Floral nectar reabsorption and a sugar concentration gradient in two long-spurred Habenaria species (Orchidaceae).

BACKGROUND: Floral nectar is the most common reward flowers offered to pollinators. The quality and quantity of nectar produced by a plant species provide a key to understanding its interactions with pollinators and predicting rates of reproductive success. However, nectar secretion is a dynamic process with a production period accompanied or followed by reabsorption and reabsorption remains an understudied topic. In this study, we compared nectar volume and sugar concentration in the flowers of two long-spurred orchid species, Habenaria limprichtii and H. davidii (Orchidaceae). We also compared sugar concentration gradients within their spurs and rates of reabsorption of water and sugars. RESULTS: Both species produced diluted nectar with sugar concentrations from 17 to 24%. Analyses of nectar production dynamics showed that as flowers of both species wilted almost all sugar was reabsorbed while the original water was retained in their spurs. We established a nectar sugar concentration gradient for both species, with differences in sugar concentrations at their spur's terminus and at their spur's entrance (sinus). Sugar concentration gradient levels were 1.1% in H. limprichtii and 2.8% in H. davidii, both decreasing as flowers aged. CONCLUSION: We provided evidence for the reabsorption of sugars but not water occurred in wilted flowers of both Habenaria species. Their sugar concentration gradients vanished as flowers aged suggesting a slow process of sugar diffusion from the nectary at the spur's terminus where the nectar gland is located. The processes of nectar secretion/reabsorption in conjunction with the dilution and hydration of sugar rewards for moth pollinators warrant further study.

Z16b, a natural compound from Ganoderma cochlear is a novel RyR2 stabilizer preventing catecholaminergic polymorphic ventricular tachycardia.

Catecholaminergic polymorphic ventricular tachycardia (CPVT) is an inherited, lethal ventricular arrhythmia triggered by catecholamines. Mutations in genes that encode cardiac ryanodine receptor (RyR2) and proteins that regulate RyR2 activity cause enhanced diastolic Ca2+ release (leak) through the RyR2 channels, resulting in CPVT. Current therapies for CPVT are limited. We found that Z16b, a meroterpenoid isolated from Ganoderma cochlear, inhibited Ca2+ spark frequency (CaSF) in R2474S/ + cardiomyocytes in a dose-dependent manner, with an IC50 of 3.2 µM. Z16b also dose-dependently suppressed abnormal post-pacing Ca2+ release events. Intraperitoneal injection (i.p.) of epinephrine and caffeine stimulated sustained ventricular tachycardia in all R2474S/+ mice, while pretreatment with Z16b (0.5 mg/kg, i.p.) prevented ventricular arrhythmia in 9 of 10 mice, and Z16b administration immediately after the onset of VT abolished sVT in 9 of 12 mice. Of translational significance, Z16b significantly inhibited CaSF and abnormal Ca2+ release events in human CPVT iPS-CMs. Mechanistically, Z16b interacts with RyR2, enhancing the "zipping" state of the N-terminal and central domains of RyR2. A molecular docking simulation and point mutation and pulldown assays identified Z16b forms hydrogen bonds with Arg626, His1670, and Gln2126 in RyR2 as a triangle shape that anchors the NTD and CD interaction and thus stabilizes RyR2 in a tight "zipping" conformation. Our findings support that Z16b is a novel RyR2 stabilizer that can prevent CPVT. It may also serve as a lead compound with a new scaffold for the design of safer and more efficient drugs for treating CPVT.

When is the circumferential stabilization necessary for subaxial cervical fracture dislocations? The posterior ligament-bone injury classification and severity score: a novel treatment algorithm.

OBJECTIVE: To propose a novel classification and scoring system called the posterior ligament-bone injury classification and severity score (PLICS) that offers a quantitative score to guide the need for posterior stabilization in addition to anterior reconstruction for subaxial cervical fracture dislocations (SCFDs). METHODS: A total of 456 patients with SCFDs were prospectively included. Patients with PLICS ≥ 7 together with extremely unstable lateral mass fracture (EULMF) were classified as high-risk group, and the other patients were classified as low-risk group. For patients in the low-risk group, anterior-only reconstruction was performed; for patients in the high-risk group, additional posterior lateral mass fixation and fusion was performed after anterior reconstruction. Clinical outcome evaluation included using the visual analogue score (VAS), the Neck Disability Index (NDI), and the American Spinal Injury Association (ASIA) impairment scale. The change in the local sagittal alignment kyphosis Cobb angle was also recorded. RESULTS: A total of 370 patients (81.1%) completed the minimal 12-month follow-ups, including 321 patients of low-risk group and 49 patients of high-risk group. Compared with the average VAS score preoperatively, the score at 12-month follow-up was significantly improved (from 6.1 + 0.3 to 1.1 + 0.2 in the low-risk group, P < 0.001; from 6.4 + 0.2 to 1.4 + 0.2 in the high-risk group, P < 0.001). The average NDI score at the 12-month follow-up was statistically low in the low-risk group (8.8 + 2.5 vs 13.8 + 3.4, P = 0.034). At least more than one grade improvement in the ASIA scale was observed in 80.5% of all patients. The local kyphosis Cobb angle at the injured segment averaged improved in both groups. CONCLUSION: A PLICS score ≥ 7 together with EULMF can be the threshold for posterior stabilization in addition to anterior reconstruction for the patients with SCFDs.

Clonal characteristics of paired infiltrating and circulating B lymphocyte repertoire in patients with primary biliary cholangitis.

BACKGROUND: PBC is a prototypical autoimmune liver disease characterized by portal lymphoplasmacyte infiltration. ALD is a prototypical environment-driven disease, featured by mild lymphocyte infiltration. We hypothesize that B cells are more involved in the pathogenesis of PBC. By analysing the infiltrating B cell repertoire, we aimed to unveil greater oligoclonal expansion and active clonal exchange between liver and periphery in PBC than in ALD patients. METHODS: Using NGS of Ig H chain genes, we analysed the liver-infiltrating and paired peripheral B lymphocyte repertoire from nine PBC and four ALD patients. RESULTS: In the liver of PBC and ALD patients, (i) roughly 10% of the B lymphocytes were clonally related and highly expressed, and there were also lineages that underwent extensive clonal expansion; (ii) there was different use of IGHV/IGHJ segments between PBC and ALD, suggesting distinct Ag exposure backgrounds, but this did not lead to a significant difference in their clonal expansion level. Analysis of data sets from paired samples further revealed, (iii) direct clonal exchange and evolutionally related B cell clones between the infiltrating and peripheral repertoire; (iv) the seeding of the infiltrating clones to periphery, and peripheral ones to the liver, for further extensive evolution. CONCLUSIONS: The oligoclonally expanded nature of the infiltrating B cell repertoire implies B cell immunity is involved in the pathogenesis of both diseases. The observed clonal exchange might provide an approach to identify and monitor the infiltrating B cells through the periphery.

Clonal Characteristics of Circulating B Lymphocyte Repertoire in Primary Biliary Cholangitis.

Primary biliary cholangitis (PBC) is an autoimmune liver disease characterized by elevated serum anti-mitochondrial Ab and lymphocyte-mediated bile duct damage. This study was designed to reveal the clonal characteristics of B lymphocyte repertoire in patients with PBC to facilitate better understanding of its pathogenesis and better management of these patients. Using high-throughput sequencing of Ig genes, we analyzed the repertoire of circulating B lymphocytes in 43 patients with PBC, and 34 age- and gender-matched healthy controls. Compared with healthy controls, PBC patients showed 1) a gain of 14 new clones and a loss of 8 clones; 2) a significant clonal expansion and increased relative IgM abundance, which corresponded with the elevated serum IgM level; 3) a significant reduction of clonal diversity and somatic hypermutations in class-switched sequences, which suggested a general immunocompromised status; 4) the reduction of clonal diversity and enhancement of clonal expansion were more obvious at the cirrhotic stage; and 5) treatment with ursodeoxycholic acid could increase the clonal diversity and reduce clonal expansion of the IgM repertoire, with no obvious effect on the somatic hypermutation level. Our data suggest that PBC is a complex autoimmune disease process with evidence of B lymphocyte clonal gains and losses, Ag-dependent ogligoclonal expansion, and a generally compromised immune reserve. This new insight into the pathogenesis of PBC opens up the prospect of studying disease-relevant B cells to better diagnose and treat this devastating disease.

Comparison of Percutaneous Kyphoplasty and Bone Cement-Augmented Short-Segment Pedicle Screw Fixation for Management of Kümmell Disease.

BACKGROUND The purpose of this study was to compare the efficacy of percutaneous kyphoplasty (PKP) and bone cement-augmented short segmental fixation (BCA+SSF) for treating Kümmell disease. MATERIAL AND METHODS Between June 2013 and December 2015, 60 patients were treated with PKP or BCA+SSF. All patients were followed up for 12-36 months. We retrospectively reviewed outcomes, including Oswestry Disability Index (ODI), visual analogue scale (VAS), and kyphotic Cobb angle. RESULTS VAS, ODI, and Cobb angle, measured postoperatively and at the final follow-up, were lower than those measured preoperatively in both groups (P<0.05). VAS, ODI, and Cobb angle measured postoperatively demonstrated no significant differences when compared with those measured at the final follow-up in the PKP group (P>0.05). In the BCA+SSF group, VAS and ODI at the final follow-up were lower than those measured postoperatively (P<0.05), but no significant difference was found in the Cobb angle (P>0.05). The PKP group had better VAS and ODI than the BCA+SSF group, postoperatively (P<0.05). No significant difference was found in VAS and ODI at the final follow-up (P>0.05) or the Cobb angle measured postoperatively and at the final follow-up (P>0.05) between the 2 groups. Operative time, blood loss, and hospital stay in the PKP group were lower than those in the BCA+SSF group (P<0.05). No significant difference was found in complications (P>0.05). CONCLUSIONS PKP patients had better early clinical outcomes, shorter operation times and hospital admission times, and decreased blood loss, but had similar complications, radiographic results, and long-term clinical outcomes compared with BCA+SSF patients.

Bone Cement-Augmented Short-Segment Pedicle Screw Fixation for Kümmell Disease with Spinal Canal Stenosis.

BACKGROUND This study aimed to explore the feasibility and efficacy of bone cement-augmented short-segmental pedicle screw fixation in treating Kümmell disease. MATERIAL AND METHODS From June 2012 to June 2015, 18 patients with Kümmell disease with spinal canal stenosis were enrolled in this study. Each patient was treated with bone cement-augmented short-segment fixation and posterolateral bone grafting, and posterior decompression was performed when needed. All patients were followed up for 12-36 months. We retrospectively reviewed outcomes, including the Oswestry disability index (ODI), visual analog scale (VAS) score, anterior and posterior heights of fractured vertebrae, kyphotic Cobb angle, and neurological function by Frankel classification. RESULTS The VAS grades, ODI scores, anterior heights of affected vertebrae, and kyphotic Cobb angles showed statistically significant differences between pre- and postoperative and between preoperative and final follow-up values (P<0.05), whereas the differences between postoperative and final follow-up values were not statistically significant (P>0.05). The differences between posterior vertebral heights at each time point were not statistically significant (P>0.05). Improved neurological function was observed in 12 cases at final follow-up. Three cases had complications, including asymptomatic cement leakage in 2 patients and delayed wound infection in 1 patient. CONCLUSIONS Bone cement-augmented short-segment pedicle screw fixation is safe and effective for treating Kümmell disease, and can achieve satisfactory correction of kyphosis and vertebral height, with pain relief and improvement in neurological function, with few complications.

The diagnostic value of confocal laser endomicroscopy for gastric cancer and precancerous lesions among Asian population: a system review and meta-analysis.

OBJECTIVE: The objective of this study is to evaluate the diagnostic value of confocal laser endomicroscopy (CLE) in detection of gastric cancer (GC), gastric intraepithelial metaplasia (GIM), and gastric intraepithelial neoplasia (GIN) lesions. METHOD: PubMed, the Cochrane Library, and Wangfang databases were searched to include eligible articles about CLE in detection of gastric lesions. After study selection, quality assessment and data extraction conducted by two reviewers independently, meta-analysis was performed by Meta-Disc 1.4. The pooled sensitivity and specificity was calculated, receiver operating characteristic (ROC) curve was constructed, and the area under ROC curve (AUC) was calculated. RESULTS: Twenty-three studies evaluating the diagnostic value of CLE were included. For the diagnosis of GC lesions, the pooled sensitivity, specificity, and AUC were 91% (88-94%), 99% (99-99%), and 0.9513, respectively. For the diagnosis of lesions, the pooled sensitivity, specificity, and AUC were 92% (90-94%), 97% (96-98%), and 0.9774, respectively. For the diagnosis of GIN lesions, the pooled sensitivity, specificity and AUC were 81% (75-85%), 98% (97-98%), and 0.9204, respectively. CONCLUSIONS: CLE can provide an accurate diagnosis with high sensitivity and specificity for GC, GIM, and GIN lesions. The results should be confirmed by well-designed, multi-centered, randomized controlled, and double blinded trials with large samples.

Endoscopic treatments for rectal neuroendocrine tumors smaller than 16 mm: a meta-analysis.

OBJECTIVE: Endoscopic mucosal resection (EMR), including conventional EMR (c-EMR) and modified EMR (m-EMR), was applied to remove small rectal neuroendocrine tumors (NETs). We aim to evaluate treatment outcomes of endoscopic submucosal dissection (ESD), m-EMR and c-EMR for rectal NETs <16 mm. METHODS: The PubMed, Cochrane Library and Elsevier Science Direct were searched to identify eligible articles. After quality assessment and data extraction, meta-analysis was performed. The main outcomes were complete resection rate, overall complication rate, procedure time and local recurrence rate. RESULTS: Compared with c-EMR, ESD could achieve higher complete resection rate (OR = 4.38, 95%CI: 2.43-7.91, p < 0.00001) without increasing overall complication rates (OR = 2.21, 95%CI: 0.56-8.70, p = 0.25). However, ESD was more time-consuming than c-EMR (MD = 6.72, 95%CI: 5.84-7.60, p < 0.00001). Compared with m-EMR, ESD did not differ from m-EMR in complete resection and overall complication rates (OR = 0.80, 95%CI: 0.51-1.27, p = 0.34; OR = 1.91, 95%CI: 0.75-4.86, p = 0.18, respectively). However, ESD was more time-consuming than m-EMR (MD = 12.21, 95%CI: 7.78-16.64, p < 0.00001). Compared with c-EMR, m-EMR could achieve higher complete resection rate (OR = 4.23, 95%CI: 2.39-7.50, p < 0.00001) without increasing overall complication rate (OR = 1.07, 95%CI: 0.35-3.32, p = 0.90). Moreover, m-EMR was not time-consuming than c-EMR (MD = 2.01, 95%CI: -0.37-4.40, p= 0.10). The local recurrence rate was 0.84% (9/1067) during follow-up. CONCLUSIONS: Both ESD and m-EMR have great advantages over c-EMR in complete resection rate without increasing safety concern while m-EMR shares similar outcomes with ESD for rectal NETs <16 mm. The results should be confirmed by well-designed, multicenter, randomized controlled trials with large samples and long-term follow-ups from more countries.

The impact of posterior temporary internal distraction on stepwise corrective surgery for extremely severe and rigid scoliosis greater than 130°.

AIM: To investigate the efficacy and safety of temporary internal distraction corrective surgery for extremely severe scoliosis. METHODS: Eleven scoliosis patients (3 males and 8 females) with curves ≥130° (mean 148.8°; range 130°-157°) who underwent a two-stage surgery, including a posterior temporary internal distraction correction and definitive posterior spinal correction with posterior pedicle screw instrumentation from 2008 to 2011 were retrospectively reviewed. Minimum follow-up was 2-years (mean 41.8 months; range 27.0-63.0 months). The analysis focused on the impact of temporary internal distraction on curve correction, pulmonary function tests (PFTs), complications and surgical outcomes. Neurosurveillance of sensory (somatosensory-evoked potentials) and motor (motor-evoked potentials) potential was performed in all cases. Posterior instrumentation was used in all patients. RESULTS: After the use of internal distraction, the preoperative major curve (mean 148.8°; range 130°-157°) was corrected to a mean of 79° (range 63°-87°), the T5-T12 kyphosis Cobb angle (mean 79°; range 30°-97°) was corrected to a mean of 59° (range 20°-75°), the coronal imbalance (mean 0.8 cm; range -3.6 to 2.8 cm) was improved to a mean of 0.6 cm (range -1.5 to 2.0 cm), the forced vital capacity percentage (FVC%) was improved from 59.3 ± 11.6 to 68.7 ± 13.7, and the forced expiratory volume in 1 s (FEV1%) was improved from 61.4 ± 13.6 to 71.3 ± 9.3. The average increase in body height was 6.7 cm, and the dorsum razor was corrected to 3-5 cm. During definitive surgery, the final major curves were corrected to a mean of 55° (range 32°-72°), the T5-T12 kyphosis Cobb was corrected to 35° (range 15°-68°), the coronal imbalance was improved to 0.5 cm (range -1.2 to 1.8 cm), the FVC% was improved to 71.2 ± 8.3, the FEV1% was improved to 76.3 ± 16.7, the increase in body height was 3.1 cm, and the dorsum razor was corrected to 1-3 cm. The mean interval time between the two surgeries was 3.5 months. None of the patients exhibited postoperative neurologic deficits or infections. No instrument complications were found during the final follow-up. CONCLUSIONS: Temporary internal distraction in a two-stage corrective surgery provided patients who had extremely severe and rigid scoliosis, an effective and safe solution for scoliosis without significant complications.

The beneficial effect of Batroxobin on blood loss reduction in spinal fusion surgery: a prospective, randomized, double-blind, placebo-controlled study.

OBJECTIVE: Our objective was to evaluate the efficacy and safety of Batroxobin on blood loss during spinal operations. METHODS: After obtaining approval from the ethics committee at the hospital along with informed written consent, we performed a double-blind, randomized, placebo-controlled study with 100 patients who were randomized equally into 2 groups (Batroxobin and placebo). Patients received either 2 ku IV 15 min before surgery and followed 1 ku IM of Batroxobin following surgery, or an equivalent volume of placebo (normal saline). Cost of Batroxobin treatment is amounted to 84.75 euros. The primary outcomes were intraoperative, 24 h postoperative, and total perioperative blood loss. Secondary outcomes were hemoglobin (Hb), red blood cell count (RBC), the volume of blood/fluid transfusion intraoperatively, and 24 h postoperatively. Safety evaluation parameters were the incidence of venous thrombosis in the lower extremities, active partial thromboplastin time, prothrombin time, thrombin time, and fibrinogen. The data were analyzed using the Statistical Package for the Social Science Version 12.0. The results were presented as mean ± SEM. The Mann-Whitney test and Independent Student t test, when appropriate, were used to compare the 2 groups, and differences were considered significant if the P value was <0.05. RESULTS: 88 patients were included in the analysis while 12 patients were withdrawn from the study due to extended surgical duration, change of surgical procedure, or after the patients' request. The total perioperative blood loss was approximately 31% lower in patients given Batroxobin versus placebo (700.5 ± 45.81 vs 485.7 ± 30.01 mL, P = 0.001). The Batroxobin group had significantly less intraoperative blood loss (326.1 ± 24.16) compared to the placebo group (556.0 ± 43.58), but there was no difference in the amount of blood/fluid transfused, postoperatively Hb, or RBC between the two groups. After the operation, coagulation parameters were not significantly different between the 2 groups at the days 1 or 3 postoperatively. No adverse events related to the use of Batroxobin were recorded. There were no cases of superficial wound infection. None of the subjects died during the study. CONCLUSIONS: In this study, prophylactic use of Batroxobin provided an effective and cheap method for reducing blood loss without coagulopathy during or after operations. The use of Batroxobin for patients undergoing one-level PLIF surgery safely and effectively reduced the total amount of perioperative blood loss.

Long non-coding RNA CARLo-5 is a negative prognostic factor and exhibits tumor pro-oncogenic activity in non-small cell lung cancer.

Despite recent advances in the management of non-small cell lung cancer (NSCLC), the prognosis of NSCLC is still relatively poor. Thus, it is of great importance to identify novel effective diagnostic and prognostic biomarker of NSCLC. A growing volume of literature has demonstrated the vital roles of long non-coding RNAs (lncRNAs) in cancer biology. Cancer-associated region long non-coding RNA (CARLo-5), a recently identified lncRNA, was initially characterized in colon cancer. However, little is known about its role in NSCLC. In the present study, a great upregulation of CARLo-5 was observed in cancer tissues compared to their adjacent normal tissues. Meanwhile, patients with high CARLo-5 expression have significantly poorer prognosis than those with low expression. Inhibition of CARLo-5 by siRNA suppressed the proliferation, migration, and invasion in NSCLC cell lines in vitro. In addition, silencing of CARLo-5 reversed the epithelial-mesenchymal transition in NSCLC cell line. Our study suggests that CARLo-5 may represent a prognostic marker and a potential therapeutic target of NSCLC.

Human leucocyte antigen alleles and haplotypes and their associations with antinuclear antibodies features in Chinese patients with primary biliary cirrhosis.

BACKGROUND & AIMS: Primary biliary cirrhosis (PBC) is an autoimmune liver disease. Genetic factors are critical in determining susceptibility to PBC. Among human leuocyte antigen (HLA) genes, an association between the DRB1*08 allele and PBC has been reported in many populations, but not in Chinese patients. METHODS: We investigated HLA-A, B, DRB1, and DQB1 alleles and haplotypes in 145 PBC patients and 500 healthy subjects. Patients were also stratified according to autoantibody features, and associations between these and HLA alleles were analyzed. RESULTS: Significant associations existed between HLA-DRB1*08:03 (22.1% vs. 9.0%, Pc < 0.0001, OR = 2.86), DQ2 (41.4% vs. 25.4%, Pc < 0.0001, OR = 2.07) and DQB1*06:01 (31.0% vs. 17.8%, Pc = 0.014, OR = 2.08) alleles and PBC. DRB1*08:03-DQB1*06:01 (22.1% vs. 8.2%, P < 0.0001, OR = 3.17) and DRB1*07:01-DQB1*02:02 haplotypes (28.3% vs. 17.6%, P = 0.005, OR = 1.85) were also associated with PBC susceptibility. In contrast, the DQB1*03:01 allele (21.4% vs. 39.2%, Pc < 0.0001, OR = 0.42) and DRB1*12:02-DQB1*03:01 haplotype (6.9% vs. 14.6%, P = 0.015, OR = 0.43) were significantly decreased in PBC patients compared with controls. DRB1*14:54 and DQ5(1) protected against antinuclear antibody (ANA) (OR = 0.25) and anti-gp210 antibody (OR = 0.39) production, respectively, while HLA-B*44:03 predisposed patients to anti-gp210 antibody (OR = 5.70) production. CONCLUSION: These results suggest that Chinese patients with PBC have a distinct genetic background in eastern Asia, and we confirmed the role of HLA genes in determining PBC susceptibility and autoantibody features in the Chinese population.

Causal association between systemic lupus erythematosus and primary biliary cholangitis: A bidirectional Mendelian randomization study.

An association has been observed between systemic lupus erythematosus (SLE) and primary biliary cholangitis (PBC) in observational studies, however, the exact causal link remains unclear. We aim to evaluate the causal relationships between SLE and PBC through bidirectional Mendelian randomization (MR). Single-nucleotide polymorphisms (SNPs) were selected as instrumental variables from publicly accessible genome-wide association studies (GWAS) in European populations. The PBC and SLE GWAS data were obtained from the MRC IEU Open GWAS database, consisting of 24,510 and 14,267 samples, respectively. After a series of quality control and outlier removal, inverse variance weighted was used as the primary approach to evaluate the causal association between SLE and PBC. The horizontal pleiotropy and heterogeneity were examined by the MR-Egger intercept test and Cochran Q value, respectively. Seven SNPs were included to examine the causal effect of SLE on PBC. Genetically predicted SLE may increase the risk of PBC development, with an odds ratio (OR) of 1.324 (95% confidence interval [CI] 1.220 ∼ 1.437, P ˂ .001). Twenty SNPs were included to explore the causal effect of PBC on SLE. Genetically predicted PBC may increase the risk of SLE development, with an OR of 1.414 (95% CI 1.323 ∼ 1.511, P ˂ .001). Horizontal pleiotropy and heterogeneity were absent (P > .05) among SNPs. The robustness of our results was further enhanced by using the leave-one-out method. Our research has provided new insights into SLE and PBC, indicating bidirectional causal associations between the 2 diseases. These findings offer valuable contributions to future clinical studies.

[Screening for EGFR mutations in lung cancer by a novel real-time PCR with double-loop probe and Sanger DNA sequencing].

OBJECTIVE: To map the frequency and types of EGFR gene mutations present in lung cancer tissues. To evaluate the clinical applicability of a novel real-time double-loop probe PCR of which the ADx-EGFR kit is based, and to compare its performance with traditional Sanger DNA sequencing in the detection of somatic mutations of tumor genes. METHODS: A total of 208 formalin-fixed paraffin-embedded (FFPE) tumor samples were tested. Genomic DNA of the tissue samples was extracted and purified, and subjected to both traditional PCR amplification, Sanger sequencing of EGFR gene in exon 18, 19, 20, 21, and ADx's EGFR mutation detection kit. The mutation rates for EGFR gene in exon 18, 19, 20, 21, as well as the frequency of each mutation detected by the two methods, were analyzed. RESULTS: The traditional Sanger DNA sequencing technique was successfully performed in 196 out of 208 (94.2%) lung cancer samples, and 22 samples (11.2%) showed EGFR gene mutations. ADx-EGFR kit was successfully used in the lung cancers of all of the 208 cases (100.0%), and 40 samples (19.2%) showed mutations. In the lung cancer samples analyzed, mutations were mainly detected in the exon 19 and exon 21 L858R point mutation, i.e. 4.8% (10/208) and 11.6% (23/208) of total mutations, respectively, and the remaining mutations were rare. CONCLUSIONS: The success rate of ADx-EGFR real-time PCR for formalin-fixed and paraffin-embedded tissues samples is significantly higher than that of Sanger sequencing (P < 0.01). There are significant differences between the two methods. ADx-EGFR real-time PCR shows a much higher successful detection rate and mutation rate of lung cancer tissues compared with that of Sanger sequencing. As a result, the real-time PCR with ADx-EGFR kit is proved to have a good clinical applicability and a strong advantage over the traditional Sanger DNA sequencing. It is an effective and reliable tool for clinical screening of somatic gene mutations in tumors.

[Detection of EML4-ALK fusion gene in non-small cell lung cancer and its clinicopathologic correlation].

OBJECTIVE: To investigate the frequency of EML4-ALK fusion gene in non-small-cell lung cancer (NSCLC) patients, and its correlation with clinicopathologic features. METHODS: Real-time PCR was used to detect the presence of EML4-ALK fusion gene in 268 cases of NSCLCs using paraffin-embedded tissue samples(among which 164 samples were re-validated by Sanger sequencing). Related clinicopathological correlation was analyzed. RESULTS: EML4-ALK fusion gene was found in 4.1% (11/268) of the cases. One hundred and sixty four samples were verified by Sanger sequencing, and the overall coincidence of the results of two methods (Sanger sequencing and Real-time PCR) was 100%. Female patients (5.9%, 5/85), ≤ 60 years of age (4.3%, 6/140), non-smokers (6.8%, 8/118) and adenocarcinomas (7.6%, 10/132) had a higher mutation rate than that in male patients (3.3%, 6/183), > 60 years of age (4.0%, 5/124), smokers (1.6%, 2/132) and squamous cell carcinomas (1.3%, 1/79), although no statistical significance in age (P = 0.918), gender (P = 0.503), smoking history (P = 0.092) and histological type (P = 0.094). CONCLUSIONS: Chinese NSCLC patients have a 4.1% detection rate of EML4-ALK fusion gene in the tumor tissues. Female, non-smoker and adenocarcinoma histological subtype tend to be associated with a higher rate of EML4-ALK gene fusion.

[Evaluation of endobronchial ultrasound-guided transbronchial needle aspiration in the diagnosis of hilar and mediastinal tumors].

OBJECTIVE: To explore the diagnostic values of endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) in patients with hilar and mediastinal tumors. METHODS: A total of 837 patients with chest CT or PET/CT confirmed mediastinal/hilar lymphadenopathy and or tumors in (or around) the trachea or bronchi, were evaluated by EBUS-TBNA examination. Pathological study or follow-up visit was carried out in the same period to make a final diagnosis, and therefore to verify the accuracy of EBUS-TBNA. RESULT: The study punctured 1631 lymph nodes totally, with an average of 1.95 times per case. The 4R group and the 7th group of lymph nodes accounted for 43% and 34%, respectively. The success rate of TBNA was 100%. The diagnosis rates of lung cancer was 94.11%, of which squamous cell carcinoma accounted for 19.22% (89 cases), adenocarcinoma 32.40% (150 cases), small cell carcinoma 23.54% (109 cases), non-small cell carcinoma of unknown histological type 9.50% (44 cases), adenosquamous carcinoma 7.56% (35 cases), and other types 7.78% (36 cases). The diagnosis rate of tuberculosis was 85.50%, and that of sarcoidosis was 55.88%. The overall sensitivity of EBUS-TBNA was 94.02%, specificity 100%, positive predictive value 100%, negative predictive value 61.91%, and the accuracy was 94.56%. Besides mild bleeding in the puncture site, no other complications occurred, and there were no severe complications such as pneumothorax, pneumomediastinum, or major vascular injury. CONCLUSION: EBUS-TBNA is of high value as a minimally invasive, convenient, and low-risk procedure for the diagnosis of mediastinal and hilar lymphadenopathy and tumors.

[Investigate circulating levels of chemokines and evaluate the correlation between these chemokines and liver function indicators in autoimmune hepatitis].

OBJECTIVE: This study investigated circulation levels of chemokines (CCL2, CCL5, CXCL8, CXCL9, CXCL10) in autoimmune hepatitis(AIH) patients and evaluated the correlation between these chemokines and liver function indicators. METHODS: A total of 5 chemokines (CCL2, CCL5, CXCL8, CXCL9, CXCL10) were measured simultaneously by cytokine beads assay(CBA) in the sera of 46 patients with AIH and 12 cases of healthy control. RESULTS: In this study we found that serum levels of CCL2 , CXCL9 and CXCL10 in AIH patients and healthy controls were 11.79:8.39 pg/ml, 11.31:2.69 pg/ml, 15.85:4.64 pg/ml, respectively , which implied these chemokines were significantly higher in AIH patients when compared to healthy control (Z=-1.958, P=0.05; Z=-4.527, P less than 0.0001; Z=-3.84, P less than 0.0001, respectively). And circulation levels of CCL2 , CXCL8 , CXCL9 and CXCL10 in pretreatment and remission stages of patients with AIH were 29.69:11.16 pg/ml, 7.2:5.38 pg/ml, 16.02:5.47 pg/ml, 90.01:13.24 pg/ml, respectively, which showed these chemokines decreased during remission from pretreatment stage levels (t=2.985, P=0.005; Z=-2.547, P=0.0112; Z=-3.187, P=0.001; t=2.12, P=0.0015, respectively). Among AIH , CXCL8 was correlated positively with lgG(r2=0.291, P=0.0039); CXCL9 was associated positively with ALT and AST(r2=0.5324 , P less than 0.0001; r2=0.3352, P less than 0.0001); CXCL10 showed a positive correlation with ALT , AST and GGT(r2=0.9551, P less than 0.0001; r2=0.8960, P less than 0.0001; r2=0.8271, P less than 0.0001). CONCLUSION: Serum levels of CCL2, CXCL8, CXCL9 and CXCL10 are significantly higher in patients with AIH, but decrease to levels in healthy controls after successful treatment , and circulation levels of CXCL9 and CXCL10 are associated positively with liver function indicators which can react inflammation activity of liver, all these may imply that chemokines can reflect the degree of liver inflammation and may be one of the main culprits in AIH pathological damage.

[Health Assessment of the Stream Ecosystem in Beijing, China].

With economic development, the health of river ecosystems is becoming severely threatened because of the increasing effects of human activities on river ecosystems. Here, 101 sites along regional river systems in Beijing rivers were investigated from autumn 2020 to summer 2021. A total of 34 metrics, including aquatic organisms, hydrology, water quality, and habitat, were calculated to be the candidate indicators. Principal component and correlation analyses were used to select the core metrics from the candidate indicators, and the weight of each core metric was estimated using the entropy method. The integrated index of stream ecological health was constructed to assess the health condition of the Beijing rivers. The results of the PCA and correlation analyses revealed that eleven metrics were selected as the core metrics to construct the integrated index of stream ecological health, including water temperature, flow velocity, BOD5, NH4+-N, Cu, the density of phytoplankton and zooplankton, the Shannon-Wiener diversity index of macroinvertebrates and fish, the BMWP index, and the qualitative habitat evaluation index. According to the health assessment results, 4.95% of the sampling sites were healthy, 23.76% were subhealthy, and 71.29% were in a fair or below healthy state. The river health status showed strong spatial heterogeneity, and the river health statuses in the northern and western regions were relatively good, whereas the river health status in the central and southeastern regions were relatively poor. The results of four aspects stream ecosystem assessment showed that the overall water quality of the rivers was "subhealthy" and the aquatic organisms and habitat were "general poor," but the hydrology was "poor." The evaluation results of five water systems demonstrated that the Chaobai River had the best health status, followed by that of the Yongding River, Daqing River, and Jiyun River, and the Beiyun River had the worst health status. Maintaining river ecological baseflow, ensuring river system connectivity, and improving and restoring the river habitat environment are the key aspects of river ecological restoration and protection in Beijing in the future.

Identification and validation of coding and non-coding RNAs involved in high-temperature-mediated seed dormancy in common wheat.

Introduction: Seed dormancy (SD) significantly decreases under high temperature (HT) environment during seed maturation, resulting in pre-harvest sprouting (PHS) damage under prolonged rainfall and wet weather during wheat harvest. However, the molecular mechanism underlying HT-mediated SD remains elusiveSeed dormancy (SD) significantly decreases under high temperature (HT) environment during seed maturation, resulting in pre-harvest sprouting (PHS) damage under prolonged rainfall and wet weather during wheat harvest. However, the molecular mechanism underlying HT-mediated SD remains elusive. Methods: Here, the wheat landrace 'Waitoubai' with strong SD and PHS resistance was treated with HT from 21 to 35 days post anthesis (DPA). Then, the seeds under HT and normal temperature (NT) environments were collected at 21 DPA, 28 DPA, and 35 DPA and subjected to whole-transcriptome sequencing. Results: The phenotypic data showed that the seed germination percentage significantly increased, whereas SD decreased after HT treatment compared with NT, consistent with the results of previous studies. In total, 5128 mRNAs, 136 microRNAs (miRNAs), 273 long non-coding RNAs (lncRNAs), and 21 circularRNAs were found to be responsive to HT, and some of them were further verified through qRT-PCR. In particular, the known gibberellin (GA) biosynthesis gene TaGA20ox1 (TraesCS3D02G393900) was proved to be involved in HT-mediated dormancy by using the EMS-mutagenized wheat cultivar Jimai 22. Similarly, a novel gene TaCDPK21 (TraesCS7A02G267000) involved in the calcium signaling pathway was validated to be associated with HT-mediated dormancy by using the EMS mutant. Moreover, TaCDPK21 overexpression in Arabidopsis and functional complementarity tests supported the negative role of TaCDPK21 in SD. We also constructed a co-expression regulatory network based on differentially expressed mRNAs, miRNAs, and lncRNAs and found that a novel miR27319 was located at a key node of this regulatory network. Subsequently, using Arabidopsis and rice lines overexpressing miR27319 precursor or lacking miR27319 expression, we validated the positive role of miR27319 in SD and further preliminarily dissected the molecular mechanism of miR27319 underlying SD regulation through phytohormone abscisic acid and GA biosynthesis, catabolism, and signaling pathways. Discussion: These findings not only broaden our understanding of the complex regulatory network of HT-mediated dormancy but also provide new gene resources for improving wheat PHS resistance to minimize PHS damage by using the molecular pyramiding approach.