MicroRNA-33a-5p suppresses colorectal cancer cell growth by inhibiting MTHFD2.

Colorectal cancer (CRC) is one of the most common malignancies with high levels of invasiveness, drug resistance and mortality, but the internal mechanisms of CRC are largely unknown. MicroRNAs (miRs) have been reported to be involved in the development of CRC, and numerous studies have demonstrated that the abnormal expression of miR-33a-5p might be associated with CRC. However, the function and downstream mechanism of miR-33a-5p in colorectal cancer (CRC) remains unclear. Methylenetetrahydrofolate dehydrogenase 2 (MTHFD2), a mitochondrial enzyme involved in folic acid metabolism, interestingly was confirmed to be one of the target genes of miR-33a-5p in the present study. We first confirmed that miR-33a-5p in CRC tissues and cell lines were downregulated (P < 0.05), and that the proliferation, clone formation capacities, G1/S progression, and migration capacities of the two CRC cell lines HCT116 cells and HT29 were suppressed by miR-33a-5p overexpression in vitro (P < 0.05). Ctrl HCT116 and miR-33a-5p-overexpressing HCT116 were injected into nude mice. In vivo tumour formation was significantly suppressed by miR-33a-5p overexpression (P < 0.05) as well as the proliferation marker Ki67 (P < 0.05). Additionally, MTHFD2 protein expression was significantly enhanced in CRC tissues. From bioinformatics predictions and a luciferase report analysis, MTHFD2 was confirmed to be one of the target genes of miR-33a-5p. In contrast to the role of miR-33a-5p overexpression, MTHFD2 overexpression promoted the proliferation and migration of HCT116 and HT29 cells (P < 0.05), which confirmed that MTHFD2 was a functional target gene of miR-33a-5p. In conclusion, miR-33a-5p inhibits the growth and migration of CRC by targeting MTHFD2.

Single-cell transcriptomic analysis of normal and pathological tissues from the same patient uncovers colon cancer progression.

The aim of the present study was to elucidate the evolutionary trajectory of colon cells from normal colon mucosa, to adenoma, then to carcinoma in the same microenvironment. Normal colon, adenoma and carcinoma tissues from the same patient were analyzed by single-cell sequencing, which perfectly simulated the process of time-dependent colon cancer due to the same microenvironment. A total of 22 cell types were identified. Results suggest the presence of dominant clones of same cells including C2 goblet cell, epithelial cell subtype 1 (Epi1), enterocyte cell subset 0 (Entero0), and Entero5 in carcinoma. Epi1 and Entero0 were Co-enriched in antibacterial and IL-17 signaling, Entero5 was enriched in immune response and mucin-type O-glycan biosynthesis. We discovered new colon cancer related genes including AC007952.4, NEK8, CHRM3, ANO7, B3GNT6, NEURL1, ODC1 and KCNMA1. The function of TBC1D4, LTB, C2CD4A, AND GBP4/5 in T cells needs to be clarified. We used colon samples from the same person, which provide new information for colon cancer therapy.

Abdominal bronchogenic cyst: A rare case report.

BACKGROUND: Bronchogenic cysts are cystic masses caused by congenital abnormal development of the respiratory system, and usually occur in the pulmonary parenchyma or mediastinum. CASE SUMMARY: A rare case of a bronchogenic cyst discovered in the abdominal cavity of a 35-year-old man is reported. Physical examination found a space-occupying lesion in the patient's abdomen for 4 d. Laparoscopic exploration found the cyst tightly adhered to the stomach and its peripheral blood vessels; therefore, intraoperative laparotomy was performed. The cystic mass was resected en bloc with an Endo-GIA stapler. The final postoperative pathological diagnosis confirmed an abdominal bronchogenic cyst. CONCLUSION: This is a rare case of a bronchogenic cyst that was discovered within the abdominal cavity of a male patient. The cyst is easily confused with or misdiagnosed as other lesions. Therefore, it is necessary to distinguish abdominal bronchogenic cyst from gastrointestinal stromal tumor, Meckel's diverticulum, enteric duplication cyst, or lymphangioma. Although computer tomography and magnetic resonance imaging were the primary diagnostic approaches, endoscopic ultrasound-guided fine-needle aspiration could assist with clarification of the cytological or histopathological diagnosis before surgery.

FERM domain-containing protein 6 exerts a tumor-inhibiting role in thyroid cancer by antagonizing oncogenic YAP1.

The emerging role of FERM domain-containing protein 6 (FRMD6) in cancer progression has been revealed in several malignancies. However, its relevance on thyroid cancer is not well understood. This work evaluated the possible role and mechanism of FRMD6 in thyroid cancer. We demonstrated that FRMD6 expression was downregulated in thyroid cancer by analyzing the Cancer Genome Atlas data. Remarkable reductions in FRMD6 expression were also confirmed in the clinical specimens and cell lines of thyroid cancer. The upregulation of FRMD6 restrained the proliferation, epithelial-mesenchymal transition, and invasion of thyroid cancer. Moreover, FRMD6 overexpression significantly increased the apoptosis and cell cycle arrest. Further molecular research demonstrated that the overexpression of FRMD6 increased the phosphorylation levels of mammalian STE20-like protein kinase 1, large tumor suppressor 1, and Yes-associated protein 1 (YAP1) and prohibited the activation of YAP1. The re-expression of constitutively active YAP1 strikingly reversed FRMD6-induced tumor-inhibiting effects. Thyroid cancer cells overexpressing FRMD6 had a weakened ability to form xenograft tumors in vivo in nude mice. Overall, the overexpression of FRMD6 produces remarkable tumor-inhibiting effects in thyroid cancer by inhibiting oncogenic YAP1.

FOCAD/miR-491-5p, downregulated by EGR1, function as tumor suppressor by inhibiting the proliferation and migration of gastric cancer cells.

Gastric cancer is a common malignant tumor in China; however, its carcinogenesis remains unknown. Focadhesin (FOCAD) is a tumor suppressor gene in gliomas, its expression, role, and mechanism in gastric cancer have not been defined. The aim of the present study was to explore the expression pattern of FOCAD in human normal tissues and cancer tissues and elucidate the role and regulatory mechanism of Early Growth Response 1 (EGR1) in FOCAD and its intron, miR-491-5p, in gastric cancer. Immuno histochemical staining revealed that FOCAD is widely and highly expressed in normal gastric mucosa, but is absent in gastric cancer tissue. Based on an association analysis FOCAD expression was found to be negatively associated with lymph node metastasis (P = 0.004); higher FOCAD levels were associated with longer survival in patients with gastric cancer (P = 0.001). MTT, colony, Transwell chamber, and flow cytometry assays revealed that siFOCAD promoted cell proliferation, growth, and migration, and inhibited apoptosis. Furthermore, bioinformatic analysis, Fluorescence reporter gene and chromatin immunoprecipitation analyses confirmed that EGR1 binds to the promoter and negatively regulates FOCAD and miR-491-5p at the transcriptional level. The overexpression of EGR1 was also found to promote cell proliferation, growth, and migration, and inhibit apoptosis. Overall, FOCAD is specifically overexpressed in the gastric mucosa and is significantly downregulated in gastric cancer. To our knowledge, this is the first study to demonstrate that FOCAD is a tumor suppressor, higher FOCAD levels might be a better prognostic marker of gastric cancer, and FOCAD/miR-491-5p may be negatively regulated by EGR1.

Molecular characterisation of the faecal microbiota in patients with type II diabetes.

The investigation provides molecular analyses of the faecal microbiota in type 2 diabetic patients. In order to characterise the gut microbiota in diabetic patients and to assess whether there are changes in the diversity and similarity of gut microbiota in diabetic patients when compared with healthy individuals, bacterial DNAs from 16 type 2 diabetic patients and 12 healthy individuals were extracted from faecal samples and characterised by PCR-denaturing gradient gel electrophoresis (DGGE) with primers specifically targeting V3 region of the 16S rRNA gene, as well as been sequenced for excised gel bands. The counts of Bacteroides vulgatus, Clostridium leptum subgroup and Bifidobacterium genus were assessed using quantitative PCR. By comparing species diversity profiles of two groups, we observed that there were no significant differences between diabetic and healthy group, although a few diabetic individuals (D6, D8) exhibited a remarkable decrease in species profiles. As for the similarity index, it was lower in inter-group than that in intra-group, which showed that the composition of gut microbiota in diabetic group might be changed due to diabetes status. Sequencing results also revealed that bacterial composition of diabetic group was different from that of the healthy group. B. vulgatus and Bifidobacterium genus were low represented in the microbiota of diabetic group, and the significant decrease was observed for Bifidobacterium by real-time PCR. Taken together, in this work we observed the characterisation of gut microbiota in diabetic patients, which suggests that the gut microbiota of diabetes patients have some changes associated with occurrence and development of diabetes.

[Effects of chronic triochloride aluminum exposure on both histocytological changes and APP695 mRNA level of rat spinal cords].

OBJECTIVE: To study the effects of chronic triochloride aluminum (AlCl3) exposure on both histocytological changes and amyloid precursor protein 695 (APP695) mRNA expression level of the rat spinal cord, in order to evaluate the relationship of the histocytological changes and the APP695 mRNA level in rat spinal cord exposed chronicly AlCl3. METHODS: Twenty six-month-old SD female rats were randomly divided into 2 groups, group normal and group exposed. The rats were exposed with intraperitoneal injection of 1 ml AlCl3 solution for continuous 3 days of the interval of one day during the period of total 72 days. Behaviors of the rats were observed by naked eyes. Histocytological examinations of the spinal cords of the rats were performed with HE dyeing. The expression levels of APP695 mRNA of spinal cords were tested with RT-PCR. RESULTS: In comparition with group normal, intraperitoneal injection of AlCl3 solution caused movement obstacle of rat hind legs for short time. In comparition with normal group in the spinal cord cinerea of exposure group, invasion of glial cells into motor neuron appeared, satellite phenomenon appeared, glial cells prominently increased in number, traverse neurofibers prominently increased both in number and marking, glial cells around the blood vessels prominently increased in number, vacuoles prominently increased in number, expressions of APP695 mRNA prominently increased. CONCLUSION: Intraperitoneal injection of AlCl3 solution could cause movement obstacle of rat hind legs for short time. Chronic aluminum exposure could cause injury or death of neurons, induced increases of glial cells, and vacuolar degeneration in rat spinal cords. Super expression of APP695 mRNA could participate the neuronal injury course.

FOXN3 suppresses the growth and invasion of papillary thyroid cancer through the inactivation of Wnt/ß-catenin pathway.

Forkhead box N3 (FOXN3) is a subtype of FOX family that has been demonstrated to be implicated in several cancers. However, the role of FOXN3 in papillary thyroid carcinoma (PTC) and its mechanisms have not yet been investigated. Our results showed that FOXN3 was markedly down regulated in PTC tissues and cell lines. Overexpression of FOXN3 suppressed the proliferation, colony formation, migration, and invasion in PTC cells. Overexpression of FOXN3 also prevented EMT process in PTC cells, as shown by the increased E-cadherin expression level and decreased expression levels of N-cadherin and vimentin. In addition, overexpression of FOXN3 inhibited tumor growth of PTC in vivo. Furthermore, overexpression of FOXN3 caused significant decreases in expression levels of ß-catenin, c-Myc, and cyclin D1. Additionally, activation of Wnt/ß-catenin pathway reversed the effects of FOXN3 on PTC cells. In conclusion, these findings indicated that FOXN3 exerted a tumor suppressive activity in PTC, which was mediated by Wnt/ß-catenin pathway.

[Antimicrobial resistance and penicillin resistance-associated genes of Streptococcus pneumoniae isolated from children with respiratory tract infection].

OBJECTIVE: To investigate the antimicrobial resistance and penicillin resistance-associated genes (TEM and pbp2B) of Streptococcus pneumoniae (S. pneumoniae) isolated from sputum specimens of Guangzhou children with respiratory tract infection. METHODS: E-test and Kirby-Bauer methods were applied to detect the antibiotic susceptibility of 44 strains of S. pneumoniae. PCR was used to detect resistance genes pbp2B and TEM, followed by DNA sequence analysis of pbp2B gene. The sequence results were compared to those of penicillin-susceptible S. pneumoniae R6. RESULTS: Of the 44 isolates of S. pneumoniae, only 5 (11.4%) were susceptible to penicillin. All strains were resistant to erythromycin but susceptible to ofloxacin and vancomycin. The resistance rate of the isolates to clindamycin and trimoxazole was more than 90%. The S. pneumoniae isolates showed a high susceptibility to amoxicillin, imipenem and ceftriaxone, with a resistance rate of 0, 2.6% and 3.9%, respectively. The sequence analysis showed that more than 99% nucleotide sequence of pbp2B gene of five penicillin-susceptible isolates was the same as penicillin-susceptible S. pneumoniae R6, without any amino acid replacement. Site mutation was found in the remaining 39 penicillin-nonsusceptible isolates with a nucleotide mutation rate ranging from 13.2% to 23.1% and amino acid replacement rate from 6.5% to 10.9%. The 39 penicillin-nonsusceptible isolates were classified into 4 types according to the mutation site between Ser391 and Thr492 of pbp2B: type I (n=30), type II (n=7), type III (n=1) and type IV (n=1). No TEM gene was detected in all the 44 S. pneumoniae isolates. CONCLUSIONS: The S.pneumoniae isolates from Guangzhou children with respiratory tract infection are resistant to penicillin and erythromycin. Amoxicillin and the third generation cephalosporin may be recommended for treating S. pneumoniae infection. The mutation of pbp2B gene plays an important role in the development of S. pneumoniae resistance to penicillin.

Antibiotic usage in Chinese children: a point prevalence survey.

BACKGROUND: Children as a population have high antimicrobial prescribing rates which may lead to high resistance of bacteria according to data from some single-center surveys of antibiotic prescribing rates in China. The acquirement of baseline data of antibiotic prescribing is the basis of developing intervention strategies on inappropriate antimicrobial prescriptions. Few studies show clearly the pattern and detailed information on classes of antibiotics and distribution of indications of antibiotic prescriptions in children in China. This study aims to assess the antibiotic prescribing patterns among children and neonates hospitalized in 18 hospitals in China. METHODS: A 24-hour point prevalence survey on antimicrobial prescribing was conducted in hospitalized neonates and children in China from December 1st, 2016 to February 28th, 2017. Information on the antibiotic use of patients under 18 years of age who were administered one or more on-going antibiotics in the selected wards over a 24-hour period was collected. These data were submitted to the GARPEC (Global Antimicrobial Resistance, Prescribing and Efficacy in Children and Neonates) web-based application ( https://pidrg-database.sgul.ac.uk/redcap/ ). For statistical analysis, Microsoft Excel 2007 and SPSS 22.0 were used. RESULTS: The antibiotic data were collected in 35 wards in 18 hospitals from 9 provinces. In total, 67.76% (975/1439) of the patients (n = 1439) were given at least one antibiotic, including 58.1% (173/298) of neonates (n = 298) and 70.3% (802/1141) of children (n = 1141). In neonates, the three most frequently prescribed antibiotics were third-generation cephalosporins (41.7%), penicillins plus enzyme inhibitor (23.8%), and carbapenems (11.2%). In children, the three most frequently prescribed antibiotics were third-generation cephalosporins (35.5%), macrolides (23.2%), and penicillins plus enzyme inhibitors (15.9%). The most common indication for antibiotics was proven or probable bacterial lower respiratory tract infection (30.9% in neonates and 66.6% in children). CONCLUSIONS: Antibiotics are commonly prescribed in the Chinese children population. It is likely that the third-generation cephalosporins and macrolides are currently overused in Chinese children. Efforts must be made to ensure safe and appropriate antibiotic prescribing to reduce and prevent the future development of antibiotic resistance.

[Molecular characteristics of methicillin resistant Staphylococcus aureus isolated from Chinese children].

OBJECTIVE: To investigate the molecular characteristics of methicillin-resistant Staphylococcus aureus (MRSA) isolates from Chinese children in seven cities. METHOD: A total of 134 MRSA isolates were collected from nine hospitals. Multilocus sequence typing and spa typing were analyzed by polymerase chain reaction (PCR), and staphylococcal chromosomal cassette mec (SCCmec) type was analyzed by multiplex PCR. The Panton-Valentine leukocidin (pvl) gene was also detected. RESULT: Most MRSA strains were isolated from pneumonia and skin and soft tissue infection (SSTIs) patients, accounting for 82.1%. Overall, 16 sequence types (STs) were obtained, and CC59 (51.7%) was found to be the most prevalent, which included ST 59 and ST 338, followed by ST239 (16.4%). SCCmec types II, III, IV, and V were also identified in the current study. SCCmec type IV was the most predominant type at 50.0%, followed by SCCmec type V at 23.9% and III at 23.9%. SCCmec subtypes IVa, IVc, and IVg were found among SCCmec type IV strains, whereas IVa was the main subtype at 77.6%. Twenty-six spa types were also identified, among which the predominant type was t437 (47.8%). The prevalence of pvl genes and the SCCmec type of strain was relevant, and the pvl gene positive rate was higher in SCCmec type IV and V-type strains than in SCCmec type II and III strains (58.6% vs. 14.3%, P < 0.05); there was a significant difference between them. In the strains isolated from pneumonia and SSTIs, ST59-MRSA-IVa(t437) was the predominant clone. There were five clones detected from the strains isolated from septicemia, with ST59-MRSA-IVa(t437) and ST59-MRSA-V(t437) as the main clones (57.1%). Various predominant clones existed in different regions. ST59-MRSA-IVa(t437) was the prevalent clone in the Guangzhou, Beijing, Chongqing, and Shenzhen areas, whereas ST239-MRSA-III(t037) was the prevalent clone in the Shanghai area. Fifty percent of the isolates from the Wenzhou area belonged to ST910-MRSA-V(t318), whereas three clinical strains isolated from the Shenyang region belonged to three different types. CONCLUSION: The results indicate that MRSA isolates from Chinese children are largely associated with the ST59-MRSA-IV(t437) and ST239-MRSA-III(t037) clones. These two may belong to community-acquired MRSA and hospital-acquired ones, respectively. Different prevalent clones were detected in different diseases and different regions. Therefore, there is a need to conduct further research on clinical isolates, which can guide the choice of antibiotic treatment and the examination of MRSA prevalence.

[Inhibition of respiratory syncytial virus replication in cultured cells by RNA-cleaving DNAzyme].

OBJECTIVE: DNAzyme/Deoxyribozyme is another novel molecular biological tool following the ribozyme. DNAzyme consists of a 15-nucleotide (nt) internal loop as its catalytic domain and two flanking substrate-recognition domains of 7 to 8 nt each which is complementary to substrate. The RNA substrate is cleaved at a particular phosphodiester located between an unpaired purine and a paired pyrimidine residue. DNAzyme has been applied in fields such as viral infectious disease, tumor, cardiovascular disease and genetic disease. But there is no report about using DNAzyme for anti-respiratory syncytial virus purpose. To observe the inhibitory effects of RNA-cleaving DNAzymes on respiratory syncytial virus (RSV) replication in cultured cells. METHODS: Anti-RSV RNA-cleaving DNAzyme DZ604 was designed to target the RSV genome at the start of the NS2 gene in an effort to inhibit the RNA replication. Microscope and electron microscope were used to observe the effects of anti-RSV genomic RNA DNAzyme on cytopathogenic effect (CPE) and ultrastructural change of 9HTE cell induced by RSV infection. Viral plaque forming reduction assay and MTT assay were used to detect the anti-RSV activity and protective function for RSV infected 9HTE cells of DNAzyme. RESULTS: Anti-RSV genomic RNA DNAzyme (DZ604) significantly improved CPE of RSV-infected 9HTE cells. The time to appearance of CPE and of total CPE was delayed by using DZ604 in a dose-dependent manner. At a 5 micro mol/L concentration of DZ604, CPE of 9HTE cells induced by RSV infection at 10 and 1 multiplicity of infection (MOI) was not improved. At smaller MOI (0.1, 0.01, 0.001, 0.0001) of RSV infection, CPE of 9HTE cells was significantly lightened by DZ604 at the same concentration. DZ604 also significantly improved ultrastructural change of 9HTE cells at early stage of RSV infection. Reduction in RSV yield was 85.56% and 8.33% at concentrations of 5 micro mol/L and 0.25 micro mol/L of DZ604. DZ604 inhibited RSV yield in a dose-dependent manner (P < 0.05). Non-specific DNAzyme did not have anti-RSV activity (P > 0.05). CONCLUSION: Anti-RSV genomic RNA DNAzyme designed and synthesized in our laboratory was capable of inhibiting respiratory syncytial virus replication specifically in cultured cells. Our data indicated that DNAzymes could be useful for the prevention against respiratory syncytial virus infection.