Effect of bladder filling status on positioning errors in post-hysterectomy cervical cancer radiotherapy.

Objective: To investigate the effect of different bladder filling states on positioning errors in radiotherapy for cervical cancer and obtain the reference range of bladder filling consistency during radiotherapy.Methods: Patients who underwent postoperative radiotherapy for cervical cancer in Nantong Tumor Hospital from October 2018 to December 2019 were selected. According to the bladder filling deviation, they were divided into group A1 (deviation < 20%) and group B1 (deviation ≥ 20%). The bladder filling variations of the two groups were compared with different positioning errors. Group A2 has a positioning error of <0.4 cm, and group B2 has a positioning error of ≥0.4 cm. The reference range of bladder filling consistency during radiotherapy is obtained by analyzing the composition ratio of different positioning errors of bladder filling deviation.Results: This study included 195 patients with cervical cancer. The error of longitudinal and vertical position in group B1 was significantly higher than that in group A1 (0.50 ± 0.34 vs. 0.26 ± 0.22 cm, p < 0.001, and 0.22 ± 0.17 vs. 0.16 ± 0.12 cm, p < 0.001). Compared with group B2, the absolute deviation of bladder filling in group A2 (54.1% ± 54.4% vs. 25.6% ± 22.7%, p < 0.001) was slight. The chi-square test showed significant differences in the proportion of the positioning state of different bladder filling forms (χ2 = 31.006, p < 0.001). In addition, there was a significant difference in the proportion of stability errors in patients with poor stability in different directions (χ2 = 118.551, p < 0.001).Conclusion: In patients with cervical cancer fixed in the supine position, a bladder capacity deviation <20% is easier to achieve excellent positioning with, and it can better control the positioning error of radiotherapy and ensure the positioning accuracy of dose distribution to the target area. It can also achieve good tumor treatment effects. This range can be used as a reference for bladder filling consistency in patients with cervical cancer undergoing radiotherapy.

Targeted Methylation of the LncRNA NEAT1 Suppresses Malignancy of Renal Cell Carcinoma.

Long-chain non-coding RNA (LncRNA) has been found to play an important role in the regulation of the occurrence and progression of renal cell carcinoma (RCC). In this study, we demonstrated that LncRNA NEAT1 expression and m6A methylation level was decreased in RCC tissues. Further, the downregulated expression level of LncRNA NEAT1 was associated with poor prognosis for RCC patients. Then we used CRIPSR/dCas13b-METTL3 to methylate LncRNA NEAT1 in RCC cells. The results showed that the expression level of LncRNA NEAT1 was upregulated after methylated by dCas13b-METTL3 in RCC cells. And the proliferation and migration ability of RCC cells was decreased after methylated LncRNA NEAT1. Finally, we examined the effect of LncRNA NEAT1 hypermethylation on the transcriptome. We found differentially expressed genes in RCC cells were associated with "cGMP-PKG signaling pathway", "Cell adhesion molecules" and "Pathways in cancer". In conclusion, CRISPR/Cas13b-METTL3 targeting LncRNA NEAT1 m6A methylation activates LncRNA NEAT1 expression and provides a new target for treatment of RCC.

Independent and joint correlation of PNPLA3 I148M and TM6SF2 E167K variants with the risk of coronary heart disease in patients with non-alcoholic fatty liver disease.

BACKGROUND: CHD is reported to be the primary cause of death in patients with NAFLD. Genetic susceptibility genes contribute to the developmental risk of NAFLD or CHD. Whether the genetic factors could affect the risk of CHD in NAFLD patients is not clear. The aim of this study was to investigate the association of PNPLA3 I148M and TM6SF2 E167K variants with the risk of CHD in NAFLD patients in Chinese Han population. PATIENTS AND METHODS: PNPLA3 I148M and TM6SF2 E167K variants were genotyped in a cohort of 189 patients with NAFLD and CHD, as well as 242 patients with NAFLD and 242 healthy controls by gene sequencing. Additionally, serum lipids profiles were determined by standard clinical laboratory methods. RESULTS: The minor allele frequency of PNPLA3 I148M and TM6SF2 E167K were 0.39 and 0.06 in this cohort, respectively. The distributions of PNPLA3 I148M genotypes and alleles were significant different in NAFLD group vs controls and in NAFLD+CHD group vs NAFLD group (all P <  0.05). NAFLD patients who carry the CG + GG genotype suffered the relative lower risk of CHD than CC genotype carriers (OR = 0.6, 95%CI: 0.40-0.90, P = 0.01). In addition, PNPLA3 I148M and TM6SF2 E167K possess the joint correlation with the decreased risk of CHD in NAFLD patients with the increased number of risk alleles. Besides, PNPLA3 I148M and TM6SF2 E167K variants associated with the decreased serum lipid levels in overall series. CONCLUSIONS: There was a joint protective correlation of PNPLA3 I148M and TM6SF2 E167K variants with the developmental risk of CHD in NAFLD patients. PNPLA3 I148M and TM6SF2 E167K variants might correlated with the decreased risk of CHD in NAFLD patients by associated with the reduced serum lipid levels.

CRISPR-Cas13-Mediated Knockdown of lncRNA-GACAT3 Inhibited Cell Proliferation and Motility, and Induced Apoptosis by Increasing p21, Bax, and E-Cadherin Expression in Bladder Cancer.

The current study is to investigate the expression pattern and biological function of long non-coding RNA Focally gastric cancer-associated transcript3 (GACAT3) in bladder cancer. Real-time quantitative qPCR was used to detect the expression level of GACAT-3 in tumor tissues and paired normal tissues. Human bladder cancer T24 and 5637 cell lines were transiently transfected with specific CRISPR-Cas13 or negative control CRISPR-Cas13. Cell migration, proliferation, and apoptosis were measured by using wound healing assay CCK-8 assay and Caspase-3 ELISA assay, respectively. The expression changes of p21, Bax, and E-cadherin after knockdown of GACAT3 were detected by using Western blot. The results demonstrated that GACAT3 was up-regulated in bladder cancer tissues than that in the paired normal tissues. Inhibition of cell proliferation, increased apoptosis, and decreased motility were observed in T24 and 5637 cell lines transfected by CRISPR-Cas13 targeting GACAT3. Downregulation of GACAT3 increased p21, Bax, and E-cadherin expression and silencing these genes could eliminate the phenotypic changes induced by knockdown of GACAT3. A ceRNA mechanism for GACAT3 was also revealed. By using CRISPR-Cas13 biotechnology, we suggested that GACAT3 may be a novel target for diagnosis and treatment of bladder cancer.

[Medication versus health education for patients with type â ¢ prostatitis-like symptoms: A prospective randomized control study].

OBJECTIVE: To investigate the necessity of medication for patients with type Ⅲ prostatitis-like symptoms for less than 3 months. METHODS: We enrolled in this study 171 outpatients with type Ⅲ prostatitis-like symptoms for less than 3 months in our hospital from November 2016 to October 2017, and randomly divided them into groups A (n = 57), B (n = 57) and C (n = 57). The patients of group A received tamsulosin, levofloxacin and health education, those of group B tamsulosin and health education, and those of group C health education only. Three months later, we evaluated the therapeutic effects according to the National Institutes of Health Chronic Prostatitis Symptom Index (NIH-CPSI) scores of the patients, 4-point reduction in the total score indicating effectiveness. RESULTS: After 3 months of treatment, the total NIH-CPSI scores of the patients in groups A, B and C were decreased by (9.0 ± 2.9), (8.2 ± 3.4) and (8.6 ± 3.2) points respectively, all indicating effectiveness, the pain scores (4.2 ± 1.8), (4.0 ± 1.9) and (4.2 ± 1.6) points, the urinary symptom scores decreased by decreased by (2.4 ± 1.2), (2.4 ± 1.4) and (2.2 ± 1.2) points, and quality of life scores decreased by (2.4 ± 1.4), (1.9 ± 1.4) and (2.2 ± 1.3) points, none with statistically significant difference among the three groups (P > 0.05). CONCLUSIONS: Health education is proved to have a therapeutic effect on type Ⅲ prostatitis-like symptoms similar to that of alpha receptor blockers.

rs11614913 polymorphism in miRNA-196a2 and cancer risk: an updated meta-analysis.

Several epidemiological studies have reported that polymorphisms in microRNA-196a2 (miR-196a2) were associated with various cancers. However, the results remained unverified and were inconsistent in different cancers. Therefore, we carried out an updated meta-analysis to elaborate the effects of rs11614913 polymorphism on cancer susceptibility. A total of 84 articles with 35,802 cases and 41,541 controls were included to evaluate the association between the miR-196a2 rs11614913 and cancer risk by pooled odds ratios (ORs) and 95% confidence intervals (CIs). The results showed that miR-196a2 rs11614913 polymorphism is associated with cancer susceptibility, especially in lung cancer (homozygote comparison, OR =0.840, 95% CI =0.734-0.961; recessive model, OR =0.858, 95% CI =0.771-0.955), hepatocellular carcinoma (allelic contrast, OR =0.894, 95% CI =0.800-0.998; homozygote comparison, OR =0.900, 95% CI =0.813-0.997; recessive model, OR =0.800, 95% CI =0.678-0.944), and head and neck cancer (allelic contrast, OR =1.076, 95% CI =1.006-1.152; homozygote comparison, OR =1.214, 95% CI =1.043-1.413). In addition, significant association was found among Asian populations (allele model, OR =0.847, 95% CI =0.899-0.997, P=0.038; homozygote model, OR =0.878, 95% CI =0.788-0.977, P=0.017; recessive model, OR =0.895, 95% CI =0.824-0.972, P=0.008) but not in Caucasians. The updated meta-analysis confirmed the previous results that miR-196a2 rs11614913 polymorphism may serve as a risk factor for patients with cancers.

Knockdown of long noncoding RNA FGFR3- AS1 induces cell proliferation inhibition, apoptosis and motility reduction in bladder cancer.

OBJECTIVES: To study the expression pattern of long non-coding RNA FGFR3 antisense transcript 1(FGFR3-AS1) and the cell proliferation inhibition, apoptosis, and motility changes induced by silencing FGFR3-AS1 in bladder cancer. METHODS: The differential expression levels of FGFR3-AS1 and FGFR3 in tumor tissues and paired normal tissues were determined using Real-Time qPCR in a total of 36 patients diagnosed with bladder cancer (urothelial carcinoma). Pearson's coefficient correlation was used for expression correlation assay. Expression differences of FGFR3-AS1 were analyzed according to grading and staging. FGFR3 protein was detected by western blot assay. Human bladder cancer T24 and 5637 cell lines were transiently transfected with FGFR3-AS1-specific siRNA or negative control siRNA. The cell proliferation changes of transfected bladder cancer cells were determined using CCK-8 assay. Apoptosis caused by knockdown of FGFR3-AS1 was evaluated using ELISA assay. Motility changes induced by knockdown of FGFR3-AS1 were measured using wound healing assay and transwell assay. RESULTS: Both FGFR3-AS1 and FGFR3 were overexpressed in bladder cancer tissues compared to matched normal tissues. They were also positively expressed in bladder cancer. FGFR3-AS1 expression levels were higher in high grade tumors than those in low grade tumors. FGFR3-AS1 expression levels were higher in invasive tumors than those in non-invasive tumors. Cell proliferation inhibition, increased apoptosis, and decreased motility were observed in FGFR3-AS1 siRNA-transfected T24 and 5637 cell lines. CONCLUSIONS: FGFR3-AS1 plays an oncogenic role in human bladder cancer. Knockdown of FGFR3-AS1 may provide a potential new therapeutic approach to this disease.

Hsa-miR-429 promotes bladder cancer cell proliferation via inhibiting CDKN2B.

BACKGROUND AND OBJECTIVES: Hsa-miR-429 is increased in bladder cancer. Its roles in bladder cancer are poorly understood. METHODS: The expression levels of hsa-miR-429 and cyclin-dependent kinase inhibitor 2B (CDKN2B) were determined using Real-Time qPCR in a total of 50 patients with bladder cancer. Bladder cancer T24 and 5637 cells were transfected CDKN2B siRNA or hsa-miR-429 mimic. CDKN2B expression levels after transfection were detected by Real-Time qPCR and Western blot assay respectively. Binding sites between hsa-miR-429 and 3'-untranslated region of CDKN2B were confirmed by Dual luciferase reporter assay. Cell proliferation was evaluated using MTT and EdU assays. Cell apoptosis was determined using ELISA assay. RESULTS: Higher hsa-miR-429 expression levels were associated with higher tumor grade and stage. All patients with low hsa-miR-429 expression survived 5 years, while with high hsa-miR-429 expression, only 58% survived. Hsa-miR-429 and CDKN2B were inversely expressed in bladder cancer. Hsa-miR-429 mimic decreased the expression of CDKN2B at both mRNA and protein levels. The binding site was confirmed between hsa-miR-429 and 3'-untranslated region of CDKN2B. Up-regulation of hsa-miR-429 or down-regulation of CDKN2B promoted cell growth and decreased apoptosis. CONCLUSIONS: Our data suggest a mechanism for hsa-miR-429 to play oncogenic roles via inhibiting CDKN2B.

Down-regulation of C12orf59 is associated with a poor prognosis and VHL mutations in renal cell carcinoma.

C12orf59 is newly identified gene in kidney. However, the relation of C12orf59 expression and clinic features is unknown. Here, our study showed that C12orf59 was broadly expressed in normal human tissues with high expression levels in kidney while its expression is beyond detectable in a panel of cancer cell lines. C12orf59 expression in RCC was significantly decreased compared with corresponding adjacent noncancerous tissues (P < 0.01). The decreased C12orf59 expression was correlated with lymph node status (P < 0.05), distant metastases (P < 0.05), poor survival (P < 0.001) (HR 3.00; 95% CI, 1.29-7.53), VHL non-sense mutations or frame-shift mutations (P < 0.01), and UMPP gene non-sense mutations or frame-shift mutations (P = 0.01). Thus, we propose that the decreased C12orf59 expression status is a prognostic biomarker of ccRCC and cooperates with the loss of VHL all the while promoting renal carcinogenesis.

Downregulated ECRG4 is associated with poor prognosis in renal cell cancer and is regulated by promoter DNA methylation.

Esophageal cancer-related gene 4 (ECRG4) has been proposed as a putative tumor suppressor gene in several tumors. However, the role and regulation of ECRG4 in the pathogenesis of human renal cancer remain largely unknown. Our current study revealed that expression of ECRG4 is downregulated in renal cell lines and renal cancer tissues. ECRG4 expression was significantly associated with histological grade of tumors (p < 0.001), primary tumor stage (p = 0.017), and distant metastasis (p = 0.017). Low expression of ECRG4 was an independent prognostic indicator for survival of renal cancer patients. Silencing of ECRG4 expression in renal cell lines was associated with its promoter methylation. Moreover, ectopic expression of ECRG4 markedly inhibited cell proliferation and invasion in renal cancer cell lines. These results indicated that ECRG4 is frequently silenced by the methylation of promoter in renal cell cancers. ECRG4 may be a tumor suppressor in renal cancer and serve as a prognostic marker.

Pyruvate-associated acid resistance in bacteria.

Glucose confers acid resistance on exponentially growing bacteria by repressing formation of the cyclic AMP (cAMP)-cAMP receptor protein (CRP) complex and consequently activating acid resistance genes. Therefore, in a glucose-rich growth environment, bacteria are capable of resisting acidic stresses due to low levels of cAMP-CRP. Here we reveal a second mechanism for glucose-conferred acid resistance. We show that glucose induces acid resistance in exponentially growing bacteria through pyruvate, the glycolysis product. Pyruvate and/or the downstream metabolites induce expression of the small noncoding RNA (sncRNA) Spot42, and the sncRNA, in turn, activates expression of the master regulator of acid resistance, RpoS. In contrast to glucose, pyruvate has little effect on levels of the cAMP-CRP complex and does not require the complex for its effects on acid resistance. Another important difference between glucose and pyruvate is that pyruvate can be produced by bacteria. This means that bacteria have the potential to protect themselves from acidic stresses by controlling glucose-derived generation of pyruvate, pyruvate-acetate efflux, or reversion from acetate to pyruvate. We tested this possibility by shutting down pyruvate-acetate efflux and found that the resulting accumulation of pyruvate elevated acid resistance. Many sugars can be broken into glucose, and the subsequent glycolysis generates pyruvate. Therefore, pyruvate-associated acid resistance is not confined to glucose-grown bacteria but is functional in bacteria grown on various sugars.

CMTM3 inhibits human testicular cancer cell growth through inducing cell-cycle arrest and apoptosis.

Human CMTM3 has been proposed as a putative tumor suppressor gene. The loss of CMTM3 has been found in several carcinomas. However, the regulation of CMTM3 expression and its function in tumor progression remain largely unknown. Here, we investigated the regulation of CMTM3 expression, function and molecular mechanism in human testicular cancer cells. CMTM3 was frequently downregulated or silenced in testicular cancer cell lines and tumor tissues but highly expressed in normal testis tissues. The re-expression of CMTM3 significantly suppressed the colony formation, proliferation, and migration capacity of testicular cancer cells by inducing a G2 cell cycle arrest and apoptosis. Moreover, the re-expression of CMTM3 activated the transcription of p53, induced p53 accumulation, up-regulated the expression of p21, and increased the cleavage of caspase 9, 8, 3, and PARP. The downregulation of CMTM3 in clinical tumor tissues was associated with the methylation of a single CpG site located within the Sp1/Sp3-responsive region of the core promoter. These results indicate that CMTM3 can function as tumor suppressor through the induction of a G2 cell cycle arrest and apoptosis. CMTM3 is thus involved in testicular cancer pathogenesis, and it is frequently at least partially silenced by the methylation of a single, specific CpG site in tumor tissues.

Modification of the RpoS network with a synthetic small RNA.

Translation of the sigma factor RpoS is activated by DsrA, RprA and ArcA, three small non-coding sRNAs (sRNA) that expose the ribosome-binding site (RBS) by opening up an inhibitory loop. In the RpoS network, no sRNAs have been found to pair with the RBS, a most common sRNA target site in bacteria. Here, we generate Ribo-0, an artificial sRNA, which represses rpoS translation by pairing with the RBS. Ribo-0 bypasses the RNA chaperon Hfq but requires the RBS to be loosely blocked. Ribo-0 interacts with DsrA and reshapes the RpoS network. Specifically, in the intact RpoS network, DsrA activates rpoS translation by freeing up the RBS. In the modified RpoS network where Ribo-0 is introduced, the DsrA-caused RBS exposure facilitates Ribo-0 binding, thereby strengthening Ribo-0 inhibition. In other words, Ribo-0 changes DsrA from an activator to an accomplice for repressing rpoS translation. This work presents an artificial mechanism of rpoS regulation, reveals mutual effects of native and synthetic players and demonstrates genetic context-dependency of their functions.

RALY RNA binding protein-like reduced expression is associated with poor prognosis in clear cell renal cell carcinoma.

The molecular mechanisms involved in the progression of clear cell renal cell carcinomas (ccRCCs) are still unclear. The aim of this study was to analyse the relationships between expression of RALYL and clinical characteristics. In 41 paired samples of ccRCCs and adjacent normal tissues, we used real-time qPCR to evaluate the expression of RALYL mRNA. RALYL protein levels were determined in 146 samples of ccRCC and 37 adjacent normal tissues by immunohistochemistry. Statistical analysis was used to explore the relationships between expression of RALYL and the clinical characteristics (gender, age, tumor size, T stage, N stage, M stage, survival times and survival outcome) in ccRCC. In addition, these patients were follow-up period 64 months (range: 4~116 months) to investigate the influence on prognosis. We found significantly differences between ccRCC tissues and normal tissues (p<0.001, paired-sample t test) in mRNA levels of RALYL. Immunohistochemistry analyses in 146 ccRCC samples and 37 adjacent normal tissues showed significantly lower RALYL protein levels in ccRCC samples (χ2-test, p<0.001), inversely correlating with tumour size (p=0.024), T stage (0.005), N stage (p<0.001) as well as M stage (p=0.019), but not age (p=0.357) and gender (p=0.348). Kaplan-Meier survival analysis demonstrated that people with lower level of RALYL expression had a poorer survival rate than those with a higher level of RALYL expression, significantly different by the log-rank test (p=0.011). Cox regression analysis indicated that RALYL expression (p=0.039), N stage (p=0.008) and distant metastasis (p<0.001) were independent prognosis factors for the overall survival of ccRCC patients. We demonstrated that the expression of RALYL was significantly low in ccRCC and correlated with a poor prognosis in a large number of clinical samples. Our findings showed that RALYL may be a potential therapeutic target as well as a poor prognostic factor.