The protective effect of Wnt3a on inflammatory response in oxygen-glucose deprivation/reoxygenation (OGD/R) astrocyte model.

Involving in the immune response after cerebral infarction, astrocytes could secrete large amounts of pro- and anti-inflammatory factors. The aim of this study is to investigate the effect of Wnt3a intervention on the inflammatory response of oxygen-glucose deprivation (OGD) followed by reoxygenation (OGD/R) astrocyte model, and to provide a new target for immunoprotective treatment of cerebral infarction. We constructed the OGD/R rat astrocyte model, the astrocytes were treated by different concentrations of glucose (25, 50, 100 mM) intervened with/without Wnt3a (25 µg/ml). Microscope was used to observe the cell survival in rat astrocytes. The relative expression of inflammatory factors (TNF-a, IL-6, HIF-a) in rat astrocytes was detected by qRT-PCR. The expression of inflammatory factors such as TNF-a, IL-6 and HIF-a in rat astrocytes was increased after OGD/R treatment. The Wnt3a intervention promoted cell survival and decreased the expression of inflammatory factors in rat astrocytes induced by OGD/R. There is a neuroprotective effect that Wnt3a intervention could reduce inflammatory response in the OGD/R rat astrocyte model.

Beware of the DeBakey type I aortic dissection hidden by ischemic stroke: Two case reports.

BACKGROUND: DeBakey type I aortic dissection is one of the rare etiologies of ischemic stroke. It is critical to identify arterial dissection before intravenous thrombolysis; otherwise, fatal hemorrhage may occur. CASE SUMMARY: In this report, we described 2 painless DeBakey type I aortic dissection cases with initial symptoms similar to ischemic stroke. Sudden onset of conscious disturbance and limb weakness within minutes occurred in both cases. Hypotension was found in both cases. Thoracoabdominal computed tomography angiography was urgently performed due to unknown reason hypotension, and DeBakey type I aortic dissection was confirmed. Intravenous thrombolysis was avoided because of timely diagnosis; however, they both eventually died of ruptured aortic dissection. CONCLUSION: Aortic dissection should always be excluded in ischemic stroke patients with unexplained hypotension or shock symptoms before intravenous thrombolytic therapy.

Pin1 facilitates isoproterenol­induced cardiac fibrosis and collagen deposition by promoting oxidative stress and activating the MEK1/2­ERK1/2 signal transduction pathway in rats.

Peptidyl­prolyl cis/trans isomerase, NIMA-interacting 1 (Pin1) is a member of a large superfamily of phosphorylation­dependent peptidyl­prolyl cis/trans isomerases, which not only regulates multiple targets at various stages of cellular processes, but is also involved in the pathogenesis of several diseases, including microbial infection, cancer, asthma and Alzheimer's disease. However, the role of Pin1 in cardiac fibrosis remains to be fully elucidated. The present study investigated the potential mechanism of Pin1 in isoprenaline (ISO)­induced myocardial fibrosis in rats. The rats were randomly divided into three groups. Echocardiography was used to evaluate changes in the size, shape and function of the heart, and histological staining was performed to visualize inflammatory cell infiltration and fibrosis. Reverse transcription­quantitative polymerase chain reaction analysis, immunohistochemistry and Picrosirius red staining were used to differentiate collagen subtypes. Additionally, cardiac­specific phosphorylation of mitogen­activated protein kinase kinase 1/2 (MEK1/2) and extracellular­signal regulated protein kinase 1/2 (ERK1/2), and the activities of Pin1 and α­smooth muscle actin (α­SMA) and other oxidative stress parameters were estimated in the heart. The administration of ISO resulted in an increase in cardiac parameters and elevated the heart­to­body weight ratio. Histopathological examination of heart tissues revealed interstitial inflammatory cellular infiltrate and disorganized collagen fiber deposition. In addition, lipid peroxidation products and oxidative stress marker activity in plasma and tissues were significantly increased in the ISO­treated rats. Western blot analysis showed significantly elevated protein levels of phosphorylated Pin1, MEK1/2, ERK1/2 and α­SMA in remodeling hearts. Treatment with juglone following intraperitoneal injection of ISO significantly prevented inflammatory cell infiltration, improved cardiac function, and suppressed oxidative stresses and fibrotic alterations. In conclusion, the results of the present study suggested that the activation of Pin1 promoted cardiac extracellular matrix deposition and oxidative stress damage by regulating the phosphorylation of the MEK1/2­ERK1/2 signaling pathway and the expression of α­SMA. By contrast, the inhibition of Pin1 alleviated cardiac damage and fibrosis in the experimental models, suggesting that Pin1 contributed to the development of cardiac remodeling in ISO­administered rats, and that the inactivation of Pin1 may be a novel therapeutic candidate for the treatment of cardiovascular disease and heart failure.

A novel Norrie disease pseudoglioma gene mutation, c.-1_2delAAT, responsible for Norrie disease in a Chinese family.

AIM: To investigate the genetic findings and phenotypic characteristics of a Chinese family with Norrie disease (ND). METHODS: Molecular genetic analysis and clinical examinations were performed on a Chinese family with ND. Mutations in the Norrie disease pseudoglioma (NDP) gene were detected by direct sequencing. Haplotypes were constructed and compared with the phenotypes in the family. Evolutionary comparisons and mutant open reading frame (ORF) prediction were also undertaken. RESULTS: Two family members with ocular manifestations were diagnosed with ND. No signs of sensorineural hearing loss were observed in either patient, while one of them showed signs of mild mental retardation. A novel heterozygous mutation in the NDP gene, c.-1_2delAAT, was detected in both patients. The mutation and the mutation bearing haplotype co-segregated with the ND phenotype in males and was transmitted from their mothers and/or grandmothers (II:2). The male without ND did not harbor the mutation. The mutation occurred at the highly conserved nucleotides. ORF finder predicted that the mutation would lead to the production of a truncated protein that lacks the first 11 N-terminal amino acids. CONCLUSION: A novel mutation, c.-1_2delAAT in the NDP gene, was identified in a Chinese family with ND. This mutation caused ND without obvious sensorineural hearing loss. Mental disorder was found in one but not the other patients. The clinical heterogeneity in the family indicated that other genetic variants and epigenetic factors may also play a role in the disease presentation.

Hydrolysis of green tea residue protein using proteolytic enzyme derived from Aspergillus oryzae.

Free amino acids are important chemical components which impact the taste of green tea infusion. The hydrolysis of water-insoluble protein in the green tea residue helps to increase the contents of free amino acids components except theanine. Studies indicate that the hydrolysis of the tea protein could be restricted due to interaction of polyphenols with protein. The experiment indicates that the hydrolysis of tea protein by protease is the main trend when the polyphenols concentration is lower than 5 mg ml(-1), however, the proteins (including tea protein and protease) would interact with polyphenoles instead of hydrolysis when the concentration of polyphenols is higher than 5 mg ml(-1). The hydrolysis of tea protein is absolutely restrained when concentration comes to 10 mg ml(-1).

[Effects of PI3K/Akt/NF-κB signal pathway on FSH facilitation on cell proliferation and invasion by human epithelial ovarian cancer].

OBJECTIVE: To explore the effects of phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt)/nuclear factor-kappa B (NF-κB) signal pathway on the process of follicle-stimulating hormone (FSH) facilitating cell proliferation and invasion in human epithelial ovarian cancer. METHODS: Ovarian cancer cell lines SKOV3 and 3AO were cultured to exponential phase, then assigned to control group, FSH group, LY294002 group and FSH + LY294002 group, respectively. Cells were treated with different concentration of FSH and LY294002, respectively. The effects of FSH on cell proliferation were observed by methylthiazolyl tetrazolium (MTT). Morphological changes were observed by phase contrast microscope. The ability of cell invasion was investigated by transwell invasion assay. The expression of FSH receptor (FSHR), Akt1/2, phosphorylated-Akt (p-Akt) and NF-κB p65 protein were detected by western blot. RESULTS: (1) FSH could promote the proliferation of SKOV3 and 3AO cells. When the cells were treated with 40 U/L FSH for 48 hours (SKOV3) and 24 hours (3AO), compared with those in control groups, they reached the highest proliferation rate (P < 0.05), respectively. (2) The morphology of SKOV3 and 3AO cells in four groups:in control group, SKOV3 cells were short spindle and 3AO cells were long spindle, the nuclei of them were both roundness or oval, the cytoplasm were bright. In FSH group, the cells changed to slightly longer or polygonal, they were full in shape, meanwhile, the cell intensity were higher than control group. In LY294002 group, some cells changed from spindle to round, and began to shrink. The cell intensity diminished. The morphology of FSH + LY294002 group was similar with control group, but the cell intensity was lower than that in FSH group. (3) The number of SKOV3 cell that passed through the membrane in control group, FSH group, LY294002 group and FSH + LY294002 group was (26 ± 6), (118 ± 19), (18 ± 5) and (38 ± 7), respectively. The number of 3AO cell was (19 ± 4), (134 ± 20), (12 ± 3) and (58 ± 11), respectively. The results showed that the number of cells in FSH group was significantly higher than that in control group (P < 0.05), while the number of cell in FSH + LY294002 group was significantly fewer than that in FSH group (P < 0.05). (4) There was no significant difference in the expression of FSHR and Akt1/2 between FSH group and control group (P > 0.05), but FSH increased the expression of p-Akt and the ratio of NF-κB p65 in the nucleus versus cytoplasm in SKOV3 and 3AO cells, there were significant differences compared with control group (P < 0.05). LY294002 reversed the effects of FSH on increasing the expression of p-Akt and the ratio of NF-κB p65 in the nucleus versus cytoplasm, there were significant differences among LY294002 group, FSH + LY294002 group and FSH group (P < 0.05). CONCLUSION: The effects of FSH on proliferation and invasion of ovarian cancer cell lines SKOV3 and 3AO may be realized by regulating the activity of NF-κB in PI3K/Akt signal pathway.

Analysis of cream formation in green tea concentrates with different solid concentrations.

The formation of tea cream in the green tea concentrates of different solid concentrations (5, 10, 20, 30, 40, 50 and 60°Brix) was investigated. The results showed a good positive correlation (γ = 0.98, p ≤ 0.05) between the amount of tea cream and the solid concentrations from 5 to 40°Brix, while the amount of tea cream in the tea concentrates of 50 and 60°Brix decreased acutely. Total sugar, caffeine and catechins were found to be the main chemical components of tea cream in the green tea concentrate. The large decrease of the amount of tea cream in the tea concentrates of 50 and 60°Brix may be induced by a sharp increase of the viscosity of the tea concentrates, which helped to improve the stability of tea concentrate. It may be indicated that the stability of green tea concentrate enhanced when the concentration higher than 50°Brix, which helped to restrain the formation of tea cream.

Identification of PRRT2 as the causative gene of paroxysmal kinesigenic dyskinesias.

Paroxysmal kinesigenic dyskinesias is a paroxysmal movement disorder characterized by recurrent, brief attacks of abnormal involuntary movements induced by sudden voluntary movements. Although several loci, including the pericentromeric region of chromosome 16, have been linked to paroxysmal kinesigenic dyskinesias, the causative gene has not yet been identified. Here, we identified proline-rich transmembrane protein 2 (PRRT2) as a causative gene of paroxysmal kinesigenic dyskinesias by using a combination of exome sequencing and linkage analysis. Genetic linkage mapping with 11 markers that encompassed the pericentromeric of chromosome 16 was performed in 27 members of two families with autosomal dominant paroxysmal kinesigenic dyskinesias. Then, the whole-exome sequencing was performed in three patients from these two families. By combining the defined linkage region (16p12.1-q12.1) and the results of exome sequencing, we identified an insertion mutation c.649_650InsC (p.P217fsX7) in one family and a nonsense mutation c.487C>T (p.Q163X) in another family. To confirm our findings, we sequenced the exons and flanking introns of PRRT2 in another three families with paroxysmal kinesigenic dyskinesias. The c.649_650InsC (p.P217fsX7) mutation was identified in two of these families, whereas a missense mutation, c.796C>T (R266W), was identified in another family with paroxysmal kinesigenic dyskinesias. All of these mutations completely co-segregated with the phenotype in each family. None of these mutations was identified in 500 normal unaffected individuals of matched geographical ancestry. Thus, we have identified PRRT2 as the first causative gene of paroxysmal kinesigenic dyskinesias, warranting further investigations to understand the pathogenesis of this disorder.

[Association of the CYP19 gene polymorphism with genetic susceptibility to endometriosis].

OBJECTIVE: To investigate whether the 115T/C, 240A/G and 1531C/T polymorphisms of CYP19 gene are associated with the risk of moderate/severe endometriosis. METHODS: Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique was used to identify the CYP19 gene polymorphism in Chinese patients with endometriosis of III-IV stage (n= 102) and individuals without endometriosis (n= 100). RESULTS: The frequencies of CYP19 gene 115T/C, TT, TC and CC were 0.9118, 0.0882 and 0 in the endometriosis group, and 0.8800, 0.1100 and 0.0100 in the control group, respectively. The frequencies of 115T and C alleles in both groups were 0.9559 and 0.0441, and 0.9350 and 0.0650, respectively (P> 0.05). The frequencies of CYP19 gene 240AA, AG and GG were 0.2745, 0.4902 and 0.2353 in the endometriosis group, and 0.4500, 0.4100 and 0.1400 in the control group, respectively. The frequencies of 240A and G alleles in both groups were 0.5196 and 0.4804, and 0.6550 and 0.3450, respectively (P< 0.05). The frequencies of CYP19 gene 1531C/T, CC, CT and TT were 0.4118, 0.4706 and 0.1176 in the endometriosis group, and 0.3800, 0.4200 and 0.200 in the control group, respectively. The frequencies of 1531C and T alleles in both groups were 0.6471 and 0.3529, and 0.5900 and 0.4100, respectively (P> 0.05). CONCLUSION: CYP19 gene 240G/G polymorphism may contribute to the susceptibility of stages III and IV endometriosis but there was no association between CYP19 gene 115T/C and 1531C/T polymorphisms and stage III and IV endometriosis.

R25G mutation in exon 1 of LMNA gene is associated with dilated cardiomyopathy and limb-girdle muscular dystrophy 1B.

BACKGROUND: Mutations of the LMNA gene encoding lamin A and C are associated with dilated cardiomyopathy (DCM), conduction system defects and skeletal muscle dystrophy. Here we report a family with a mutation of the LMNA gene to identify the relationship between genotype and phenotype. METHODS: All 30 members of the family underwent clinical and genetic evaluation. A mutation analysis of the LMNA gene was performed. All of the 12 exons of LMNA gene were extended with polymerase chain reaction (PCR) and the PCR products were screened for gene mutation by direct sequencing. RESULTS: Ten members of the family had limb-girdle muscular dystrophy (LGMD) and 6 are still alive. Two patients suffered from DCM. Cardiac arrhythmias included atrioventricular block and atrial fibrillation; sudden death occurred in 2 patients. The pattern of inheritance was autosomal dominant. Mutation c.73C > G (R25G) in exon 1 encoding the globular domains was confirmed in all of the affected members, resulting in the conversion of arginine (Arg) to glycine (Gly). CONCLUSIONS: The mutation R25G in exon 1 of LMNA gene we reported here in a Chinese family had a phenotype of malignant arrhythmia and mild LGMD, suggesting that patients with familial DCM, conduction system defects and skeletal muscle dystrophy should be screened by genetic testing for the LMNA gene.

[Recent advances of genetic research on paroxysmal kinesigenic dyskinesias].

Paroxysmal kinesigenic choreoathetosis/dyskinesias (PKC/PKD) is one of the most common types of praoxysmal dyskinesia. It is characterized by recurrent episodic dystonia and/or choreoathetotic attacks triggered by sudden voluntary movement. Some patients have a history of febrile infantile convulsion. PKD commonly occurs sporadically or as an autosomal-dominant familial trait with variable penetrance. It has been linked to 16p12-q12 or 16q13-q22 loci in various families of different populations, which suggests a genetic heterogeneity. The exact etiology and pathogenesis of PKD await further elucidation, although ion channelopathy is suggested as a probable underlying etiology. Here, the recent advances of the genetic research on PKD will be reviewed.

[Clinical observation of microwave endometrial ablation for treatment of abnormal uterine bleeding].

OBJECTIVE: To evaluate the efficacy, indication, and complication of microwave endometrial ablation (MEA) in treating abnormal uterine bleeding (AUB). METHODS: One hundred and sixty-eight women with AUB due to benign causes received MEA treatment. Pre-operative endometrial thinning was carried out using uterine curettage. Then, the applicator radiating microwaves was moved by progressive withdrawal as well as "W" shape motion inside uterine cavity. All the patients were followed-up. The change of menstrual cycle, the amount of flow, dysmenorrhoea, anemia after treatment at 1, 3, 6, 12 and 24 months was recorded. RESULTS: The mean operating time was (286 +/- 75) seconds. Average follow-up time was (22 +/- 6, range 6-36) months. Of these patients, 156 women (92.9%) were premenopausal, 97 cases (62.2%) were amenorrhea, 56 cases (35.9%) were hypomenorrhoea or eumenorroea, and 3 cases (1.9%) had irregular bleeding. The overall satisfaction of this treatment reached 98.1% (153/156). The follow-up of 119 cases was up to 24 months after operation. The concentration of hemoglobin in 107 women with anemia increased significantly from (83 +/- 24) g/L to (117 +/- 18) g/L 3 months after operation (P < 0.01). Dysmenorrhoea was relieved in 74.5% (35/47) patients. No bleeding occurred in any one of 12 postmenopausal patients after MEA. There was no intraoperative complication in any case. The procedure was successful in all of 47 patients with severe medical disorders. After operation, 12 cases were complicated with endometritis, 2 with hematometra, and one case was performed with hysterectomy due to postablation tubal sterilization syndrome. CONCLUSIONS: MEA is a simple, safe and effective treatment of patients with abnormal uterine bleeding, especially suitable for those women associated with severe medical complications. Complete endometrial ablation is one of the most important determinants of treatment success. Stringent selection of patients may reduce the rate of complications.

[Study on gene mutations of alpha-thalassemia in the South of China].

There is a high prevalence of thalassemia in the South of China. To explore the genotype of alpha-thalassemia as well as the distribution of alpha globin gene mutation in the South of China, 356 patients with heterozygote alpha(+) thalassemia, heterozygote alpha(0) or homozygote alpha(+) thalassemia and 78 patients with HbH were analyzed. The gene diagnosis methods including Gap-PCR, nested-PCR, PCR-RE, PCR-SSCP, 4P-ASPCR and DNA sequence analysis were used. The results showed that among 356 patients, 295 patients with --SEA/alphaalpha (82.87%), 1 patient with alphaalpha/alpha-alpha(3.7) (0.28%), 3 patients with alphaalpha/alpha-alpha(4.2) (0.84%), 3 patients with alphaalpha/alpha(CS)alpha (0.84%), 1 patient with alphaalpha/alphaalpha(QS) (0.28%) and 2 patients with alphaalpha/alpha(Westmead) alpha (0.56%) were found. The homozygote with -alpha(4.2) or -alpha(3.7) was not found. In 78 patients with HbH, 29 patients with --SEA/alphaalpha(-3.7) (37.2%), 20 patients with --SEA/alphaalpha(-4.2) (25.6%), 19 patients with --SEA/alphaalpha(CS) (24.3%), 2 patients with --SEA/alphaalpha(QS) (2.6%) were detected, and other remaiming 8 patients were needed to be defined. Among the non-defined 8 patients, the synonymous mutation with C-->G transversion (GCC-GCG) at codon 65 in the exon 2 of alpha 2-globin gene was detected in 2 unrelated HbH patients came from Guangxi province. Whether it correlated with the phenotype of HbH disease or it is only a single nucleotide polymorphism site (SNPs), should be confirmed in the future. In addition, a set of gene diagnosis methods based on PCR to screen deletion and non-deletion genotypes of alpha-thalassemia in Chinese was improved. A new method, 4P-ASPCR, to detect Hb CS and Hb QS was also developed. The method was verified to be more accurate, time-saving and economic. In conclusion, the genotypes of alpha-thalassemia in Chinese are very complicated, the genotypes of alpha-thalassemia in Chinese need to be further studied, the results of this research probably have practical significance for the gene diagnosis or antenatal diagnosis of alpha-thalassemia in the South of China.

[Detection and analysis of the sub-types of -alpha3.7 in Chinese].

-alpha3.7 is a common deletional alpha-thalassemia-2 in China. According to different recombination sites,-alpha3.7 can be divided into -alpha3.7I,-alpha3.7IIand -alpha3.7III. The frequency and population distribution of these -alpha3.7 are quite different. In this study,we detected 56 patients among Chinese population of -alpha3.7 defect in alpha globin gene by PCR method, then the PCR product was digested by the restriction enzyme ApalI and BalI. The sub-typing result shows that in the 56 cases of -alpha3.7 defect,54 out of 56 is -alpha3.7I,2 out of 56 is -alpha3.7II and none of -alpha3.7III is detected. This result enriches the data about the alpha thalassemia genotypes of Chinese people.