[Screening of biomarkers in exhaled breath of mice exposed to benzene].

Objective: To screen the biomarkers in the exhaled breath of mice exposed to benzene by using exhaled breath online analysis system. Methods: Thirty 8-week-old male C57BL/6 mice were randomly divided into six groups (0, 3, 32, 324, 648, and 1 296 mg/m3) and treated with benzene vapour for 28 days. At the end of the exposure, the peripheral blood cell counts and blood glutathione (GSH) were detected. The content of malondialdehyde (MDA) in HL60 cells treated by mice plasma was examined. Exhaled breath data from mice were collected by Secondary electrospray ionization source high resolution mass spectrometry (SESI-HRMS). Targeted analysis underlying benzene metabolites and oxidative stress metabolites was performed to screen the biomarkers in exhaled breath. Results: After benzene exposure, the number of peripheral blood cells was decreased in different degrees, particularly in the white blood cells (WBC) number. The WBC in 32 and 324 mg/m3 groups was declined by 27.76% and 52.87%, respectively compared to that in control group (P<0.05). Meanwhile, compared with the control group, the GSH content of peripheral blood cells from 324 mg/m3 group decreased by 13.16% (P<0.05). In addition, MDA content was increased by 18.11% in HL60 cells treated with plasma from 324 mg/m3 group mice (P<0.05). The phenol, hydroquinone/catechol, benzenetriol and trans, trans-Muconic acid (t,t-MA) in the exhaled gas of mice could be used as biomarkers for benzene exposure (R2>0.8, P<0.001). The peak intensity of five small molecular metabolites related to oxidative stress (ω-carboxylic fatty acid C5H10O3, ω-carboxylic fatty acid C6H12O3, glutamate, cysteine and MDA) increased with the increase of benzene concentration (P<0.05), which was negatively correlated with WBC decline (P<0.001), suggesting that these molecules mignt be used as biomarkers of benzene-induced toxicity. Conclusions: Phenol, hydroquinone/catechol, benzenetriol and trans, trans-Muconic acid (t,t-MA) in exhaled breath of mice could be used as biomarkers for benzene exposure; ω-carboxylic fatty acid C5H10O3, ω-carboxylic fatty acid C6H12O3, glutamate, cysteine and MDA might be used as markers of benzene-induced toxicity.

[A case of Caroli's disease confirmed by pathology, atypical symptoms and images].

Caroli's disease is a rare congenital disease characterized by non-obstructive dilatation of the intrahepatic bile ducts, with a prevalence of one in a million in the general population[1]. Most of it is considered to be an autosomal recessive genetic disease, but in many cases, the typical genetic family history cannot be traced back. There are two forms of Caroli's disease: simple type (commonly called Caroli disease) and Caroli syndrome (characterized by congenital liver fibrosis and/or polycystic kidney disease). PKHD1 gene is considered to be the causative gene of Caroli's disease, congenital liver fibrosis and/or polycystic kidney disease [2]. Here, we introduce a case of Caroli's disease confirmed by pathology, atypical symptoms and images in our hospital.

[The effects of ethylbenzene on HEI-OC1 cells proliferation and oxidative stress level].

Objective: To study the changes of proliferation and oxidation indexes of Cochlear hair cell line (HEI-OC1 cells) exposed to ethylbenzene. Methods: From July to December 2019, 11 groups with ethylbenzene concentrations of 0, 30, 60, 90, 300, 600, 900 µmol/L and 3, 6, 9, 10 mmol/L, were used to determine the proliferation activity of HEI-OC1 cells exposed to ethylbenzene for 24 hours, and the cells were treated with 0, 1, 2, 4, 8, 16, 32, 64 mmol/L ethylbenzene for 24 hours, then the 50% inhibitory concentration of ethylbenzene was calculated. After HEI-OC1 cells were exposed to 0, 6, 9 and 12 mmol/L ethylbenzene for 24 hours, the malondialdehyde (MDA) content, superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) activities were measured. Results: Compared with 0 mmol/L concentration group, the survival rate of HEI-OC1 cells at 6, 9, 12 mmol/L concentration was significantly decreased (P<0.01) . The 50% inhibitory concentration of ethylbenzene on HEI-OC1 cells was 12.86 mmol/L (R(2)=99.05) . There were significant differences in SOD and GSH-Px activity in HEI-OC1 cells treated with ethylbenzene at different concentrations (0, 6, 9, 12 mmol/L) for 24 hours (F=65.11, 6.48, 22.85, P<0.05) . Compared with 0 mmol/L concentration group, the MDA content of HEI-OC1 cells was significantly increased in 9 and 12 mmol/L concentration groups, the SOD activity was significantly decreased in 12 mmol/L concentration group, and the GSH-Px activity was significantly decreased in 6 and 12 mmol/L concentration groups. Conclusion: Ethylbenzene can inhibit the proliferation of HEI-OC1 cells and cause oxidative damage.

[The association between mode of action and adverse outcome pathway and its application in risk assessment].

Risk assessment is a necessary technical means to protect human health and environmental safety. Traditional risk assessment based on toxicity data obtained from animal experiments was difficult to meet the need for risk assessment for a large number of chemicals due to the low throughput, long cycle, high cost and uncertainty of extrapolation to human exposure dose. The proposed risk assessment frameworks, the model of action (MOA) and the adverse outcome pathway (AOP), pointed the way for us to develop new and efficient evaluation methods. In this review, the basic concepts and contents of MOA and AOP, as well as the relationship between them, were introduced. Taking acrylamide (AA) as an example, this review briefly described the application of MOA/AOP framework in chemical risk assessment, so as to provide theoretical guidance for better application of MOA/AOP framework in chemical risk assessment.

[Clinicopathologic features of Cronkhite-Canada syndrome and the significance of IgG4-positive plasma cells infiltration].

Objective: To analyze the clinical and pathological features of Cronkhite-Canada syndrome (CCS), and to investigate the significance of IgG4 positive plasma cell infiltration. Methods: Clinical presentations, endoscopic appearances and morphological features of 18 patients diagnosed with CCS at Peking Union Medical College Hospital during 2000-2016 were included in the study.There were 11 male and 7 female patients. IgG4 and IgG immunohistochemical stains were performed in total of 55 biopsies from the patients (36 polyps, 10 adenoma and 9 surrounding mucosa) and a control group of 58 cases (19 colonic mucosa, 7 colonic hyperplastic polyps, 9 inflammatory bowel disease and 23 adenoma). Results: Average age of CCS was 59 years (range 47-69 years) and the male to female ratio was 11∶7. All patients had at least one ectoderm lesion. Fourteen cases had testicular abnormalities. Digestive tract symptoms were encountered in most cases. Four patients had a history of connective tissue disease. Endoscopically, multiple polyps were found to involve entire gastrointestinal tract except the esophagus. Morphologically, CCS polyps were characterized by prominent mucosal edema, mild to moderate inflammation, glands hyperplasia and cystic dilatation. Ten cases had colonic adenoma and one case had rectal adenocarcinoma. While none of the cases reached the diagnostic criteria of IgG4 disease, the positive rate of IgG4 positive plasma cells in adenoma of CCS was significantly higher than that in polyps and surrounding mucosa of CCS (60.0%∶13.9%∶0, P<0.01). The positive rate of IgG4 positive plasma cells of adenoma and polyp of CCS had an increased tendency compared to that of adenoma and polyp of control group (60.0%∶34.8%; 13.9%∶0). Conclusion: The findings in the study do not support that CCS is a variant of IgG4 disease, although IgG4 positive plasma cells may be involved in the pathogenesis of CCS, which may be explored in future investigations.

[Value of DNA content in comet tail in predicting changes in blood cell counts in workers exposed to benzene].

Objective: To investigate the value of DNA content in comet tail (TailDNA) in predicting the changes in peripheral blood cell counts in workers exposed to benzene. Methods: In 2011, cluster sampling was used to select 150 male workers exposed to benzene in a petrochemical factory. Cubital venous blood and urine samples were collected for routine blood rest, comet assay, and measurement of s-phenylmercapturic acid (SPMA) and urine creatinine. The population was divided into groups according to urinary SPMA or TailDNA, and routine blood test results within 3 years were collected to analyze the changes in blood cell counts. Results: The low-SPMA group had significantly higher white blood cell and neutrophilcounts in all years than the high-SPMA group (P<0.05) . The low-Tail DNA group had a significant increase in platelet count from 2012 to 2014 (P<0.05) , while the high-Tail DNA group had no significant change (P>0.05) . During the 4-year period, the high-TailDNA group had a significantly lower red blood cell count than the low-TailDNA group (P<0.05) . The high-TailDNA group showed a gradual reduction in white blood cell count over time (ß=-0.113, P<0.05) , and the low-TailDNA group showed no trend of the reduction in white blood cell count (P>0.05) . Conclusion: TailDNA can be used to predict the changes in blood cell counts in workers exposed to benzene.

Micropapillary bladder cancer: a clinico-pathological characterization and treatment analysis.

PURPOSE: Micropapillary bladder cancer (MPBC) is a very rare and aggressive variant of urothelial carcinoma (UC). The aim of this study was to investigate the clinico-pathological characteristics, treatment, and prognosis of MPBC to improve the understanding of this invasive disease. METHODS: We reviewed the records of 6 patients with MPBC who were evaluated and treated at our hospital between 2009 and 2015, and additionally reviewed 38 cases reported in the literature. RESULTS: In 44 cases, 36 cases (81.8%) were male and 8 cases (18.2%) were female, with a male:female ratio of 4.5:1; the median age of the patients was 68 years (range 45-91 years). A majority (81.8%) of patients with cT1 above or with lymph node and distant metastasis (cT2N0 in 18.2%, cT3-4N0 in 13.6%, cTanyN+ in 43.2%, and cTanyM+ in 6.8%). There was a high grade in 70.5% of patients. Lymphovascular invasion (LVI) was present in 61.4% of patients, and LVI in cT2 was more common than in cT1 (71.4 vs 22.2%). 52.3% of patients were treated with radical cystectomy (RC). After a mean follow-up of 16.2 months, 77.3% of patients developed distant metastases, and 47.7% of patients died of the disease. The mean overall survival (OS) was 28.9 months and the median OS was 20 months, and the amount of micropapillary (MPP) is correlated inversely with prognosis. CONCLUSIONS: Micropapillary bladder cancer is a rare variant of UC associated with a poor prognosis, which often presents at an advanced stage with LVI and distant metastases. The optimal treatment strategy is early RC combined with chemotherapy.

[Comparison of long-term dental treatment effects of children treated under general anesthesia and passive restraint].

Objective: To compare the long-term dental treatment effects, oral health habits and oral-health-related qualities of life of children treated under general anesthesia (GA) and passive restraint (PR), respectively. Methods: Twenty seven 2 to 4-year-old children treated under GA and thirty four children treated under PR were recruited in the Department of Pediatric Dentistry, Peking University School and Hospital of Stomatology. Up to 2 years after the treatment, a follow up assessment was conducted. The data of general information, dental plaque level and the unplanned treatments were recorded and analyzed. The questionnaire of oral health habits and early childhood oral health impact scale (ECOHIS) for each child was also completed and analyzed. The survival rate and median survival time of the deciduous teeth were calculated. Multivariate analysis was performed by using Cox proportional hazard model. Results: Twenty-five children under GA and 32 under PR were finally included, with a total of 1 098 deciduous teeth. The postoperative dental plaque indicesin both GA and PR groups had significantly improved than that of before the treatments (P=0.019, P<0.001). The oral health habits had also improved, and the improvement in PR group was more obvious than that in GA group. Totally 128 teeth (27.0%) appeared unplanned treatments in GA group and 232 teeth (37.2%) in PR group during the follow-ups. The new caries and recurrent caries in PR group were significantly more than that in GA group (P<0.001, P=0.012). No significant differences were found between the two groups in restoration failure, secondary caries and endodontic diseases (P=0.129, P=0.822, P=0.642). However, the time of occurrence of endodontic disease and secondary caries in GA group were significantly longer than that in PR group (P<0.01, P<0.001). The median survival time of teeth in GA group was 1 018 days comparing to 944 days in PR group. The difference was statistically significant (P<0.05). The survival rate was associated with such factors as decayed-missing-filled tooth (dmft), anterior or posterior teeth, feeding frequency, brushing habits and behavior management techniques. Conclusions: The long-term dental treatment effects of children treated under GA was significantly better than that of PR group. Continuous reinforcement of proper dietary and oral hygiene habits might help in maintaining the long-term treatment effect.

[The multicenter study on the registration and follow-up of low anticoagulation therapy for the heart valve operation in China].

OBJECTIVE: To investigate the optimal anticoagulation methods and monitoring strategy for Chinese patients undergoing heart valve replacement, which is potentially quite different from western populations. METHODS: In this multicenter prospective cohort study, the anticoagulation and monitoring strategy data was acquired from 25 773 in-hospital patients in 35 medical centers and 20 519 patients in outpatient clinic in 11 medical centers from January 1st, 2011 to December 31th, 2015. RESULTS: As for in-hospital patients, mean age of study population was (48.6±11.2) years old; main etiology of valve pathology was rheumatic (87.5%) origin among study cohort; 94.8% of study population received mechanical valve implantation; international normalized ratio (INR) monitoring (in all the study centers) and low-intensity anticoagulation strategy (31 hospitals chose target INR range of 1.5-2.5, and actual values of INR among 89.2% of 100 069 in-hospital monitoring samples were 1.5-2.5), with mean actual INR values of 1.84±0.53, and warfarin dosage of (2.82±0.93) mg/d were widely adopted among the study centers; strategies of in-hospital warfarin administration were similar in all the study centers; complication rates of low-intensity anticoagulation strategy were low in severe hemorrhage (0.02%), thrombosis (0.05%), and thromboembolism (0.05%) events, without anticoagulation-related death.As for 18 974 outpatient clinic patients, the follow-up rate was 92.47%, with a total of 30 012 patient-years (Pty). Anticoagulation-related morbidity and mortality rates were 0.67% and 0.15% Pty; major hemorrhage morbidity and mortality rates were 0.25% and 0.13% Pty; thromboembolism morbidity and mortality rates were 0.45% and 0.03% Pty.The mean dosage of warfarin daily dosage was (2.85±1.23) mg/d and INR value was 1.82±0.57.No significant regional difference in the intensity of anticoagulation therapy was noted during the study. CONCLUSIONS: INR can be used as a normalized indicator for intensity of anticoagulation therapy in China.The optimal anticoagulation intensity with INR range from 1.5 to 2.5 is safe and effective for Chinese patients with heart valve replacement, and there is no significant regional difference in the intensity of anticoagulation therapy.

[Children's intelligence quotient following general anesthesia for dental care: a clinical observation by Chinese Wechsler young children scale of intelligence].

OBJECTIVE: It has been demonstrated that anesthetics exposure may lead to neurocognitive impairment in developing brain of animal models. However, for the limitation that the animal models cannot fully mimic the dose and duration in clinical settings especially for dental general anesthesia, the clinical significance of anesthetics exposure on developing central nervous system remains undetermined. Therefore, we conducted the current study in order to observe the fluctuation of intelligence quotient (IQ) after the administration of dental general anesthesia comparing to that before surgery. We conducted the current study in order to observe the fluctuation of intelligence quotient (IQ) after the administration of dental general anesthesia compared with that before surgery. METHODS: Thirty two patients, ASA I, who were exposed to dental general anesthesia in Department of Pediatric Dentistry Peking University School and Hospital of Stomatology, aged 4 to 6.5 years, were enrolled in this prospective study. Patients with severe learning difficulties or communication disorders were excluded. Written and informed consent was obtained from each patients' family which was fully explained of the purpose and method of study. Their intelligence quotients were evaluated with the Chinese Wechsler young children scale of intelligence (Urban version) before and 2 weeks after dental anesthesia. They were treated by experienced pediatric dentists and the sevoflurane, propofol and nitrous oxide were used for general anesthesia by anesthetist. Articaine hydrochloride and epinephrine tartrate injections were used for their pulp treatment or extraction. The examiners and scorers for IQ had technical training in the test administration. All the patients were tested by the same examiner and with standardized guide language. Each subtest was scored according to the tool review. Verbal IQ and performance IQ consisted of relevant 5 subtests and full scale IQ. Statistical analyses were performed by SPSS 18.0. All the scores of subtests and 3 types of IQ were expressed as mean±standard deviation. Paired two-tailed t test was applied and P<0.05 was accepted as statistically significant. RESULTS: The results of intelligent assessment from 28 subjects were collected. The anesthetic time was (163.4±32.6) min and the number of treated teeth was 12.1±2.3, mean age (4.60±0.41) years; age range=4.04 to 5.44 years. Among the patients, there were 13 girls and 15 boys. There was no significant difference in gender distribution. The postoperative full IQ (128.46±10.85) was higher than that before surgery (124.64±11.46, P= 0.017). We found that the elevation of performance IQ, to a large extent, contributed to this change in full IQ (P= 0.007). Correspondingly, there was no statistical difference in the verbal IQ, which was 119.68±11.74 to 120.21±15.61 (P=0.854). CONCLUSION: Dental general anesthesia has no negative effect on the intelligence of preschool children, who were treated under general anesthesia by sevoflurane, propofol and nitrous oxide for 2 to 4 hours. The full IQ and performance IQ were slightly enhanced after treatment which can be explained by the memory effect.

Focusing polycapillary to reduce parasitic scattering for inelastic x-ray measurements at high pressure.

The double-differential scattering cross-section for the inelastic scattering of x-ray photons from electrons is typically orders of magnitude smaller than that of elastic scattering. With samples 10-100 µm size in a diamond anvil cell at high pressure, the inelastic x-ray scattering signals from samples are obscured by scattering from the cell gasket and diamonds. One major experimental challenge is to measure a clean inelastic signal from the sample in a diamond anvil cell. Among the many strategies for doing this, we have used a focusing polycapillary as a post-sample optic, which allows essentially only scattered photons within its input field of view to be refocused and transmitted to the backscattering energy analyzer of the spectrometer. We describe the modified inelastic x-ray spectrometer and its alignment. With a focused incident beam which matches the sample size and the field of view of polycapillary, at relatively large scattering angles, the polycapillary effectively reduces parasitic scattering from the diamond anvil cell gasket and diamonds. Raw data collected from the helium exciton measured by x-ray inelastic scattering at high pressure using the polycapillary method are compared with those using conventional post-sample slit collimation.

New developments in high pressure x-ray spectroscopy beamline at High Pressure Collaborative Access Team.

The 16 ID-D (Insertion Device - D station) beamline of the High Pressure Collaborative Access Team at the Advanced Photon Source is dedicated to high pressure research using X-ray spectroscopy techniques typically integrated with diamond anvil cells. The beamline provides X-rays of 4.5-37 keV, and current available techniques include X-ray emission spectroscopy, inelastic X-ray scattering, and nuclear resonant scattering. The recent developments include a canted undulator upgrade, 17-element analyzer array for inelastic X-ray scattering, and an emission spectrometer using a polycapillary half-lens. Recent development projects and future prospects are also discussed.

Involvement of the mitfa gene in the development of pigment cell in Japanese ornamental (Koi) carp (Cyprinus carpio L.).

A colored phenotype is an important feature of ornamental fish. In mammals, microphthalmia-associated transcription factor (MITF) was found to regulate the development of melanocytes. In this study, the mitfa cDNA was first cloned from the Japanese ornamental (Koi) carp (Cyprinus carpio L.), an important ornamental freshwater fish. The full-length cDNA of the mitfa gene contains 1634 bp, coding for 412 amino acids in Koi. The identity degree of mitfa amino acid sequences between the Koi carp and zebrafish is 92.9%. We tested the expression of the mitfa gene in several varieties of Koi using reverse transcription-polymerase chain reaction and found that the mitfa gene is highly expressed in the skin tissues of the Taisho sanke and the Procypris merus. Interestingly, the mitfa gene was also expressed in the Kohaku and Yamabaki ogon, although melanocytes were not observed in the skin. Koi carp embryos were transparent and colorless, while after hatching, different types of pigment cells successively emerged in a fixed order. In Taisho sanke, melanocytes first appeared in the trunk at approximately 12 days of age. Subsequently, there was a large area of melanocytes by 30 days of age. The expression level of the mitfa mRNA was low in early embryos and newly hatched larvae, and increased to high levels in 30-day-old fry. The results show that the mitfa gene is involved in regulating fish body color in the development of both melanocytes and pigment cells.

Identification and analysis of the jnk1 gene in polyploid hybrids of red crucian carp (Carassius auratus red var.) and common carp (Cyprinus carpio L.).

c-Jun N-terminal kinase (JNK) is an important member of the mitogen-activated protein kinase superfamily. The allotetraploid crucian carp is a product of distant hybridization of female red crucian carp with male common carp. It is the first natural case of an allotetraploid with stable genetic characters, including fertility of both female and male animals. In this study, 2 jnk1 cDNAs (including jnk1a and jnk1b) have been cloned from the polyploid crucian carp system, consisting of the allotetraploid crucian carp, the triploid crucian carp, and their original parents (red crucian and common carp). We show that jnk1a and jnk1b represent 2 splice forms arising from the jnk1 gene. On the basis of the genetic structure of jnk1a gene in the polyploid crucian carp system, we demonstrated that the allotetraploid crucian carp is phylogenetically closer to its paternal parent (common carp) than to its maternal parent. We further show a similarity between the triploid crucian carp and its original female parent (red crucian carp). Comparisons of genetic structures indicated that the jnk1b genes of allotetraploid and triploid crucian carp are more similar to those of the original paternal parent rather than the original female parent (red crucian carp). RT-PCR analysis indicated that both the jnk1a and jnk1b genes are widely expressed in fish embryos and in the adult organs, displaying distinct features of embryonic-stage and organ specificity in the polyploid crucian carp system.

Magnetic and structural transitions of SrFe2As2 at high pressure and low temperature.

One of key issues in studying iron based superconductors is to understand how the magnetic phase of the parent compounds evolves. Here we report the systematic investigation of paramagnetic to antiferromagnetic and tetragonal to orthorhombic structural transitions of "122" SrFe2As2 parent compound using combined high resolution synchrotron Mössbauer spectroscopy and x-ray diffraction techniques in a cryogenically cooled high pressure diamond anvil cell. It is found that although the two transitions are coupled at 205 K at ambient pressure, they are concurrently suppressed to much lower temperatures near a quantum critical pressure of approximately 4.8 GPa where the antiferromagnetic state transforms into bulk superconducting state. Our results indicate that the lattice distortions and magnetism jointly play a critical role in inducing superconductivity in iron based compounds.

Pressure-decoupled magnetic and structural transitions of the parent compound of iron-based 122 superconductors BaFe2As2.

The recent discovery of iron ferropnictide superconductors has received intensive concern in connection with magnetically involved superconductors. Prominent features of ferropnictide superconductors are becoming apparent: the parent compounds exhibit an antiferromagnetic ordered spin density wave (SDW) state, the magnetic-phase transition is always accompanied by a crystal structural transition, and superconductivity can be induced by suppressing the SDW phase via either chemical doping or applied external pressure to the parent state. These features generated considerable interest in the interplay between magnetism and structure in chemically doped samples, showing crystal structure transitions always precede or coincide with magnetic transition. Pressure-tuned transition, on the other hand, would be more straightforward to superconducting mechanism studies because there are no disorder effects caused by chemical doping; however, remarkably little is known about the interplay in the parent compounds under controlled pressure due to the experimental challenge of in situ measuring both of magnetic and crystal structure evolution at high pressure and low temperatures. Here we show from combined synchrotron Mössbauer and X-ray diffraction at high pressures that the magnetic ordering surprisingly precedes the structural transition at high pressures in the parent compound BaFe2As2, in sharp contrast to the chemical-doping case. The results can be well understood in terms of the spin fluctuations in the emerging nematic phase before the long-range magnetic order that sheds light on understanding how the parent compound evolves from a SDW state to a superconducting phase, a key scientific inquiry of iron-based superconductors.

A versatile medium-resolution x-ray emission spectrometer for diamond anvil cell applications.

We present design and performance details for a polycapillary-coupled x-ray spectrometer that provides very high collection efficiency at a moderate energy resolution suitable for many studies of nonresonant x-ray emission spectroscopy, especially for samples of heavy elements under high pressures. Using a single Bragg analyzer operating close to backscattering geometry so as to minimize the effect of the weak divergence of the quasicollimated exit beam from the polycapillary optic, this instrument can maintain a typical energy resolution of 5 eV over photon energies from 5 keV to 10 keV. We find dramatically improved count rates as compared to a traditional higher-resolution instrument based on a single spherically bent crystal analyzer.

The PP2A-Aß gene is regulated by multiple transcriptional factors including Ets-1, SP1/SP3, and RXRα/ß.

Protein phosphatase-2A (PP-2A) is a major serine/threonine phosphatase abundantly expressed in eukaryotes. PP-2A is a heterotrimer that contains a 65 kD scaffold A subunit, a 36 kD catalytic C subunit, and a regulatory B subunit of variable isoforms ranging from 54-130 kDs. The scaffold subunits, PP2A-Aα/ß, act as platforms for both the C and B subunits to bind, and thus are key structural components for PP-2A activity. Mutations in both genes encoding PP2A-Aα and PP2A-Aß lead to carcinogenesis and likely other human diseases. Our previous work showed that the gene coding for PP2A-Aα is positively regulated by multiple transcription factors including Ets-1, CREB, and AP-2α but negatively regulated by SP-1/SP-3. In the present study, we have functionally dissected the promoter of the mouse PP2A-Aß gene. Our results demonstrate that three major cis-elements, including the binding sites for Ets-1, SP1/SP3, and RXRα/ß, are present in the proximal promoter of the mouse PP2A-Aß gene. Gel mobility shifting assays reveal that Ets-1, SP1/SP3, and RXRα/ß all bind to PP2A-Aß gene promoter. In vitro mutagenesis and reporter gene activity assays demonstrate that while Ets-1 displays negative regulation, SP1/SP3 and RXRα/ß positively regulate the promoter of the PP2A-Aß gene. Co-expression of the cDNAs encoding Ets-1, SP1/SP3, or RXRα/ß and the luciferase reporter gene driven by PP2A-Aß promoter further confirm their control over the PP2A-Aß promoter. Finally, ChIP assays demonstrate that Ets-1, SP1/SP3, and RXRα/ß can all bind to the PP2A-Aß gene promoter. Together, our results reveal that multiple transcription factors regulate the PP2A-Aß gene. Moreover, our results provide important information explaining why PP2A-Aα and PP2A-Aß display distinct expression levels.

The p53-Bak apoptotic signaling axis plays an essential role in regulating differentiation of the ocular lens.

The tumor suppressor p53 is a master regulator of apoptosis and also plays a key role in cell cycle checking. In our previous studies, we demonstrated that p53 directly regulates Bak in mouse JB6 cells (Qin et al. 2008. Cancer Research. 68(11):4150) and that p53-Bak signaling axis plays an important role in mediating EGCG-induced apoptosis. Here, we demonstrate that the same p53-Bak apoptotic signaling axis executes an essential role in regulating lens cell differentiation. First, during mouse lens development, p53 is expressed and differentially phosphorylated at different residues. Associated with p53 expression, Bak is also significantly expressed during mouse lens development. Second, human p53 directly regulates Bak promoter and Bak expression in p53 knockout mice (p53-/-) was significantly downregulated. Third, during in vitro bFGF-induced lens cell differentiation, knockdown of p53 or Bak leads to significant inhibition of lens cell differentiation. Fourth, besides the major distribution of Bak in cytoplasm, it is also localized in the nucleus in normal lens or bFGF-induced differentiating lens cells. Finally, p53 and Bak are co-localized in both cytoplasm and nucleus, and their interaction regulates the stability of p53. Together, these results demonstrate for the first time that the p53-Bak apoptotic signaling axis plays an essential role in regulating lens differentiation.

Upregulation of miR-27a contributes to the malignant transformation of human bronchial epithelial cells induced by SV40 small T antigen.

The introduction of the Simian virus 40 (SV40) early region, the telomerase catalytic subunit (hTERT) and an oncogenic allele of H-Ras directly transforms primary human cells. SV40 small T antigen (ST), which forms a complex with protein phosphatase 2A (PP2A) and inhibits PP2A activity, is believed to have a critical role in the malignant transformation of human cells. Recent evidence has shown that aberrant microRNA (miRNA) expression patterns are correlated with cancer development. Here, we identified miR-27a as a differentially expressed miRNA in SV40 ST-expressing cells. miR-27a is upregulated in SV40 ST-transformed human bronchial epithelial cells (HBERST). Suppression of miR-27a expression in HBERST cells or lung cancer cell lines (NCI-H226 and SK-MES-1) that exhibited high levels of miR-27a expression lead to cell growth arrested in the G(0)-G(1) phase. In addition, suppression of miR-27a in HBERST cells attenuated the capacity of such cells to grow in an anchorage-independent manner. We also found that suppression of the PP2A B56γ expression resulted in upregulation of miR-27a similar to that achieved by the introduction of ST, indicating that dysregulation of miR-27a expression in ST-expressing cells was mediated by the ST-PP2A interaction. Moreover, we discovered that Fbxw7 gene encoding F-box/WD repeat-containing protein 7 was a potential miR-27a target validated by dual-luciferase reporter system analysis. The inverse correlation between miR-27a expression levels and Fbxw7 protein expression was further confirmed in both cell models and human tumor samples. Fbxw7 regulates cell-cycle progression through the ubiquitin-dependent proteolysis of a set of substrates, including c-Myc, c-Jun, cyclin E1 and Notch 1. Thus, promotion of cell growth arising from the suppression of Fbxw7 by miR-27a overexpression might be responsible for the viral oncoprotein ST-induced malignant transformation. These observations demonstrate that miR-27a functions as an oncogene in human tumorigenesis.