[Implication of BRAF V600E and CTNNB1 gene mutations in the pathological classification of craniopharyngioma].

Objective: To investigate the clinicopathological significance of BRAF V600E and CTNNB1 gene mutations in adamantinomatous craniopharyngiomas (ACP) and papillary craniopharyngiomas (PCP). Methods: The retrospective study included a total of 67 craniopharyngiomas diagnosed from October 2009 to August 2018 at Xuanwu Hospital, Capital Medical University. The immunohistochemical staining for ß-catenin and BRAF V600E expression, Sanger sequencing of exon 3 of CTNNB1, BRAF mutation analysis by scorpions amplification refractory mutation system (ARMS) fluorescence quantitative PCR were performed. Univariate survival analysis was used to correlate with tumor recurrence. Results: Of the 67 patients, 53 were ACPs and 14 were PCPs. Four patients underwent multiple operations and one of them presented with malignant transformation into squamous cell carcinoma. Histologically, ACPs were characterized by whorl-like cell clusters, peripheral palisaded layer, stellate reticulum, finger-shaped protrusions, ghost cells and wet keratinous substances. While PCPs usually consisted of mature squamous epithelium associated with fibrovascular stroma resulting in papillary appearance. The nuclear immunopositivity for ß-catenin was observed in 73.6% (39/53) of ACPs, and it was absent in PCPs (0/14). The nuclear translocation of ß-catenin usually presented at whorl-like structures or around ghost cells. Of all the cases, mutations analysis in exon 3 of ß-catenin gene CTNNB1 were successful in 46 cases and 42.1% (16/38) of ACP showed CTNNB1 gene mutation, while none of the PCPs harbored CTNNB1 gene mutation (0/8). The cytoplasmic immunopositivity for BRAF V600E mutant protein was found in all PCPs (14/14) and negative in all ACPs (0/53). ARMS-PCR results showed that BRAF V600E mutations were observed in 13/14 of PCPs but not seen in ACPs (0/53). Follow-up data were available in 35 patients with duration of 2 to 120 months. Ten patients experienced recurrences after the first surgery. Upon univariate survival analysis, only subtotal excision was found to be associated with increased recurrence (P=0.032), while pathological type, postoperative radiotherapy and CTNNB1 gene mutation were not (P>0.05). Conclusions: There is significant difference in the expression of BRAF V600E and CTNNB1 genes between ACP and PCP, and their immunohistochemical and molecular detection therefore can be used in the diagnosis and differential diagnoses of craniopharyngiomas.

[Correlation between prognosis and laboratory parameters of complete deafness in patients with sudden deafness].

Objective: To study the correlation between the prognosis of sudden deafness and laboratory indicators, and to provide a theoretical basis for clinical evaluation of prognosis and selection of appropriate interventions.Method: A retrospective analysis of 123 cases of total deafness was conducted(all frequencies decreased with 250-8 000 Hz average hearing threshold ≥ 81 dB HL). All patients admitted to the hospital received laboratory test within 24 hours and underwent a unified treatment: alprostadil + mecobalamin + citicoline sodium phosphate + Ginkgo biloba extract + batroxobin + methylprednisolone (patients with hypertension or diabetes were given retro-auricular injection). After 2 weeks of comprehensive treatment, the pure tone audiometry was reviewed and the clinical efficacy was evaluated. According to the prognosis, the patients were divided into the healing group, the marked effective group, the effective group and the ineffective group. The laboratory indicators included: white blood cell count, neutrophil percentage, neutrophil lymphocyte ratio(NHR), erythrocyte sedimentation rate, blood lipid, red blood cell count, hemoglobin(Hb), plasma fibrinogen content(FIB). Result: There were no significant differences in white blood cell count, neutrophil percentage, NHR, erythrocyte sedimentation rate, blood lipid index and red blood cell count between different prognosis groups(all P>0.05). The differences of FIB and Hb were statistically significant(P<0.05). The overall prognosis was negatively correlated with FIB and positively correlated with Hb. Patients with vertigo had a worse prognosis and a higher plasma FIB level than those without vertigo(P<0.05), but there was no significant difference of Hb between patients with and without vertigo(P>0.05). Conclusion: FIB and Hb can be used as correlative indexes to evaluate the prognosis of patients with sudden deafness, and the effects of vertigo and other associated symptoms on the prognosis should be comprehensively analyzed. .

Drug-resistance reversal in colorectal cancer cells by destruction of flotillins, the key lipid rafts proteins.

Multi-drug resistance (MDR) of tumor cells attenuates the efficacy of anticancer drugs and has become the main reason for chemotherapy failure. It is indispensable to establish an effective way to reverse multi-drug resistance. Our previous work has shown that down-regulation of the ERK/MAPK signaling pathway activity can reverse the drug-resistance of resistant cells. Further-more, the effect of signal transduction is strongly associated with lipid rafts. The drug-resistance is reversed successfully after lipid rafts are destroyed by heptakis(2, 6-di-O-methyl)-ß-cyclodextrin (MßCD). However, the reversal of the drug-resistance is not associated with down-regulation of the expression of ERK1/2. Cell membrane permeability may increase when lipid rafts are destroyed by MßCD, causing the reversal of drug-resistance due to an increase in accumulation of the drugs in the cytoplasm. To minimize the influence of MßCD on the cell membrane structure, we selected flotillin, a marker protein of lipid rafts, as the target molecule, to further investigate the mechanism of changes in drug resistance after destruction of the lipid rafts. The effect of flotillin on the reversal of the drug resistance was examined using an RNA interference (RNAi) in a retrovirus system in human drug-resistant strains of colorectal cancer cell line HCT-15. The results demonstrate that flotillin-1 downregulation by RNAi (Flot1-RNAi) reduced the drug resistance, caused cell cycle arrest and decreased the expression of ERK1/2; however, apoptosis was not significantly affected. Knockdown of flotillin-2 by RNAi (Flot2-RNAi) had effects similar to those of Flot1-RNAi except that the effects on expression of ERK1/2 and apoptosis were different. Screening of multiple pathways indicated that the PI3K/Akt signaling pathway was closely related. This experiment demonstrates the association between PI3K and drug resistance through the activation of PI3K and suggests that PI3K may play a key role during the development of resistance in CRC. The results reveal that the levels of IRS-1 and PI3K proteins in the Flot1-RNAi and Flot2-RNAi groups were significantly down-regulated. Knockdown of flotillins by RNAi reduced the resistance of HCT-15/ADM cells; the results investigations of the Akt pathway indicate a decrease in resistance after lipid raft destruction. These data confirm that knockdown of flotillin reduces the resistance of HCT-15/ADM cells, and the mechanism may be relevant to the PI3K/Akt pathway. Additionally, flotillin may be used as a potential target for chemotherapy in the treatment of colorectal cancer.

A genome-wide association study reveals candidate genes for the supernumerary nipple phenotype in sheep (Ovis aries).

Genome-wide association studies (GWASs) have been widely applied in livestock to identify genes associated with traits of economic interest. Here, we conducted the first GWAS of the supernumerary nipple phenotype in Wadi sheep, a native Chinese sheep breed, based on Ovine Infinium HD SNP BeadChip genotypes in a total of 144 ewes (75 cases with four teats, including two normal and two supernumerary teats, and 69 control cases with two teats). We detected 63 significant SNPs at the chromosome-wise threshold. Additionally, one candidate region (chr1: 170.723-170.734 Mb) was identified by haplotype-based association tests, with one SNP (rs413490006) surrounding functional genes BBX and CD47 on chromosome 1 being commonly identified as significant by the two mentioned analyses. Moreover, Gene Ontology enrichment for the significant SNPs identified by the GWAS analysis was functionally clustered into the categories of receptor activity and synaptic membrane. In addition, pathway mapping revealed four promising pathways (Wnt, oxytocin, MAPK and axon guidance) involved in the development of the supernumerary nipple phenotype. Our results provide novel and important insights into the genetic mechanisms underlying the phenotype of supernumerary nipples in mammals, including humans. These findings may be useful for future breeding and genetics in sheep and other livestock.

[Progress of sino-nasal IgG4-related disease].

IgG4-related disease is a newly recognized systemic fibro inflammatory disorder that affects the sino-nasal region. It is a rare and emerging entity that can present with bony and soft-tissue invasion,the final diagnosis of this disease mainly depends on pathological examination and majority of patients receiving corticosteroids responded very well to treatment. Thus,Our goal was to highlight the sino-nasal presentation of this unique disease and to review previously reported cases from 2010 to 2016.We hope that clinical physicians to enhance understanding of the disease in order to ensure early diagnosis and early intervention to prevent serious injury and fibrosis of organs.

Serum microRNA-210 as a potential noninvasive biomarker for the diagnosis and prognosis of glioma.

BACKGROUND: MicroRNA-210 (miR-210) is an oncogenic miRNA previously associated with prognosis in human gliomas, an incurable tumour type of the central nervous system. Here miR-210 was investigated as a potential serum biomarker in the diagnosis and prognosis of glioma. METHODS: Serum was immediately prepared from blood samples collected from patients with glioma grades I-IV at primary diagnosis (n=136) and healthy controls (n=50) from February 2007 to March 2014 in the Department of Neurosurgery of the First Affiliated Hospital of Wannan Medical College (Wuhu, China). Total RNA was isolated from serum. cDNA was synthesised with primers specific for miR-210 and miR-16-1 (internal control), and quantitative real-time RT-PCR was performed. Results were statistically analysed to determine the role of miR-210 in the diagnosis and prognosis of human glioma patients. RESULTS: An approximately seven-fold increase in miR-210 expression was detected in serum samples from glioblastoma patients relative to healthy controls. A threshold expression value (2.259) was chosen from receiver operator characteristic curves (ROC), and the low and high miR-210 expression groups were analysed by multivariate Cox proportional hazard regression and Kaplan-Meier analyses. Results revealed an association of high serum miR-210 expression with tumour grade and poor patient outcome (P-values <0.001). CONCLUSIONS: Serum miR-210 is a promising diagnostic and prognostic biomarker that can be detected in the peripheral blood of patients with glioma.

Pathways involved in the spread of buccal carcinoma on contrast-enhanced multislice CT.

OBJECTIVES: To explore how buccal carcinoma spread, using contrast-enhanced multislice CT (CEMSCT). METHODS: We retrospectively analysed the extent of lesions in 56 patients with primary buccal squamous cell carcinoma (SCCA). Abnormal manifestations on CEMSCT at oral subsites and involved adjacent structures were documented and evaluated, which were compared with the results of surgery and histopathology. RESULTS: Infiltration and spread to oral subsites and/or adjacent structures was confirmed in 33 patients (58.9%). The opening of the Stensen duct was the most commonly invaded oral subsite (72.7%); other sites included the gingivobuccal sulcus (60.6%), pterygomandibular raphe (54.5%), gingiva (24.2%), retromolar trigone (24.2%), orbicularis oris (18.2%) and the floor of mouth (15.2%). Of the involved adjacent structures, the buccal space was the most common site of spread (69.7%), followed by the masticatory muscles and spaces (57.6%), bone (54.5%), skin and subcutaneous fat (39.4%), pharynx (30.3%), investing fascia (15.2%) and the base of the skull (6.1%). CEMSCT manifestations of the involvement in buccal SCCAs had correlations with pathological findings (p < 0.05). The sensitivities, specificities and accuracies of two radiologists' evaluation on buccal carcinoma involvement were 50.00%, 23.21% and 73.21%; and 51.79%, 32.14% and 83.93%, respectively. CONCLUSIONS: Buccal SCCAs could superficially and deeply spread to multiple oral subsites and/or adjacent structures. CEMSCT could delineate their spread pathways and extents.

Radiation hybrid QTL mapping of Tdes2 involved in the first meiotic division of wheat.

Since the dawn of wheat cytogenetics, chromosome 3B has been known to harbor a gene(s) that, when removed, causes chromosome desynapsis and gametic sterility. The lack of natural genetic diversity for this gene(s) has prevented any attempt to fine map and further characterize it. Here, gamma radiation treatment was used to create artificial diversity for this locus. A total of 696 radiation hybrid lines were genotyped with a custom mini array of 140 DArT markers, selected to evenly span the whole 3B chromosome. The resulting map spanned 2,852 centi Ray with a calculated resolution of 0.384 Mb. Phenotyping for the occurrence of meiotic desynapsis was conducted by measuring the level of gametic sterility as seeds produced per spikelet and pollen viability at booting. Composite interval mapping revealed a single QTL with LOD of 16.2 and r (2) of 25.6 % between markers wmc326 and wPt-8983 on the long arm of chromosome 3B. By independent analysis, the location of the QTL was confirmed to be within the deletion bin 3BL7-0.63-1.00 and to correspond to a single gene located ~1.4 Mb away from wPt-8983. The meiotic behavior of lines lacking this gene was characterized cytogenetically to reveal striking similarities with mutants for the dy locus, located on the syntenic chromosome 3 of maize. This represents the first example to date of employing radiation hybrids for QTL analysis. The success achieved by this approach provides an ideal starting point for the final cloning of this interesting gene involved in meiosis of cereals.

A Novel Retrotransposon Inserted in the Dominant Vrn-B1 Allele Confers Spring Growth Habit in Tetraploid Wheat (Triticum turgidum L.).

Vernalization genes determine winter/spring growth habit in temperate cereals and play important roles in plant development and environmental adaptation. In wheat (Triticum L. sp.), it was previously shown that allelic variation in the vernalization gene VRN1 was due to deletions or insertions either in the promoter or in the first intron. Here, we report a novel Vrn-B1 allele that has a retrotransposon in its promoter conferring spring growth habit. The VRN-B1 gene was mapped in a doubled haploid population that segregated for winter-spring growth habit but was derived from two spring tetraploid wheat genotypes, the durum wheat (T. turgidum subsp. durum) variety 'Lebsock' and T. turgidum subsp. carthlicum accession PI 94749. Genetic analysis revealed that Lebsock carried the dominant Vrn-A1 and recessive vrn-B1 alleles, whereas PI 94749 had the recessive vrn-A1 and dominant Vrn-B1 alleles. The Vrn-A1 allele in Lebsock was the same as the Vrn-A1c allele previously reported in hexaploid wheat. No differences existed between the vrn-B1 and Vrn-B1 alleles, except that a 5463-bp insertion was detected in the 5'-UTR region of the Vrn-B1 allele. This insertion was a novel retrotransposon (designated as retrotrans_VRN), which was flanked by a 5-bp target site duplication and contained primer binding site and polypurine tract motifs, a 325-bp long terminal repeat, and an open reading frame encoding 1231 amino acids. The insertion of retrotrans_VRN resulted in expression of Vrn-B1 without vernalization. Retrotrans_VRN is prevalent among T. turgidum subsp. carthlicum accessions, less prevalent among T. turgidum subsp. dicoccum accessions, and rarely found in other tetraploid wheat subspecies.

Identification of novel tan spot resistance loci beyond the known host-selective toxin insensitivity genes in wheat.

Tan spot, caused by Pyrenophora tritici-repentis, is a destructive foliar disease of wheat causing significant yield reduction in major wheat growing areas throughout the world. The objective of this study was to identify quantitative trait loci (QTL) conferring resistance to tan spot in the synthetic hexaploid wheat (SHW) line TA4152-60. A doubled haploid (DH) mapping population derived from TA4152-60 x ND495 was inoculated with conidia produced by isolates of each of four virulent races of P. tritici-repentis found in North America. QTL analysis revealed a total of five genomic regions significantly associated with tan spot resistance, all of which were contributed by the SHW line. Among them, two novel QTLs located on chromosome arms 2AS and 5BL conferred resistance to all isolates tested. Another novel QTL on chromosome arm 5AL conferred resistance to isolates of races 1, 2 and 5, and a QTL specific to a race 3 isolate was detected on chromosome arm 4AL. None of these QTLs corresponded to known host selective toxin (HST) insensitivity loci, but a second QTL on chromosome arm 5BL conferred resistance to the Ptr ToxA producing isolates of races 1 and 2 and corresponded to the Tsn1 (Ptr ToxA sensitivity) locus. This indicates that the wheat-P. tritici-repentis pathosystem is much more complex than previously thought and that selecting for toxin insensitivity alone will not necessarily lead to tan spot resistance. The markers associated with the QTLs identified in this work will be useful for deploying the SHW line as a tan spot resistance source in wheat breeding.

Molecular mapping of hybrid necrosis genes Ne1 and Ne2 in hexaploid wheat using microsatellite markers.

Hybrid necrosis is the gradual premature death of leaves or plants in certain F1 hybrids of wheat (Triticum aestivum L.), and it is caused by the interaction of two dominant complementary genes Ne1 and Ne2 located on chromosome arms 5BL and 2BS, respectively. To date, molecular markers linked to these genes have not been identified and linkage relationships of the two genes with other important genes in wheat have not been established. We observed that the F1 hybrids from the crosses between the bread wheat variety 'Alsen' and four synthetic hexaploid wheat (SHW) lines (TA4152-19, TA4152-37, TA4152-44, and TA4152-60) developed at the International Maize and Wheat Improvement Center (CIMMYT) exhibited hybrid necrosis. This study was conducted to determine the genotypes of TA4152-60 and Alsen at the Ne1 and Ne2 loci, and to map the genes using microsatellite markers in backcross populations. Genetic analysis indicated that Alsen has the genotype ne1ne1Ne2Ne2 whereas the SHW lines have Ne1Ne1ne2ne2. The microsatellite marker Xbarc74 was linked to Ne1 at a genetic distance of 2.0 cM on chromosome arm 5BL, and Xbarc55 was 3.2 cM from Ne2 on 2BS. Comparison of the genetic maps with the chromosome deletion-based physical maps indicated that Ne1 lies in the proximal half of 5BL, whereas Ne2 is in the distal half of 2BS. Genetic linkage analysis showed that Ne1 was about 35 cM proximal to Tsn1, a locus conferring sensitivity to the host selective toxin Ptr ToxA produced by the tan spot fungus. The closely linked microsatellite markers identified in this study can be used to genotype parental lines for Ne1 and Ne2 or to eliminate the two hybrid necrosis genes using marker-assisted selection.

Heroin abuse and nitric oxide, oxidation, peroxidation, lipoperoxidation.

To further reveal the risks of heroin abuse to human body, and to determine the injuries of oxidation, peroxidation and lipoperoxidation induced by nitric oxide and other free radicals to heroin abusers, we determined and compared plasma values of lipoperoxides (LPO), nitric oxide (NO), vitamin C (VC), vitamin E (VE), beta-carotene (beta-CAR) and erythrocyte values of LPO, superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH-Px) in 114 heroin abusers and 100 healthy volunteers. Using linear regression and correlation as well as stepwise regression and correlation, we also analyzed the effect of the abusing duration, and daily abusing quantity on the above-mentioned biochemical parameters in the heroin abusers. The results showed that, compared with the healthy volunteer groups, the average plasma values of LPO, and NO, and the average erythrocyte value of LPO in the heroin abuser group were significantly increased (P < 0.0001), and the average plasma values of VC, VE, and beta-CAR and the average erythrocyte values of SOD, CAT, and GSH-Px were significantly decreased (P < 0.0001). Analysis of linear regression and correlation showed that with prolonged heroin abusing and with increased daily quantity in the heroin abusers, the plasma values of LPO, and NO, and the erythrocyte value of LPO were gradually increased (P < 0.001), whereas the plasma values of VC, VE, and beta-CAR and the erythrocyte values of SOD, CAT, and GSH-Px were gradually decreased (P < 0.001). Analysis of stepwise regression and correlation indicated that the plasma values of NO, VC and VE were closely correlated with the abusing duration and daily abusing quantity. These results indicate that the balance between oxidation and antioxidation in the heroin abusers was seriously disturbed, and the injuries induced by nitric oxide and other free radicals, through oxidation, peroxidation and lipoperoxidation to the bodies of heroin abusers exacerbated. It is therefore necessary that in abstaining from heroin dependence, the heroin abusers should acquire sufficient quantities of antioxidants such as VC, VE and beta-CAR.