity and functional responses of periphyton after removal of Cu from wastewater. Results showed that the cultivated periphyton was dominated by filamentous algae, and high Cu removal efficiencies by periphyton were obtained after 108 h treatments. Although 2 mg/L Cu2+ changed the microalgal growth (decreasing the contents of total chlorophyll-a (Chla), the carbon source utilization and microbial metabolic activity in periphyton were not significantly affected and even increased by 2 mg/L Cu2+. Moreover, chemical oxygen demand (COD) removal rates were sustained after 0.5 and 2 mg/L Cu2+ treatments. Our work showed that periphyton had strong tolerance and resistance on Cu stress and is environmentally friendly in dealing with wastewater containing heavy metals, as the microbial functions in pollutant removal could be maintained.

Antitumor Drug Combretastatin-A4 Phosphate Aggravates the Symptoms of Dextran Sulfate Sodium-Induced Ulcerative Colitis in Mice.

Ulcerative colitis (UC) is an idiopathic inflammatory bowel disease (IBD) that causes long-lasting inflammation and ulcers in the innermost lining of the colon and rectum. Previous studies demonstrated that resveratrol suppresses colitis and colon cancer associated with colitis by improving glucose metabolism, but resveratrol use is limited by its low oral bioavailability. Combretastatin-A4 phosphate (CA4P) is a vascular-disrupting agent with antitumor activity. CA4P is structurally similar to resveratrol, but whether CA4P has the same effect as resveratrol on UC is not clear. In this study, we examined the pharmacological effects of CA4P administration on dextran sulfate sodium (DSS)-induced inflammation in a mouse model of UC. C57BL/6 mice were administered 2.5% DSS in the drinking water to induce acute UC. CA4P (11 mg/kg/d) was injected intraperitoneally daily. The Disease Activity Index (DAI) score and histological score were evaluated to determine the severity of UC. Colon tissues and blood samples were collected for histological analyses. The results show that CA4P plus DSS significantly decreased colon length (P < 0.05 versus DSS+PBS group) and body weight (P < 0.001 versus PBS group), while increased spleen weight (P < 0.01 versus DSS+PBS group), DAI score (P < 0.01 versus DSS+PBS group), and histological score (P < 0.01 versus DSS+PBS group). Moreover, CA4P exacerbated the pathological features of colitis and significantly increased proinflammatory cytokines (IL-1ß, IL-6, TNF-α) and inflammatory cells (neutrophil, lymphocyte, monocyte). These findings reveal that CA4P aggravates the symptoms of DSS-induced UC and provide a key reference for the potential of CA4P as an anticancer drug.

Rigidity effect on phase behavior of symmetric ABA triblock copolymers: A Monte Carlo simulation.

The phase behavior of symmetric ABA triblock copolymers containing a semiflexible midblock is studied by lattice Monte Carlo simulation. As the midblock evolves from a fully flexible state to a semiflexible state in terms of increase in its persistence length, different phase behaviors are observed while cooling the system from an infinite high temperature to a temperature below T(ODT) (order-disorder transition temperature). Within the midblock flexibility range we studied (l(p)N(c)

Interfacial Synthesis of Conjugated Two-Dimensional N-Graphdiyne.

We explored the interfacial synthesis of 2D N-graphdiyne films at the gas/liquid and liquid/liquid interfaces. Triazine- or pyrazine-based monomers containing ethynyl group were polymerized through the Glaser coupling reactions at interfaces. Several layered, highly ordered and conjugated 2D N-graphdiyne were obtained. Their structures were characterized by TEM, SEM, AFM, XPS, and Raman spectra. Thin films with minimum thickness of 4 nm could be prepared.

An allosteric conduit facilitates dynamic multisite substrate recognition by the SCFCdc4 ubiquitin ligase.

The ubiquitin ligase SCFCdc4 mediates phosphorylation-dependent elimination of numerous substrates by binding one or more Cdc4 phosphodegrons (CPDs). Methyl-based NMR analysis of the Cdc4 WD40 domain demonstrates that Cyclin E, Sic1 and Ash1 degrons have variable effects on the primary Cdc4WD40 binding pocket. Unexpectedly, a Sic1-derived multi-CPD substrate (pSic1) perturbs methyls around a previously documented allosteric binding site for the chemical inhibitor SCF-I2. NMR cross-saturation experiments confirm direct contact between pSic1 and the allosteric pocket. Phosphopeptide affinity measurements reveal negative allosteric communication between the primary CPD and allosteric pockets. Mathematical modelling indicates that the allosteric pocket may enhance ultrasensitivity by tethering pSic1 to Cdc4. These results suggest negative allosteric interaction between two distinct binding pockets on the Cdc4WD40 domain may facilitate dynamic exchange of multiple CPD sites to confer ultrasensitive dependence on substrate phosphorylation.

Expression of Pdx-1 in bone marrow mesenchymal stem cells promotes differentiation of islet-like cells in vitro.

Bone marrow mesenchymal stem cells (BMSCs) have the ability of self-renewal and multi-directional differentiation. Recent reports showed that BMSCs could differentiate into endocrine cells of pancreas. However, the differentiation is not efficient enough to produce insulin-producing cells for the future therapeutic use. Pdx-1 is a crucial regulator for pancreatic development. Therefore we constructed a eukaryotic expression vector containing Pdx-1 to determine the effect of Pdx-1 expression on differentiation of BMSCs in vitro. The results showed that BMSCs could self-assemble to form functional pancreatic islet-like structures after differentiation in vitro. The proportion of insulin-producing cells differentiated from Pdx-1+BMSCs was 28.23% +/- 2.56%, higher than that from BMSCs transfected with vacant vector and Pdx-1-BMSCs (7.23% +/- 1.56% and 4.08% +/- 2.69% respectively) by flow cytometry. Immunocytochemical examination also testified the expression of multiple beta-cells-specific genes such as insulin, glucagons, somatostatin in differentiated BMSCs. The results also revealed that the expressions of genes mentioned above in Pdx-1+BMSCs were higher than that in Pdx-1-BMSCs, which was confirmed by Western blotting analysis and RT-PCR. Glucose-induced insulin secretion from Pdx-1+BMSCs in 5 mmol/L and 25 mmol/L glocuse was (56.61 +/- 4.82) microU/mL and (115.29 +/- 2.56) microU/mL respectively, which were much higher than those from Pdx-1-BMSCs ((25.53 +/- 6.49) microU/mL and (53.26 +/- 7.56) microU/mL respectively). Grafted animals were able to maintain their body weight and survive for relatively longer periods of time than hyperglycemic sham-grafted controls, which demonstrated an overall beneficial effect of the grafted cells on the health of the animals. These findings thus suggested that exogenous expression of Pdx-1 should provide a promising approach for efficiently producing islet-like cells from BMSCs for the future therapeutic use in diabetic patients.

[Study on molecular mechanism of inhibitory effect of baicalin on proliferation of insulinoma cell line in rats].

OBJECTIVE: To investigate the effect of baicalin on insulinoma cell line and the molecular mechanism involved. METHODS: Light microscope, MTT assay, flow cytometry, gene analysis and Western Blot were applied to investigate the effects of baicalin on the cell proliferation, the cell cycle and the involved molecular mechanism. RESULTS: After treatment with baicalin, the number of cells in mitotic stage and the survival rate of cells obviously decreased, and cell proliferation was inhibited in a drug concentration- and acting time-dependent manner, with the appearance of apoptotic insulinoma cells. During the apoptotic process, the activity of caspase-3 was elevated by baicalin in a time-dependent manner; with the increase of the concentration of baicalin, the number of cells in S-phase obviously decreased from 38.2% to 9.4%, while the percentage of cells in G0/G1 phase increased from 56.4% to 85.9%, indicating cells were arrested in G1-phase. Meanwhile, the activity of cyclin gene promoter obviously declined, and the expression of cyclin reduced remarkably. CONCLUSION: Baicalin could induce apoptosis of insulinoma cells, which might be correlated with the activity of caspase-3, and inhibiting proliferation of insulinoma cells in a concentration- and time-dependent manner, in which the action of baicalin in down-regulating the gene transcription and expression of cyclin may play an important role.

[Protective effects of baicalin pretreatment on hypoxic-ischemic brain damage in neonatal rats].

OBJECTIVE: Previous research suggests that dexamethasone (Dex) pretreatment protects neonatal rats against hypoxic-ischemic brain damage (HIBD). Some of the pharmacological effects of baicalin (a traditional Chinese medicine extracted from Scutellaria baicalensis Georgi) are similar to Dex. This study was designed to explore the effect of baicalin on the neuronal apoptosis following HIBD in neonatal rats. METHODS: Six-day-old Sprague-Dawley rats were randomly assigned into Control (without HI), HIBD, Dex-pretreatment and post-treatment, Baicalin-pretreatment and -post-treatment groups. HIBD was induced by ligating the left common carotid artery, followed by exposure to hypoxia. In the pretreatment groups either baicalin (16 mg/kg) or Dex (0.1 mg/kg) was administered to the rats 24 hrs before HIBD; in the post-treatment groups baicalin or Dex was given 30 minutes after HIBD. The rat pups were sacrificed on postnatal day 10, and brain tissues were harvested. Brain water content was determined, morphological changes were observed under a light microscope, and neuronal apoptosis was measured by terminal deoxynucleotidyl transferase mediated dUTP biotin nick end labeling (TUNEL) staining. RESULTS: The brain water content and the number of apoptotic cells were significantly higher in the HIBD group than those of the Control group (P < 0.05). Both baicalin and Dex pretreatment decreased the brain water content from 88.9 +/- 1.7 % (HIBD group) to 87.4 +/- 0.7% (baicalin) or 87.3 +/- 0.6% (Dex) (P < 0.05) and the number of apoptotic cells were reduced from 251 +/- 28 (HIBD group) to 102 +/- 47 (baicalin) or 75 +/- 26 (Dex) (P < 0.05). Baicalin and Dex post-treatment had no effects on the brain water content and the number of apoptotic cells. Loss and degeneration of neurons could be observed in the HIBD group. Baicalin and Dex pretreatment significantly alleviated neuronal injury, but post-treatment did not. CONCLUSIONS: Pretreatment with baicalin, as with Dex, has a protective effect against HIBD in neonatal rats, but baicalin or Dex post-treatment do not reverse the neuronal injuries.